key: cord- -elfroq f authors: hakim, f. a.; tleyjeh, i. m. title: severe adenovirus pneumonia in immunocompetent adults: a case report and review of the literature date: - - journal: eur j clin microbiol infect dis doi: . /s - - -z sha: doc_id: cord_uid: elfroq f adenovirus is a frequent cause of mild self-limiting upper respiratory tract infection, gastroenteritis, and conjunctivitis in infants and young children. fatal infections (severe pneumonia progressing to respiratory failure, septic shock and/or encephalitis) are rare among immunocompetent adults. we report a case of severe adenovirus pneumonia in a young immunocompetent male who presented with sudden onset respiratory distress that progressed rapidly to respiratory failure and made a successful recovery on supportive measures. systematic review of the literature identified cases of severe adenovirus pneumonia (defined as respiratory failure requiring ventilatory support at any point during the course of illness) in otherwise healthy immunocompetent adults both in epidemic and community settings. we describe the clinical characteristics, radiological features, and outcome of identified cases. adenovirus is a frequent cause of mild self-limiting upper respiratory tract infection, gastroenteritis, and conjunctivitis in infants and young children. epidemics of self-limited infections by adenovirus (especially serotype ) have been reported from time to time in military recruits. severe adenovirus infections causing significant morbidity and mortality are well described in immunocompromised patients but are extremely rare in healthy immunocompetent adults [ ] . among the known serotypes of human adenoviruses, serotypes and account for most of these severe infections. such infections first gained attention during the epidemics of the s and early s in military trainees, when no vaccine was available [ , ] . occurrence of such infections among the military recruits dropped significantly with the introduction of a vaccine in . however, severe adenovirus infections among immunocompetent adults raised concern when ryan et al. reported related deaths in two military recruits in [ ] . sporadic cases of severe infection have been reported from the community as well [ , ] . we report a case of severe adenovirus pneumonia in a previously healthy immunocompetent male who presented to us with rapidly developing respiratory failure and made a successful recovery on supportive measures. we also summarize the clinical features, radiological findings, and outcome of severe adenovirus pneumonia cases that have been reported in the literature. immunodeficiency virus (hiv). there was no history of travel outside riyadh city in the recent past. he had no exposure to birds or contact with sick persons. he had no pets at home. past medical history and family were unremarkable. on his arrival to the er, the patient was febrile (temperature . °c), tachypneic (respiratory rate /min), and tachycardic (pulse rate /min). his blood pressure was / mmhg and oxygen saturation was % by pulse oximetry while breathing room air. there was mild pharyngeal and tonsillar erythema but no conjunctival injection, rash, or lymphadenopathy. there was no lower limb edema or jugular venous distension. cardiovascular examination was normal and auscultation of the chest revealed bilateral scattered rhonchi. the rest of the systemic examination was unremarkable. laboratory investigations at admission revealed polymorpholeukocytosis (white cell count . × /μl with . % neutrophils). hemoglobin ( . g/dl), platelets ( × /μl), sedimentation rate ( mm st h), and urinalysis were normal. chest x-ray was normal and arterial blood gases revealed hypoxemia and mild respiratory alkalosis. serum biochemical panel including random glucose, urea/electrolytes, liver function tests, and amylase were normal. serum cardiac enzymes and coagulation profile were normal. doppler ultrasound of leg veins was negative for venous thrombosis and ventilation perfusion scan revealed multiple bilateral ventilation perfusion defects. computed tomogram (ct) scan of the chest showed no parenchymal lung disease and spiral ct angiogram scan did not reveal pulmonary embolism. six hours after presentation, the patient developed worsening respiratory distress and was transferred immediately to the intensive care unit (icu) and mechanically ventilated. given the septic shock picture, empiric antimicrobial therapy was started. over the next h the patient's respiratory failure worsened and he developed fluidresistant shock. inotropes and vasopressors were used to maintain his mean arterial pressure between and mmhg. he continued to have low-grade fever, developed new conjunctival congestion, and had coarse crackles on chest auscultation. repeat portable chest x-ray showed bilateral fluffy and confluent air space shadowing in the mid and lower zones. ct scan of the chest was consistent with bilateral consolidation and bilateral pleural effusion. transesophageal echocardiography was normal. cultures drawn at admission were negative. legionella urine antigen, mycoplasma serology, and hiv serology were negative. gram staining, acid fast staining, mycobacterium tuberculosis dna probe, cytomegalovirus, and pneumocystis carinii performed on bronchoalveolar lavage were negative. repeat cultures and urine drug screen were negative. the patient developed nonoliguric acute renal failure. urinalysis was negative for active urine sediment and proteins and renal ultrasound was normal. immunologic tests including ana, anti-dsdna, c-anca, and p-anca and cryoglobulins were negative. interestingly, the patient developed bilateral conjunctival chemosis and hemorrhages with mild watery discharge on day of illness ( fig. ). tracheal and nasopharyngeal aspirates sent for immunofluorescent viral studies were reported as strongly positive for adenovirus (magen direct antibody fluorescence). adenovirus serology and serotyping were not performed. droplet precautions were observed to prevent infection of the caring staff. ophthalmological evaluation revealed hemorrhagic conjunctivitis. the systemic empirical antibiotics were withdrawn and supportive measures for respiratory and renal failure continued with continuous monitoring of vitals and serum biochemical panels. no antiviral therapy was used. inotropes and vasopressors were gradually withdrawn. the patient's respiratory failure and renal failure improved over the next days and he was successfully weaned off the ventilator and extubated. he was discharged home on day after admission. he was doing very well at the -month follow-up. we reviewed and analyzed the clinical features, radiological findings, and outcome of severe adenovirus pneumonia in immunocompetent patients reported between and by searching medline for english reports and using "severe," "adenovirus," "pneumonia," and "immunocompetent" as search terms. we defined severe adenovirus pneumonia if associated with respiratory failure requiring ventilatory support at any point during the course of illness and immunocompetent adults as individuals with no acquired or congenital immunodeficiency state with or without associated premorbid conditions. we identified cases. seven of these cases occurred in epidemics among fig. photograph of the patient showing bilateral hemorrhagic conjunctivitis military recruits and a mental health care center and the remaining seven were reported from the community. table summarizes age, sex, presenting complaints, physical findings, radiological findings, and outcome of each patient. adenovirus serotype (wherever performed), time to ventilation, and time to death from onset of illness are also included when available. the majority of the patients ( / , . %) were males and the mean age was . years (range: - years). most patients had no premorbid conditions. respiratory symptoms were present in all patients; three had associated diarrhea. radiograph of the chest revealed focal or diffuse infiltrate or consolidation. serotyping was done in six patients: three had serotype , three had serotype , and one had both serotypes and . mean time from onset of symptoms to admission to icu for respiratory failure was . days (range: - days). of the patients identified, ( . %) died due to worsening respiratory failure. three of the deceased patients had associated renal failure and disseminated intravascular coagulopathy (dic) and one had meningitis. an antiviral agent (cidofovir) was used in one patient. most of these patients developed hypotension during the course of their disease and six of these progressed to fluid-resistant shock. adenovirus is a frequent cause of mild upper respiratory tract illness. severe adenovirus infection is rare in immunocompetent adults. outbreaks of fulminant respiratory illness due to adenovirus have been reported in military camps in and [ , ] , in hospitalized patients in [ ] , in a mental health care center in [ ] , and among civilians in [ ] . several sporadic cases of adenovirus infections in healthy immunocompetent individuals with case fatalities have also been reported [ , ] . our review suggests that most patients have upper respiratory symptoms prior to the onset of respiratory distress and progression to respiratory failure is unpredictable, occurring over hours to days. the presence of flu-like symptoms, diarrhea, and conjunctival infection may be a clue to the viral etiology of the illness. hemorrhagic conjunctivitis was present in only one of the identified patients; when present it is strongly associated with adenovirus infection. however, it may not be apparent early in the course of the disease as is illustrated in our case. the majority of such patients are febrile and tachycardic at presentation. chest auscultation may be normal or may reveal localized or bilateral rales, rhonchi, or signs of consolidation. fluid responsive hypotension and a transient drop in leukocyte count (relative leukopenia and lymphopenia) are common during the course of illness. initial chest x-ray may be normal but reveal focal reticulonodular infiltrate or a lobar consolidation with the progression of the infection. lobar consolidation is rare in other types of viral pneumonia [ ] [ ] [ ] [ ] . the majority of patients with severe adenovirus infection died due to respiratory failure and shock. disseminated intravascular coagulopathy, renal failure, and fluid-resistant shock are also common causes of death and should be always looked for in such patients. meningitis is a rare but fatal complication of severe adenoviral pneumonia. the whole clinical scenario may be mistaken for bacterial sepsis. whenever considered, the diagnosis of adenovirus infection is established by isolating the virus on cultures or detecting viral particles on fluorescence antibody testing from material obtained from nasopharyngeal and tracheal aspirate, bronchoalveolar lavage, or lung biopsy. a fourfold or more rise in adenovirus-specific antibody titers during the course of illness suggest acute infection and alternatively establishes the diagnosis. treatment of severe adenovirus infection is supportive and no antiviral agent has been shown to alter the unpredictable course of the disease. cidofovir and intravenous immunoglobulin have been used with some success in patients with liver and bone marrow transplant [ ] . in our review, it was used in only one patient who did not survive. it appears that severe adenovirus infections even in immunocompetent patients are associated with high mortality and morbidity. the spectrum of presentation and characteristics of severe adenovirus infections could be affected by publication bias. adenovirus produces cytolysis which accounts for tissue damage and subsequent dysfunction of the affected organs. it also induces interleukin (il- ) production, which causes a severe systemic inflammatory response causing systemic vasodilatation and triggers dic. damage to pulmonary capillaries also contributes to respiratory failure. cellular immunity is important in viral infection in general. hence, severe adenovirus infections are common in patients with defective or deficient t-cell immunity such as those with allogenic stem cell transplantation, solid organ transplants, and hiv infection. infants and young children also are at risk of severe adenovirus infections due to immature t-cell function. why severe adenovirus infection should occur in immunocompetent individuals is not well understood at this point in time. a unique interplay between viral factors (virulence) and host factors (genes) have been well described in adenovirus infections. virulent strains of adenovirus (serotypes and , etc.) in a genetically susceptible host impair cytokine, t-cell function, and expression of major histocompatibility complex expression [ , ] . however, the relationship between virulence and genotype needs to be explored. to conclude, the clinical scenario of severe adenovirus pneumonia resembles severe bacterial pneumonia and the two may be indistinguishable on clinical and radiological grounds. diagnosis of adenovirus infection should be always considered in patients with severe pneumonia with negative cultures and failure to respond to antibiotics. since this can rapidly progress to respiratory failure, the diagnosis should be looked for early in the course to avoid unnecessary drugs and to apply aggressive supportive measures in the icu setting which might improve outcome in such patients. more work needs to be done to identify viral determinants and genetic factors triggering severe adenovirus infection in immunocompetent individuals and to better understand the pathophysiology and immunology of such infections. we also need to identify subsets of patients with severe adenovirus who could benefit from antiviral drugs. adenovirus infections in immunocompromised patients association of type adenovirus with acute respiratory illness in military recruits fatal pneumonia associated with adenovirus type in three military trainees two fatal cases of adenovirus-related illness in previously healthy young adults--illinois fatal type adenoviral pneumonia in immunocompetent adult identical twins severe adenovirus infection in an immunocompetent adult-a case report type adenoviral pneumonia occurring in a respiratory intensive care unit multiple cases of life-threatening adenovirus pneumonia in a mental health care center severe adenovirus pneumonia in an adult civilian successful treatment of adenovirus disease with intravenous cidofir in an unrelated stem cell transplant recipient molecular basis of immune evasion strategies by adenovirus adenovirus-pulsed dendritic cells stimulate human virus-specific t-cell responses in vitro key: cord- -woref g authors: fragoso-saavedra, sergio; iruegas-nunez, david a.; quintero-villegas, alejandro; garcía-gonzález, h. benjamín; nuñez, isaac; carbajal-morelos, sergio l.; audelo-cruz, belem m.; arias-martínez, sarahi; caro-vega, yanink; calva, juan josé; luqueño-martínez, verónica; gonzález-duarte, alejandra; crabtree-ramírez, brenda; crispín, josé c.; sierra-madero, juan; belaunzarán-zamudio, pablo f.; valdés-ferrer, sergio i. title: a parallel-group, multicenter randomized, double-blinded, placebo-controlled, phase / , clinical trial to test the efficacy of pyridostigmine bromide at low doses to reduce mortality or invasive mechanical ventilation in adults with severe sars-cov- infection: the pyridostigmine in severe covid- (pisco) trial protocol date: - - journal: bmc infect dis doi: . /s - - - sha: doc_id: cord_uid: woref g background: severe acute respiratory syndrome coronavirus (sars-cov- ) infection, the causative agent of coronavirus disease (covid- ), may lead to severe systemic inflammatory response, pulmonary damage, and even acute respiratory distress syndrome (ards). this in turn may result in respiratory failure and in death. experimentally, acetylcholine (ach) modulates the acute inflammatory response, a neuro-immune mechanism known as the inflammatory reflex. recent clinical evidence suggest that electrical and chemical stimulation of the inflammatory reflex may reduce the burden of inflammation in chronic inflammatory diseases. pyridostigmine (pdg), an ach-esterase inhibitor (i-ach-e), increases the half-life of endogenous ach, therefore mimicking the inflammatory reflex. this clinical trial is aimed at evaluating if add-on of pdg leads to a decrease of invasive mechanical ventilation and death among patients with severe covid- . methods: a parallel-group, multicenter, randomized, double-blinded, placebo-controlled, phase / clinical trial to test the efficacy of pyridostigmine bromide mg/day p.o. to reduce the need for invasive mechanical ventilation and mortality in hospitalized patients with severe covid- . discussion: this study will provide preliminary evidence of whether or not -by decreasing systemic inflammation- add-on pdg can improve clinical outcomes in patients with severe covid- . trial registration: clinicaltrials.gov nct (registered on april , ). severe acute respiratory syndrome coronavirus (sars-cov- ) infection, the causative agent covid- , may result in severe systemic inflammatory response. about one third of hospitalized patients with covid- develop acute respiratory distress syndrome (ards) [ ] , while % require invasive mechanical ventilation associated to a high mortality rate [ ] . the two main causes of death in patients with severe covid- are respiratory and multiple-organ failure as a result of overwhelming inflammatory response [ , ] . therefore, patients with severe covid- will theoretically benefit from therapeutic interventions that modulate the inflammatory response [ ] . pyridostigmine, an acetylcholinesterase inhibitor (i-ach-e), increases acetylcholine (ach) half-life by inhibiting its peripheral degradation. pyridostigmine has been used for decades in the symptomatic treatment of myasthenia gravis [ ] and as pre-exposure prophylaxis against nerve gas (chemical) warfare [ ] . pyridostigmine has well-characterized pharmacokinetic and safety profiles. recently, pyridostigmine has been shown to reduce persistent inflammation in people living with hiv- infection [ ] [ ] [ ] . the proposed anti-inflammatory effect occurs after the ach binds to nicotinic receptors on the surface of immune cells and this interaction causes a decrease in the production of pro-inflammatory cytokines. this so-called inflammatory reflex, originally described in response to vagus nerve stimulation [ ] , leads to the release of ach with a resulting reduction in acute [ ] and chronic inflammation [ ] . our primary objective is to evaluate whether or not add-on pyridostigmine to best medical management of hospitalized covid- patients will result in reduced need for invasive mechanical ventilation and death. our aim is to tests the efficacy of pyridostigmine use as an immunomodulator to reduce the incidence of complications leading to critical illness or death in hospitalized adults with severe covid- . in order to test this, we propose a randomized, double-blinded, placebocontrolled trial. participants will be randomly allocated in a : ratio to receive either oral pyridostigmine at a dose of mg/day or a matching placebo for a maximum of days in parallel groups. we will compare the need of invasive mechanical ventilation and fatality rates during the days following randomization (fig. ). unblinding will be permissible in case of severe adverse events at the request of the treating group of physicians, or at the request of the external data and safety monitoring board (dsmb). the study is planned in two parts: a phase aimed at determining security, followed by a phase part aimed at evaluating the effect -or lack thereof-of pyridostigmine in patients with severe covid- . phase started recruiting on may . during the security (phase ) part, we aimed at evaluating the safety and feasibility of the study and explore in a preliminary way the magnitude of the effect of the intervention. safety was evaluated according to the frequency of outcomes as well as of reported adverse events. enrollment for the security phase was limited to patients hospitalized at instituto nacional de ciencias médicas y nutrición salvador zubirán (incmnsz) in mexico city. on july , a pre-appointed dsmb performed an ad interim analysis after the first participants ( % of the calculated sample) had been recruited and, as results derived from this security part indicated that pyridostigmine was not associated with an increased frequency of outcomes or adverse events (safety outcome), the dsmb recommended to proceed to a multi-center, phase trial. during this ongoing phase component of the rct, the primary outcome to be evaluated will be a composite outcome including ) the requirement of invasive mechanical ventilation, ) an increase in the sofa scale ≥ points, or ) death. the dsmb also suggested to repeat an ad interim analysis every time a %-recruiting milestone is reached. we are including adult (≥ -year-old), hospitalized patients with confirmed sars-cov- infection based on a positive rt-pcr test for sars-cov- rna in a respiratory specimen (nasopharyngeal or nasal swab) and an imaging study compatible with pneumonia, and at least one high-risk criteria of death (see table ). exclusion criteria include one or more of the following: allergy to pyridostigmine; pregnancy or breastfeeding status; concomitant autoimmune disease; diagnosed immunodeficiencies (including hiv infection); need for mechanical ventilation, admission to the icu, or meeting criteria for septic shock before providing signed, informed consent; inability to receive orally or enterally administered drugs; use of immunosuppressants or immune-modulators (including chemotherapy and corticosteroids) in the preceding -day period unless recommended by the treatment medical team as part of the therapeutic approach for sars-cov- infection; and participation in clinical trials of any kind in the previous days. participants will be randomized in a : ratio, with parallel assignment to receive either placebo or pyridostigmine as an add-on medical treatment to the best medical care available for severe covid- in participating centers. the block-randomization process will be performed using the publicly available online resource (www.randomizer.org). patients in the treatment group will receive pyridostigmine bromide, mg/day per os. participants randomized to the control group will receive matching placebo (identical in appearance) made of pharmaceutical grade starch. participants will be receiving the assigned intervention until the occurrence of either ) any of the prespecified outcomes; ) hospital discharge; or ) a maximum period of in-hospital days (fig. ) . the primary outcomes are a composite of requirement of invasive mechanical ventilation, an increase of ≥ points in the sofa scale, or all-cause mortality, during the -day period following enrollment; and, ) safety of the study drug. the secondary outcome is the change in interleukin (il)- levels (Δ il- ) between baseline samples and those taken on days , , , and (for an outline of the protocol, please refer to fig. ; for the timeline of interventions and measurements, please refer to table ). we will collect demographic information from participants at baseline, including age, sex assigned at birth, presence of comorbidities which will include diabetes mellitus, systemic arterial hypertension, obesity, cardiovascular disease, and lung disease, and other chronic medical conditions from the clinical charts. safety of the intervention will be actively evaluated by daily interrogation of the following common adverse effects of pyridostigmine [ ] : abdominal pain or cramps; diarrhea; nausea, vomiting, or both; hypersalivation/drooling; urinary incontinence; muscle weakness or fasciculations; and, blurred vision. on day , patients will be contacted by telephone to assess their vital and functional status ( fig. ; table ). all collected data will be safeguarded on a coded database with access limited to project investigators. only the principal investigators and the dsmb will have access to the final trial dataset. the final results will be published for generalized access, regardless of the outcome. • abg x • il- measurement x x x x x • mass cytometry x x x x x follow-up phone call x a blood sampling will be performed only while participants are hospitalized. protocol does not require participants to return for further blood sampling after hospital discharge. abbreviations: cbc complete blood count, abg arterial blood gases, il- interleukin currently, recruiting for this study is undergoing at, instituto nacional de ciencias médicas y nutrición salvador zubirán (incmnsz), and instituto nacional de cardiología ignacio chávez, two covid- -designated hospitals in mexico city, mexico. we estimate that a sample size of participants ( in each group) would produce a one-sided % confidence limit that would exclude us finding a %-point difference that would be statistically significant in the complete trial [ ] . however, calculating a % loss, we will recruit participants for this part of the study. we estimate that a sample size of participants ( per group) estimating a % reduction in the occurrence of the primary outcome in the intervention group to be clinically significant. based on recent evidence from china, we estimate that % of patients hospitalized with severe sars-cov- infection will develop complications leading to the need of invasive mechanical ventilation or death [ ] accordingly, we estimate that this sample size will allow us to identify with an % power a reduction in the need of invasive mechanical ventilation or death of % in the group receiving pyridostigmine in comparison with the group on placebo, using a two-sided ttest at the . significance level. primary analysis will be performed by intention-to-treat analysis comparing the proportion of outcome events between groups using x test. we will also compare point estimates and its corresponding confidence intervals between groups. in a secondary analysis, we will use multivariate logistic regression models to explore variables associated with the primary outcomes. this is an ongoing study. recruiting started on may ; at submission, we have recruited participants. no results have been made available, and the therapeutic arms remain double blinded. therefore, no results have been submitted for publication or published. here, we propose to evaluate the potential usefulness of pyridostigmine as add-on therapy to best medical care of patients admitted to a hospital due to severe covid- . recent evidence indicates that between and % of patients hospitalized for covid- required care in intensive care units (icu) for severe hypoxemia. the reported mortality in those first cases that required hospital management is %, but in those with severe disease, the reported mortality is between and % [ , ] , and we assume that it will be similar elsewhere. severity and mortality of covid- appear to be mediated not by infection, but by the disproportionate inflammatory response of the host. hence, finding novel immunomodulatory strategies is a promising strategy to reduce severity and mortality of covid- . furthermore, the repurposing of drugs with well characterized safety profiles and readily available production lines, might lead to faster development of anti-covid- therapies if proven efficacious in well-designed, randomized clinical trials. in mammals, the central nervous system has mechanisms to control the inflammatory response. during inflammatory states, the vagus nerve can inhibit the synthesis and release of inflammatory cytokines [ ] , thereby reducing both local damage and mortality secondary to severe systemic inflammation in murine models as diverse as sepsis, ischemia and re-perfusion damage, or obesity [ ] [ ] [ ] [ ] . the vagus nerve can be stimulated electrically and chemically. chemical stimulation using cholinergic agonists has shown promising effects in murine and cellular models of inflammation [ , ] . acetylcholine esterase inhibitors (i-ach-e) are a family of drugs used regularly by millions of patients, including older adults with alzheimer disease and other dementias, as well as in patients with myasthenia gravis and dysautonomia [ , [ ] [ ] [ ] [ ] . these drugs inhibit the enzymatic degradation of endogenous ach, resulting in greater bioavailability and, therefore, an increase in the possibility of binding to both nicotinic and muscarinic receptors. in addition to the approved uses of i-ach-e in human pathology, there is evidence in various murine models of their efficacy in experimental sepsis and severe inflammatory response [ , , ] , suggesting that i-ach-e drugs have a potential immunomodulatory effect in patients with severe systemic inflammatory response syndrome. pyridostigmine, an acetylcholinesterase inhibitor, has been previously shown to decrease inflammation in people living with human immunodeficiency virus (hiv) infection [ ] [ ] [ ] ; therefore, it is possible that pyridostigmine may lead to a decrease in the production of pro-inflammatory cytokines in patients with covid- at high risk of severe disease. regarding safety concerns, at the proposed dose of pyridostigmine, the rate of adverse events is less than - % with no reported serious adverse effects [ ] . from this perspective, we consider that pyridostigmine can function as an immunomodulator and reduce morbidity and mortality in these patients. the reduction in the frequency of the need for mechanical ventilation would contribute to reducing mortality and the demand for these services. clinical features of patients infected with novel coronavirus in wuhan clinical course and risk 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dementia pyridostigmine treatment trial in neurogenic orthostatic hypotension publisher's note springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations not applicable. this study is funded by peer-reviewed, competitive grants from consejo nacional de ciencia y tecnología (conacyt): grants and , both to sivf. the final datasets will be made publicly available in the final manuscripts, supplemental materials, or public repositories. all protocol documents are available in spanish upon reasonable requests. this study is being carried out in accordance with the recommendations of the institutional ethics in human research committees. all participants will have given written informed consent to one of the study investigators in accordance with the declaration of helsinki. the protocol was approved by the ethics in human research committees of instituto nacional de ciencias médicas y nutrición salvador zubirán (incm nsz), and instituto nacional de cardiología ignacio chávez, both in mexico city, mexico; and comisión federal para la protección contra riesgos sanitarios (cofepris), the federal mexican pharmacological regulatory commission. not applicable. pablo f belaunzarán-zamudio is an associated editor for the hiv and coinfections section of bmc infectious diseases. key: cord- -obupqcua authors: chierakul, wirongrong title: leptospirosis date: - - journal: manson's tropical infectious diseases doi: . /b - - - - . - sha: doc_id: cord_uid: obupqcua nan leptospirosis is a worldwide zoonotic disease caused by pathogenic leptospira species. the disease presents in both tropical and temperate zones. the major reservoirs of the organisms are cattle, horses, canines and rodents. humans are accidentally infected through contact with contaminated water. the symptoms range from mild or asymptomatic to severe fatal illness. the severe illness, characterized by febrile illness with jaundice, acute renal injury and bleeding, is recognized as weil's disease, though many different local names have been used such as fort bragg, mud, swamp and sugar cane fevers. specific treatment with antibiotics is valuable at all stages of the illness and prevents development of severe disease. in severely ill patients, intensive care supportive treatment is crucial. molecular biology studies have led to major advances in our understanding of leptospira spp. during the past decade. leptospira are spiral bacteria (spirochaete) in the family of leptospiraceae. there are eight pathogenic, seven non-pathogenic leptospira species, and five intermediate species with unknown ability for causing disease. leptospira interrogans and l. borgpetersenii are the two most common species causing diseases in human and animals. the schematic classification of the organisms is shown in figure . . the old phenotypic classification of leptospira, based on serology using the cross-agglutination absorption test (caat) is still in use. approximately pathogenic serovars, grouped by related antigenicity into serogroups, have been identified to date. the concept of a serovar is complicated and may fail to define epidemiologically important strains. serovars of the same serogroup may distribute between different species identified by dna-dna hybridization (table . ). , the current recommendation for leptospira nomenclature is using species name followed by the term 'serovar' and serovar name with initial capital letter and non-italic, e.g. leptospira interrogans serovar autumnalis. the whole genomes of two pathogenic (l. interrogans and l. borgpetersenii) and a non-pathogenic species (l. biflexa) have been sequenced. l. interrogans has - % gc content in two circular chromosomes of approximately mb and kb in size. l. borgpetersenii has a smaller genome ( . mb) and a larger proportion of pseudogenes ( %), compared with l. interrogans. this may impair the ability of l. borgpetersenii to survive in the external environment, and so require direct contact between hosts to maintain the transmission cycle. , recent developments in molecular typing have characterized relationships between pathogenic strains and assisted outbreak investigation and epidemiology. several typing techniques have been developed. one is the sequence-based approach, multilocus sequence typing (mlst). using mlst the major outbreak of leptospirosis in thailand during - was shown to have been caused by one successive clone, strain type (st) of l. interrogans serovar autumnalis. (mlst has been used to create a standard global database for typing and mapping strains of leptospira, see: http://leptospira.mlst.net). a wide range of mammalian species, including rodents, cattle, pigs, domestic and wild animals, are the major reservoirs and carriers, whereas humans are mostly accidental and dead end hosts. the infecting organism is sustained naturally by chronic infection of the renal tubules of maintenance hosts after primary • leptospirosis is caused by eight pathogenic leptospira species, which have many mammalian animals as reservoirs. humans are accidental and dead-end hosts, who contact directly or indirectly with leptospiracontaminated water or animal products. • many infections in endemic areas are asymptomatic or oligosymptomatic. • clinical manifestations are those of a nonspecific acute febrile illness. complications such as cholestatic jaundice, aseptic meningitis, acute renal injury, haemorrhage especially in the lung and myocarditis can occur and lead to a fatal outcome. overall mortality is less than %. • clinical diagnosis is important but nonspecific. laboratory diagnosis is not practical for patient care, but very important for epidemiology, since both culture and serology take a relatively long time. • differential diagnoses are dengue and other haemorrhagic fevers, malaria, scrub typhus, hepatitis, yellow fever, hantavirus (both hps and hfrs), enteric fever and other bacterial sepsis, especially in patients with severe complications. • antibiotic treatment should be given as early as possible. doxycycline is the drug of choice in uncomplicated cases; penicillin, doxycycline, ceftriaxone and cefotaxime are efficacious alternatives in severe cases. • no effective human vaccine available, protection from contact is crucial. weekly doxycycline chemoprophylaxis in very high-risk groups is also effective. to assess the global burden of human leptospirosis, and thereby hopefully, will inform rational deployment of effective control and preventive measures. according to the second report of lerg in , the median global incidence of human leptospirosis, excluding outbreak cases, was five cases per population. ) and . ( . - . ) cases/ population, respectively. there were no data available from the eastern mediterranean in this report. leptospira serovars show specific, but not exclusive, host preferences. for example, l. borgpetersenii serovar hardjo predominates in farm animals especially cattle, l. interrogans serovar canicola circulates mainly in dogs. the incidence of leptospirosis is seasonal in several countries, mostly related to rainfall. after skin or mucosal penetration, the organisms cross the tissue barriers through intercellular junctions. leptospires can be detected in the bloodstream shortly after penetration into the body, and from some organs within three days after infection. leptospires are rarely observed within host cells. however, they can reside transiently within cells while crossing cell barriers. the organisms can evade host immune response during the initial phase of infection through unclear mechanisms. they are resistant to the alternative pathway of complement activation and acquire complement factor h and related fluid-phase regulators through ligands such as leptospiral endostatin-like a (lena) proteins. recent data show that pathogenic leptospires can bind human plasminogen (plg), and plasmin activity on the bacterial surface interferes with complement c b and igg deposition, therefore decreasing opsonization of the organisms. lipopolysaccharide (lps) of pathogenic leptospires can activate toll-like receptor (tlr ) but not tlr pathway, and activate the production of pro-inflammatory cytokines mediating inflammation and damaging end-organ tissues. comparison of putative pathogenicity factors with the saprophyte, l. biflexa has identified several pathogenic processes. these can be divided into three components; adhesins (proteoglycans, lena, kda outer surface membrane protein, and kda laminin binding protein (lsa , lsa ), leptospiral immunoglobulin-like proteins (lig a, lig b and lig c proteins); evasion of natural defences or phagocytosis; and resistance to complement as described above. unlike other gram-negative bacteria, the lipopolysaccharides (lps) of leptospires cause minor endotoxicity, while leptospiral outer membrane lipoproteins (omps), such as lip , loa play a major role as virulence factors. although the early phase of the disease does not cause much inflammation, hepatocellular and tubular damage can occur. liver pathology ranges from no appreciable changes, unicellular damage with oedema, to spotty necrosis surrounded by mononuclear cells without hepatocyte ballooning or swelling. complete liver cell necrosis is not unusual. kupffer cells and sinusoidal lining cells may show swollen cytoplasm. biliary stasis with bile droplets in hepatocytes and bile thrombi in the canaliculi is prominent in the centrolobular areas, with infection. the organisms are usually transferred from animal to animal by direct contact. the maintenance hosts or carriers can excrete leptospira in their urine for long periods of time or for their entire lives. leptospires survive weeks or months in moist and warm soil, stagnant water at neutral or slightly alkaline ph. humans are infected via direct contact with infected animal urine, animal abortion products or most commonly through indirect contact with infected urine-contaminated water. the oral route of transmission had been reported in water-borne fatal outbreaks in portugal, greece, russia, italy and india,. , , breast milk transmission may occur. sexual and vertical transmission in humans occurs rarely. humans at risk for leptospirosis are those with occupational exposure. these include farmers, fishermen, miners, animal slaughterers, veterinarians, sewage and canal workers, sugar cane workers, soldiers, etc. special events and activities clearly related to the diseases are recreational water sports, including triathlon, canoeing and white-water rafting, and natural disasters, such as hurricanes, floods, etc. laboratory-acquired infection may occur when dealing with high concentrations of organisms in culture. the true global burden is largely unknown both in human and animals. leptospirosis occurs in tropical, subtropical and temperate regions. the disease is often under-recognized and neglected because of the difficulties confirming the diagnosis. furthermore many infections in endemic areas are asymptomatic or oligosymptomatic. the six highest incidences reported are from the indian ocean and caribbean sea islands (seychelles, trinidad and tobago, barbados, jamaica, costa rica and sri lanka). countries in south-east asia, except thailand and singapore, have no official incidence data, so the true incidence is unknown. the leptospirosis burden epidemiology reference group (lerg) set up by world health organization (who) is trying renal pathology is mainly tubulointerstitial nephritis. in severe cases, tubular necrosis and medullary tubular cell desquamation can occur. glomeruli contain exudates and inflammatory cell infiltration, but are usually spared. primary injury of the proximal convoluted tubules is regarded as the hallmark of renal pathology, especially affecting sodium and water transportation. recently developed in situ hybridization assays and immunohistochemistry suggest that cell membrane damage in both the liver and kidney may be important in pathogenesis. vasculitis is a prominent feature of leptospirosis. endothelial damage and increased capillary fragility lead to internal organ haemorrhage. disseminated intravascular coagulation (dic) is not obvious in the pathology in fatal cases. however, with new definitions and sensitive markers for detection of dic, patients with leptospirosis do have significantly elevated fibrinogen, d-dimer, thrombin-antithrombin iii complexes, and prothrombin fragment , . using dic scores, defined by the dic scientific subcommittee of the international society of thrombosis and haemostasis, overt dic was evident in nearly half of patients. fatal bleeding usually occurs in the lung, with fibrin aggregation suggestive of adult respiratory distress syndromes (ards) in nearly % of fatal cases. capillary lesions characterized by swollen endothelial cells with an increase in pinocytotic vesicles and haemorrhagic pneumopathy with septal capillary lesions were present. the average incubation period is - days, with a range of - days. the clinical spectrum is extremely broad; ranging from subclinical or asymptomatic, mild insignificant, selflimited symptoms, to severe fatal manifestations. a substantial proportion of people in endemic areas infected with leptospirosis may have subclinical disease evidenced only by serology. leptospirosis is an important cause of acute undifferentiated fever. in the seychelles, % and % of screened people had past and recent infections respectively, though none of them reported any recognizable symptoms of leptospirosis. the reasons for protean manifestations are unclear. there were some reports of serovar-specific presentations and serovarrelated severity, such as serovar icterohaemorrhagiae and hepatorenal syndrome (pulmonary haemorrhage and renal failure), serovar copenhageni and uveitis, serovar bataviae with neurological preference. the classical manifestation of leptospirosis has been described as 'biphasic' pattern, an early nonspecific leptospiraemic phase lasts for a week followed by an immune phase with complications during the second week of illness. this biphasic course of leptospirosis dictates the selection of specimens for laboratory diagnosis, since it describes the phase of leptospires and the immunologic response in human hosts. the fever too may be biphasic. complications such as jaundice, acute renal injury, haemorrhage, especially pulmonary haemorrhage, aseptic meningitis, myocarditis, shock, occur early during the course of illness. direct invasion of leptospires and the hostresponse cause organ damage although specific immune responses may not be measurable until the second week of illness. certain distinctive complications, such as uveitis, usually occur as a late complication months later, sometimes without detectable clinical symptoms during the acute infection. the majority of patients are mild, self-limited 'flu-like' symptoms and may not seek medical attention. a small proportion of patients present with sudden onset of febrile illness and non-specific symptoms, such as headache, myalgia, backache, abdominal pain, conjunctival suffusion, chills, diarrhoea, anorexia, transient rash, cough, sore throat, etc. myalgia can be mild or severe accompanied by muscle tenderness and elevated muscle enzyme (creatinine phosphokinase; cpk) levels in serum. hepatomegaly may develop, but splenomegaly is unusual. severe leptospirosis is a multi-system disease. nearly % of patients develop the following complications during the course of illness. shock is a common presenting sign of leptospirosis, and may occur in % of patients. this results from several factors, including hypovolaemia from low fluid intake compounded by increased capillary permeability and vasodilatation from high fever, microvascular dysregulation, inflammatory responses to the high levels of bacteraemia, and low cardiac output from myocarditis and dysrhythmias. the majority of shocked patients will response to fluid replacement and low-dose inotropic agents, but in severe myocarditis and patients with sepsis-like syndrome, the prognosis of profound shock is poor. jaundice is a notable feature of severe leptospirosis. the proportion of patients with jaundice was reported as ranging widely from less than % up to more than %. extreme elevations of bilirubin may occur. jaundice in leptospirosis is associated with cholestasis rather than hepatocellular damage, reflected by a characteristic dissociation between the slight elevations in transaminase and alkaline phosphatase concentrations despite the exceedingly high bilirubin levels. the prothrombin time may be slightly prolonged. severe hepatic necrosis and fatal liver failure does occur rarely. jaundice usually appears on days - (up to days) after the onset of illness, and is not associated with pruritus. haemolysis may also contribute to the degree of jaundice. liver enlargement can be prominent in severe cases. renal involvement is an important manifestation of severe leptospirosis. in mild cases, the only abnormal findings are in the urinary sediment. these include albuminuria, microscopic haematuria, pyuria and the presence of granular casts in freshly examined urine. renal impairment may be attenuated by dehydration from low fluid intake and high fever. therefore, careful supportive treatment is very important. presenting symptoms can be non-oliguric, oliguric or anuric in severe cases. renal insufficiency commonly occurs together with jaundice, and is usually evident within the first - days of illness, followed by a rapid rise of plasma urea and creatinine often requiring renal replacement therapy. hyperkalaemia from acute renal injury can occur, but hypokalaemia due to impairment of sodium transporters in the proximal tubules and spared function of the distal tubules is more common. the polyuric phase may develop after - days, and the serum creatinine begins to fall at the end of the second week and normalizes within - weeks after the onset of illness as in other causes of acute tubulointerstitial nephritis. renal injury from leptospirosis is never permanent. pulmonary involvement occurs in - % of cases. this is mostly mild and often overlooked, but can be serious, leading to death. the severity of the pulmonary involvement is unrelated to liver dysfunction. the most common symptom is cough, with or without secretions. blood-tinged sputum or obvious haemoptysis may occur. abnormal radiological findings are found in more than half of patients despite the absence of respiratory symptoms. many different abnormalities can be found, including localized lesions such as lobar, confluent, or patchy infiltrations, or diffuse reticulonodular or interstitial infiltration. pulmonary haemorrhage usually presents with nodular or patchy infiltration, and sometimes with localized confluent consolidation, isolated interstitial infiltration is uncommon. pulmonary oedema with cardiomegaly due to volume overload or congestive heart failure from myocarditis and diffuse ground glass appearance without cardiomegaly reflecting ards are both observed and sometimes are difficult to distinguish. this fatal complication can occur as early as - days after onset of fever. pulmonary haemorrhage can be minimal or severe diffused leading to respiratory failure and is the leading cause of fatal outcome. cardiac involvement in leptospirosis is relatively common but often goes unnoticed. the most common finding is slight pr interval prolongation on the electrocardiogram (ecg), although other more severe conduction abnormalities may occur. atrial fibrillation is the most common dysrhythmia. fatal arrhythmias such as ventricular tachycardia seldom occur. these ecg changes and arrhythmias usually resolve after treatment. myocarditis, pericarditis, aortitis, arteritis of coronary and cerebral arteries and vasculitis have been reported, and may contribute to an unfavourable outcome. conjunctival suffusion is prominent in patients with leptospirosis, and subconjunctival haemorrhage is a common sign. subconjunctival haemorrhage is found more frequently compared with patients with scrub typhus (which commonly coexists in endemic areas of east asia), whereas red eyes happen equally in both diseases. more serious complications to the eyes are vitreous oedema or haemorrhages, and retinal haemorrhages which occur as early complications, or uveitis and anterior iridocyclitis as late complications, which may lead to permanent disturbance of vision. several forms of central nervous system involvement have been reported. the most common neurological complication is aseptic lymphocytic meningitis which occurs in - % of patients. patients usually present with severe headache, photophobia and nuchal rigidity accompanying the onset of fever. leptospira can be isolated from cerebrospinal fluid (csf) within days after onset of fever. the usual findings are a raised csf opening pressure, raised protein with normal csf glucose level and lymphocytic pleocytosis. the csf is culture negative for fungi or other aerobic bacteria. thus the main differential diagnosis is viral meningitis. seizure is rare and usually occurs late after the onset of other complications. other uncommon reported neurological manifestations include encephalomyelitis, polyneuropathies, guillain-barré syndrome, mononeuritis multiplex, cranial or peripheral nerve palsies. patients may present with a psychiatric syndrome characterized by mania, or may have mood instability as a rare complication which may persist for years. besides pulmonary haemorrhage, abnormal bleeding may be observed in other areas, such as gastrointestinal bleeding, epistaxis, gum bleeding, vaginal bleeding, and skin bleeding presenting as petechiae or ecchymoses. bleeding in vital organs with fatal sequelae have been reported in the adrenal glands and subarachnoid space. the mechanism of bleeding is unclear, but several factors contribute. these include thrombocytopenia, capillary endothelial damage, and coagulation defects resulting from hepatic dysfunction, consumptive coagulopathy and dic. patients with serositis may occasionally present with severe abdominal pain and peritonism. sometimes the abdominal pain is severe and indistinguishable from appendicitis, acute cholecystitis or cholangitis, especially in jaundiced patients, leading to unnecessary intra-abdominal surgery. acute pancreatitis has been reported rarely, although serum amylase may be raised in up to % of patients with severe disease due to renal impairment. leptospirosis in pregnancy may result in abortion, postpartum haemorrhage and intrauterine fetal death. congenital leptospirosis occurs rarely. other rare complications include erythema nodosum, life-threatening rhabdomyolysis associated with renal failure, and reactive arthritis. these complications appear in variable proportions and can overlap. weil's disease was first described by a german doctor, professor adolf weil, in in four patients with a febrile illness together with severe nervous symptoms, hepatosplenomegaly, jaundice and signs of renal disease which recovered rapidly. nowadays, the term 'weil's syndrome' usually refers to the extremely severe form of leptospirosis, characterized by the combination of jaundice, renal dysfunction, and haemorrhagic diathesis, especially pulmonary haemorrhage. this syndrome occurs in less than % of patients and carries a mortality rate of - %. overall mortality of patients hospitalized with leptospirosis varies among countries and series reports, and usually does not exceed %. anaemia is a prominent finding especially in severe cases, partly due to blood loss and haemolysis. white blood cell count can be normal ( %) or elevated with the median wbc count around - × /l and more than % neutrophil, with elevation of erythrocyte sedimentation rate and c-reactive protein concentrations. white blood cell counts higher than /mm are associated with poorer outcomes. the bleeding, prothrombin and activated partial thromboplastin times can be has similar epidemiology and clinical manifestations as leptospirosis. in addition, dual infection of leptospirosis and hantavirus has been reported. the clinical diagnosis of leptospirosis cannot be certain because of the nonspecific symptoms and signs which overlap with those of other common febrile diseases. in endemic areas, awareness should be raised during the rainy season. in some circumstances, such as natural disasters with fresh water exposure, health care workers should be alert for outbreaks. a definite diagnosis of leptospirosis is based on the isolation of leptospira from clinical specimens (mainly from blood or csf), or a positive microscopic agglutination test (mat), both of which are laborious and not widely available. although leptospira can survive in some commercial aerobic bacteria culture bottles, the recommended media used for isolation is the semi-solid ellinghausen, mccullough, johnson, and harris (emjh) medium. cultures are usually kept at - °c without co incubation. it may take - weeks for leptospira to multiply to detectable densities. cultures should be examined weekly by dark field microscopy. this technique is not routine practice in a standard microbiological laboratory. recently, a new solid medium called lvw agar was developed. it can increase the growth rate of leptospira. using this prolonged in % of patients. thrombocytopenia, a platelet count ≤ × /l, occurs in - % of patients. thrombocytopenia is an indicator of severe disease and risk of bleeding. acute febrile illness accompanied by jaundice and renal failure should always include leptospirosis in the differential diagnosis. a history of either indirect exposure to possible contaminated water or direct contact with animals occupationally or recreationally increases the probability of the diagnosis. many patients from the same community presenting with similar symptoms within a short period of time alerts to the likelihood of an outbreak and intervention may be needed to minimize the number of severe and fatal cases. however, the symptoms and signs of leptospirosis are not specific and can mimic many other acute febrile illnesses in endemic areas. in patient with marked jaundice, viral hepatitis has to be excluded. biliary tract infection may mimic leptospirosis especially in patients with severe upper abdominal pain. since the disease is found worldwide, the list of differential diagnoses is broad and may differ among different geographical areas. table . shows the differential diagnosis of leptospirosis according to symptoms and regions. of these, rickettsioses, especially scrub typhus, carries the same clinical spectrum and geographical distribution of the disease and completely overlaps with leptospirosis in asia-oceania. there have been several reports of co-infections from the south-east asia regions. sepsis is difficult to rule out in patients with severe forms, and co-infections can also occur. hantavirus, which cause haemorrhagic fever with renal syndrome and is associated with rodent exposure, several other serological tests have been developed and used in many places, although the sensitivity, specificity and accuracy of the tests are generally unsatisfactory. all of these tests are non-serovar-specific, so they have no epidemiological usefulness, and are used for screening purposes only. examples of these tests are the immunochromatographic test (ict) or cassette test, microcapsule agglutination test (mcat), and latex agglutination test (la). standard pcr diagnostics are not available for leptospirosis. pcr methods using various target genes have been developed. real time pcr may be a good candidate for future confirmatory tests. leptospira are broadly sensitive to many antibiotics; however the efficacy of antibiotic treatment has been confirmed in only small series. because of the biphasic course of the disease, the usefulness of antibiotic treatment has been doubted when given in the second immune phase. there are insufficient randomized controlled trial data to provide clear evidence on the efficacy of treatment at different stages of the infection. there have been seven randomized controlled trials; four trials with patients compared an antibiotic with placebo or no intervention. penicillin shortened the fever time, times to normalization of creatinine level and hospitalization in a small randomized, placebo-controlled trial even late in the course of disease in thailand, but showed a non-significant trend to reduction in mortality in brazil when given after days of illness. despite these uncertainties effective treatment should be given in every patient as soon as possible at any stage of the disease. the recommended drugs and doses are shown in table . . all regimens, except -day azithromycin, should be given for days. penicillin, cefotaxime and ceftriaxone can be switched to oral amoxicillin if patients improve and can tolerate oral drug treatment. oral doxycycline remains the drug of choice in uncomplicated cases of leptospirosis. alternative choices for patients with novel agar, simple antimicrobial susceptibility testing, using the e-test can provide clear cut-offs and easy-to-read minimal inhibitory concentrations (mic) of antimicrobials currently recommended for treatment (figure . ). the novel lvw agar may open a new era in the clinical practice for diagnosis and rapid antimicrobial susceptibility testing of leptospirosis. isolation of leptospires should be performed from blood or csf of untreated patients who present not later than days after onset of illness. antibiotic pre-treatment substantially decreases the yield of the isolation. isolation from urine is less useful, since leptospiruria usually occurs during the second week in which antibiotics have been given and antibodies are readily detectable. the acidity of the urine reduces the sensitivity of culture. the gold standard serological test is mat. the test is serovarspecific and useful for epidemiological studies. it detects mixed igg and igm. the mat test is considered positive if there is either a fourfold rise of the convalescent titre compared to the acute titre, or there is a titre of more than or equal to : in single or paired sera. this may not be useful for patient care, people at risk of exposure and people living in endemic areas should be informed about the disease and the risks. the education of people at risk and awareness of healthcare workers enable early detection and treatment. reducing rodent populations in housing and working areas may reduce the risk of exposure. however, the disease circulates mainly in mammals other than rodents. immunization of domestic farm animals and pets reduces the overall risks to humans significantly. the use of personal protective measures such as long boots, gloves and protective clothing are recommended and should be emphasized in all workers at risk. chemoprophylaxis is recommended in people at particularly high risk. doxycycline is effective for prevention when given at mg weekly during high-risk exposure. the main adverse effect is gastrointestinal irritation causing vomiting, so the drug should be administered with food. human serovar-specific, short-term protective vaccines are available in china, cuba, france and russia. the efficacy of the vaccines in other areas have never been evaluated. genome sequencing of all pathogenic leptospira species may provide the opportunity to identify new vaccine candidates which ultimately aim to protect against all eight pathogenic leptospira species. hypersensitivity and pregnant or lactating woman are oral amoxicillin or azithromycin. in severe cases, penicillin, doxycycline, ceftriaxone and cefotaxime have similar efficacy. however, in south-east asia, the use of doxycycline or a cephalosporin in combination with doxycycline are recommended because scrub typhus co-infection is common. supportive care in severely ill patients is critical in leptospirosis. careful fluid resuscitation should be given in patients with hypovolaemic shock. inotropic drug administration should be given to patients with refractory shock despite effective fluid replacement. haemodialysis should be started early in patients with hypercatabolic renal injury. daily haemofiltration or dialysis may be needed in some patients, and renal replacement should be continued until clear improvement. bleeding precautions are needed; however, routine use of proton-pump inhibitors and h blockers is not recommended and should be used with caution, especially in patients who are intubated, as they may increase the occurrence of ventilator-associated pneumonia. respiratory support is mandatory in patients with respiratory failure either from ards or severe pulmonary haemorrhage. ventilation should be supervised in the intensive care unit care to maintain adequate oxygenation. the use of high-dose steroids is not indicated and may be harmful predisposing to superimposed bacterial infections. a century of leptospira strain typing emergence, control and re-emerging leptospirosis: dynamics of infection in the changing world new insights into the pathogenicity of leptospires: evasion of host defences rapid isolation and susceptibility testing of leptospira spp. using a new solid medium, lvw agar antibiotic prophylaxis for leptospirosis a century of leptospira strain typing genome reduction in leptospira borgpetersenii reflects limited transmission potential genome sequence of the saprophyte leptospira biflexa provides insights into the evolution of leptospira and the pathogenesis of leptospirosis diagnostic accuracy of real-time pcr assays targeting s rrna and lipl genes for human leptospirosis in thailand: a case-control study a waterborne outbreak of leptospirosis an institutional outbreak of leptospirosis in chennai, south india human-to-human transmission of leptospira interrogans by milk the globalization of leptospirosis: worldwide incidence trends emergence, control and re-emerging leptospirosis: dynamics of infection in the changing world report of the second meeting of the leptospirosis burden epidemiology reference group leptospira: the dawn of the molecular genetics era for an emerging zoonotic pathogen in vitro evidence for immune evasion activity by human plasmin associated to pathogenic leptospira interrogans leptospiral lipopolysaccharide activates cells through a tlr -dependent mechanism new insights into the pathogenicity of leptospires: evasion of host defences liver biopsy in human leptospirosis: a light and electron microscopy study kidney biopsies in human leptospirosis: a biochemical and electron microscopy study acute kidney injury in human leptospirosis: an immunohistochemical study with pathophysiological correlation immunohistochemical and in situ hybridization studies of the liver and kidney in human leptospirosis activation of the coagulation cascade in patients with leptospirosis lung lesions in human leptospirosis: microscopic, immunohistochemical, and ultrastructural features related to thrombocytopenia melbourne: medisci factors associated with clinical leptospirosis: a populationbased case-control study in the seychelles (indian ocean) rapid isolation and susceptibility testing of leptospira spp. using a new solid medium, lvw agar an open randomized controlled trial of desmopressin and pulse dexamethasone as adjunct therapy in patients with pulmonary involvement associated with severe leptospirosis antibiotic prophylaxis for leptospirosis access the complete references online at www.expertconsult.com. key: cord- - xmxcm authors: walden, andrew p; clarke, geraldine m; mckechnie, stuart; hutton, paula; gordon, anthony c; rello, jordi; chiche, jean-daniel; stueber, frank; garrard, christopher s; hinds, charles j title: patients with community acquired pneumonia admitted to european intensive care units: an epidemiological survey of the genosept cohort date: - - journal: crit care doi: . /cc sha: doc_id: cord_uid: xmxcm introduction: community acquired pneumonia (cap) is the most common infectious reason for admission to the intensive care unit (icu). the genosept study was designed to determine genetic influences on sepsis outcome. phenotypic data was recorded using a robust clinical database allowing a contemporary analysis of the clinical characteristics, microbiology, outcomes and independent risk factors in patients with severe cap admitted to icus across europe. methods: kaplan-meier analysis was used to determine mortality rates. a cox proportional hazards (ph) model was used to identify variables independently associated with -day and six-month mortality. results: data from patients admitted to centres across countries was extracted. median age was years, % were male. mortality rate at days was %, rising to % at six months. streptococcus pneumoniae was the commonest organism isolated ( % of cases) with no organism identified in %. independent risk factors associated with an increased risk of death at six months included apache ii score (hazard ratio, hr, . ; confidence interval, ci, . - . ), bilateral pulmonary infiltrates (hr . ; ci . - . ) and ventilator support (hr . ; ci . - . ). haematocrit, ph and urine volume on day one were all associated with a worse outcome. conclusions: the mortality rate in patients with severe cap admitted to european icus was % at six months. streptococcus pneumoniae was the commonest organism isolated. in many cases the infecting organism was not identified. ventilator support, the presence of diffuse pulmonary infiltrates, lower haematocrit, urine volume and ph on admission were independent predictors of a worse outcome. community acquired pneumonia (cap) is common, affecting between and individuals per , of the adult population each year [ , ] and is the commonest cause of sepsis, severe sepsis and septic shock [ ] . between and % of patients require hospital admission [ , , ] of whom to % will be admitted to an icu [ ] [ ] [ ] . hospital and icu admission rates for cap are increasing for all ages [ ] . an ageing, more vulnerable population, earlier recognition of deteriorating patients and better availability and use of intensive care beds may in part explain this increase. a number of investigators have examined the clinical and microbiological factors that might affect the outcome from severe cap [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] , but these studies have often been relatively small, with considerable heterogeneity in both the inclusion criteria and outcome measures. as a consequence reported mortality rates have varied and there has been uncertainty regarding the most important risk factors for death. a number of more recent, larger studies have focussed on identifying patients with cap at increased risk of severe sepsis and death, as well as those who may require ventilator or vasopressor support [ , [ ] [ ] [ ] . one of these studies provides outcome data at days for the smaller subgroup of patients with severe sepsis admitted to the icu [ ] , and another [ ] reports microbiological findings and -day mortality in a subgroup of patients admitted to the icu. the aim of the study reported here was to define the clinical characteristics, microbiological aetiology, outcomes and independent risk factors for mortality in a large, contemporary cohort of patients with severe cap admitted to icus across europe. by using such a large database with clear censor data for mortality we hoped to overcome some of the deficiencies of previous studies. the genosept study was conceived by the european critical care research network (eccrn), the research arm of the european society for intensive care medicine (esicm) with the aim of defining genetic influences on the host response and outcomes in patients with sepsis. genosept is a pan-european study that has recorded comprehensive phenotypic data and obtained dna from a large cohort of patients admitted to icu with sepsis due to cap, peritonitis, meningococcal disease or pancreatitis. ethics approval was granted nationally, for individual centres, or for both. written, informed consent was obtained from all patients or a legal representative. patients reported here were recruited into genosept from centres across countries (see additional file for contributors) over a -year period between september and october . inclusion criteria were as follows: admission to a high dependency unit (hdu) or icu with cap, and age over years. the diagnosis of cap was defined as a febrile illness associated with cough, sputum production, breathlessness, leucocytosis and radiological features of pneumonia acquired in the community or within less than days of hospital admission [ ] . the diagnosis of sepsis was based upon the international consensus criteria published in [ ] . patients were further sub-classified according to the criteria for severe sepsis and septic shock. exclusion criteria were as follows: patient or legal representative unwilling or unable to give consent; patient under yrs of age; patient pregnant; advanced directive to withhold or withdraw life-sustaining treatment or admitted for palliative care only (please see also additional file ). patients were observed until death or for a maximum of months. in those who had died between icu discharge and -month follow-up the date of death was recorded. specific data was recorded in the electronic case report form (ecrf) to allow calculation of the acute physiology and chronic health evaluation ii (apache ii) and sequential organ failure assessment (sofa) scores [ , ] . the infectious diseases society of america/american thoracic society (idsa/ats) criteria were used for the diagnosis of severe cap [ ] . co-morbidities were recorded according to the modified charlson scoring system [ ] . chest radiograph appearances were recorded as: ) lobar, ) localised or ) diffuse bilateral. investigators were also asked to consider the differential diagnosis of cardiogenic pulmonary oedema in those with diffuse pulmonary infiltrates. microbiological investigations were performed according to local policies and practices. investigators recorded the microbiological findings, including the organism(s) isolated, the source of the organism and the use of serology. mortality at days and at months were chosen as primary endpoints. univariate analysis was performed using a cox proportional hazards model, adjusted for age and gender. variables considered by the investigators to be clinically relevant were chosen for analysis (see additional file ). a test for proportional hazards using the schoenfeld residuals was performed, and for covariates indicating evidence of non-proportionality, spline smooth estimates of time-dependent hazard ratios (hrs) with point-wise confidence bands were calculated [ ] . variables that were significant in the univariate analysis after bonferroni adjustment for multiple testing (p-value < . /k where k is the number of variables tested) were entered into a multivariate cox proportional hazards model to determine independent risk factors for death. a final set of predictors for each of -day and -month mortality was selected from these variables via a stepwise cox proportional hazards regression model. r software version . . was used for all data analysis. between september and october , , patients were enrolled. four individuals were excluded because of missing or inconsistent data. patient characteristics are shown in table . on admission, , patients ( %) met idsa/ats criteria for severe cap, on major criteria and on minor criteria. median age was years (range to , iqr to ): % were male, % were caucasian and % mediterranean, and patients were hospitalised for more than hours prior to icu admission. of these the mean time from hospital admission to icu admission was . days. median length of stay in the icu was days (iqr to ); median length of stay in hospital was days (iqr to ) ( table ) . a total of ( %) patients had one or more co-morbid conditions, with cardiac and respiratory disease affecting and patients, respectively. chronic obstructive pulmonary disease was documented in ( . %) patients, diabetes mellitus in . % and a chronic neurological condition in . %. on the day of admission ( %) patients required mechanical ventilation, with the number increasing to patients ( %) within the first week of admission ( table ). median duration of mechanical ventilation was days: patients ( %) satisfied the criteria for septic shock. the median duration of inotrope/vasopressor support was days. renal replacement therapy was required during the first week of admission in % of patients. median sofa score on admission was (iqr to ) and median apache ii score was (iqr to ). chest radiograph appearances were recorded as lobar consolidation in . %, patchy localised consolidation in . % and diffuse, bilateral changes in . %. a total of ( . %) patients had died within months of enrolment, ( . %) in the icu, in hospital after icu discharge and between hospital discharge and months following icu admission. the in-hospital mortality rate was . % and the -day mortality rate was . % (see table and figure ). mortality was higher in mechanically ventilated patients, ( . % at days and . % at months) and in patients with septic shock receiving haemodynamic support with inotropes or vasopressors ( . % and . % respectively). the standardised mortality ratio derived from the apache ii score was . ( % ci . , . ) [ ] . no causative organism was identified in over a third of patients (table ) . streptococcus pneumoniae (pneumococcus) was the most commonly isolated organism ( % of cases), positive microbiology was obtained from lung secretions/lavage in % of cases; blood culture in %; urinary antigen testing in %; blood serology in %; pleural fluid in . % and other methods in %. bacteraemia was present in ( %) of patients and pleural infection/empyema in ( %). pneumococcus was the commonest organism causing bacteraemia and empyema, accounting for episodes ( %) and episodes ( %) respectively. atypical organisms and viruses were rarely identified. isolation of staphylococcus aureus, presence of septic shock, need for mechanical ventilation and renal replacement therapy all showed a strong association with outcome (table ) . the factors most strongly associated with -day mortality were the apache ii score calculated on day (hr = . [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . the considerable heterogeneity in admission policies, study design, guidelines compliance [ ] , and severity scoring in these studies probably accounts for the wide range of reported mortality rates and makes meaningful comparisons difficult. most of these studies have used icu admission rather than severity scores to indicate severe disease. only three studies [ , , ] defined censor points for death, which is important as in-hospital mortality increases following icu discharge by between and % [ ] . admission practices in different countries may also lead to large ranges in mortality. take for instance one study of patients admitted to a spanish respiratory icu in the s the mortality rate was % but with rates of mechanical ventilation and septic shock of % and % respectively [ ] , whereas in a uk study published in the mortality rate was %, with mechanical ventilation and septic shock rates of % and % respectively [ ] . the mortality rates reported here are more in keeping with other recent, large cohorts. the capuci group analysed patients admitted to over spanish icus between and and found icu mortality rates of % with associated apache ii scores of [ ] . they included both immunocompetent and immunosuppressed patients. in the immunocompetent individuals the rate of death at icu discharge was slightly lower at %, a figure that is closer to the % seen in the genosept cohort, (in which immunocompromised patients were [ ] . in the port study, of , patients were admitted to the icu with an in-hospital mortality rate of . % [ ] . the standard censor point for most interventional icu studies is days, although it is recognised that there is a significant attrition rate post icu discharge. it is also well-recognised that in cap patients there is an increased death rate in the months following discharge [ ] , and in patients with sepsis there is significant excess mortality for at least five years [ ] . in one study, the death rate of icu patients between days and months was % in patients with sepsis, similar to the % seen in icu patients without sepsis [ ] and the % found in the present study. this compares to an increase in mortality from . % at days to . % at days in the subgroup of icu patients in the port study [ ] and an increase from . % to . % at months in the genims cohort of icu patients [ ] . although the microbiological methodology was not standardised, our findings are consistent with previous studies of severe cap. notably, streptococcus pneumoniae accounted for % of all cases and no aetiological agent was identified in over a third [ ] . within the genosept cohort the rate of streptococcus pneumoniae was higher than previously reported and was mirrored by a decrease in the number of cases where no aetiological agent was identified, suggesting that detection rates may have improved, rather than there being a true increase in all values are significant at a type i error rate of % after a bonferroni correction to take account of the multiple testing of variables (a p-value < . was considered to be statistically significant). results are adjusted for age and sex. all day- variables unless specified otherwise. rrt, renal replacement therapy; mv, mechanical ventilation; niv, non-invasive ventilation; sbp, systolic blood pressure; map, mean arterial pressure. incidence. pneumococcal antigen testing in urine and other bodily fluids has become the standard of care in many institutions and has good positive and negative predictive value both in hospitalised cap patients [ ] and in those admitted to icu [ ] . amongst the patients with confirmed pneumococcal pneumonia in the present study, a total of positive results were obtained. of these ( %) were positive urinary antigen tests and ( %) were based on positive blood serology. further evidence to support this apparent increase in detection rates is provided by a recent study that reported detection rates of % for streptococcus pneumoniae, with % of cases being diagnosed using antigen testing [ ] . in comparison, the bts study from reported a rate of streptococcus pneumoniae infection of %, with positive antigen testing in only % of patients [ , ] . in hospitalised patients with cap, as many as to % may involve a viral infection, the virus being the only organism isolated in % [ ] . failure to respond to standard antimicrobial therapy means that more patients with viral pneumonia are likely to be admitted to icu. certainly patients with co-morbidities have a higher incidence of viral infection. in the present study viral infections were rarely identified. this raises the question as to whether a more aggressive search for viral pathogens should be conducted in all ventilated patients, coupled with more frequent and targeted use of antiviral therapy. multivariate analysis identified four variables (apache ii score, haematocrit, mechanical ventilation and ph) that were independently associated with outcome at both days and months the need for mechanical ventilation was related to a worse outcome at both days and months, consistent with other data showing respiratory failure to be an independent predictor of mortality in many categories of critically ill patients [ , ] . the persistence of this relationship for up to months reflects the fact that many patients who have received ventilator support will continue to have significant neuromuscular weakness and be at risk for a prolonged period after discharge from the icu [ ] . it may be that over-aggressive use of intravenous fluids, reflected in a dilutional reduction in haematocrit worsens lung injury and thus prolongs the need for mechanical ventilation. conservative fluid management has been associated with better outcomes, albeit in the later phases of critical illness [ ] . a key element of early goal-directed therapy in patients with sepsis is blood transfusion to maintain the haematocrit, perhaps accounting for the positive association between better outcomes and a higher haematocrit in the present, and other studies [ ] . similarly the positive independent relationship between a higher ph on admission and a better outcome may reflect more effective early resuscitation. although there was no association between the admission p:f ratio and outcome, diffuse bilateral changes on the chest radiograph (suggesting a diagnosis of acute lung injury/acute respiratory distress syndrome) independently predicted a worse outcome at months. this is consistent with other studies showing a mortality rate for ards much higher than that seen in our cohort of patients with cap [ ] . urine volume, renal failure and the need for renal replacement therapy were associated with worse outcome in the univariate analysis and there was a clear independent association between urine output and mortality at months. acute kidney injury (aki) has been shown to be independently associated with higher icu and in hospital mortality rates [ , ] . the need for ongoing renal support in those with aki is estimated to be %, perhaps explaining the association with outcome at months. other studies have attempted to determine independent risk factors for death from cap. for example, the presence of septic shock has been associated with odds ratios for risk of death ranging from to but inevitably with wide confidence intervals due to the small numbers of patients included in these analyses [ , , , , ] ; also the lack of a consistent censor point complicates interpretation of these findings. we found an association between septic shock and outcome on univariate analysis but this effect was not seen in multivariate analysis, perhaps reflecting improvements in the acute management of shock. this study has two important limitations. firstly participating centres were at liberty to decide which patients they would enrol; subjects were not, therefore, enrolled consecutively, thereby introducing a potential for selection bias. also there was considerable variation in the number of patients recruited in each country and some centres contributed only small numbers of patients. nevertheless there was a wide range of ages, severity of physiological derangement and co-morbidities, whereas apache ii scores and ventilation rates were similar to previous studies, suggesting that a significant, systematic selection bias is unlikely. secondly, microbiological protocols were not standardised. on the other hand our observations therefore reflect current approaches to microbiological diagnosis in routine clinical practice across europe. the icu mortality rate in this contemporary cohort of patients admitted to icus across europe with severe cap was %, rising to % at hospital discharge and % at months. streptococcus pneumoniae was the commonest cause of cap, but in many cases the infecting organism was not identified. the need for ventilator support, and the presence of diffuse bilateral infiltrates on the chest radiograph, as well as lower haematocrit, urine volume and ph on admission to icu were independent predictors of a worse outcome. prospective study of the aetiology and outcome of pneumonia in the community incidence of community-acquired pneumonia in the population of four municipalities in eastern finland understanding the inflammatory cytokine response in pneumonia and sepsis: results of the genetic and inflammatory markers of sepsis (genims) study community-acquired pneumonia: the annual cost to the national health service in the uk severe community-acquired pneumonia. epidemiology and prognostic factors prognostic factors of pneumonia requiring admission to the intensive care unit the british thoracic society research committee and the public healthlaboratory service: the aetiology, management and outcome of severe community-acquired pneumonia on the intensive care unit increasing hospital admissions for pneumonia combination antibiotic therapy with macrolides improves survival in 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in community-acquired pneumonia sccm/esicm/accp/ats/sis international sepsis definitions conference apache ii: a severity of disease classification system the sofa (sepsis-related organ failure assessment) score to describe organ dysfunction/failure. on behalf of the working group on sepsis-related problems of the european society of intensive care medicine infectious diseases society of america/american thoracic society consensus guidelines on the management of community-acquired pneumonia in adults a new method of classifying prognostic comorbidity in longitudinal studies: development and validation modeling survival data: extending the cox model communityacquired pneumonia on the intensive care unit: secondary analysis of , cases in the icnarc case mix programme database inflammatory markers at hospital discharge predict subsequent mortality after pneumonia and sepsis long-term mortality and quality of life in sepsis: a systematic review quality of life of survivors from severe sepsis and septic shock may be similar to that of others who survive critical illness community-acquired pneumonia in europe: causative pathogens and resistance patterns evaluation of the immunochromatographic binax now assay for detection of streptococcus pneumoniae urinary antigen in a prospective study of community-acquired pneumonia in spain evaluation of the binax now streptococcus pneumoniae urinary antigen assay in intensive care patients hospitalized for pneumonia viral infection in adults hospitalized with community-acquired pneumonia: prevalence, pathogens, and presentation canadian critical care trials group: one-year outcomes in survivors of the acute respiratory distress syndrome comparison of two fluid-management strategies in acute lung injury sepsis in european intensive care units: results of the soap study beginning and ending supportive therapy for the kidney (best kidney) investigators: acute renal failure in critically ill patients: a multinational, multicenter study effect of acute renal failure requiring renal replacement therapy on outcome in critically ill patients submit your next manuscript to biomed central and take full advantage of: • convenient online submission • thorough peer review • no space constraints or color figure charges • immediate publication on acceptance • inclusion in pubmed, cas, scopus and google scholar • research which is freely available for redistribution the genosept project was supported by the european critical care research network and the european society of intensive care medicine. we would like to acknowledge all the co-investigators and research nurses who were involved in the project. a list of their names can be found in the additional file . the mortality rate from severe cap in patients admitted to icu is % at six months streptococcus pneumonia remains the most commonly isolated organism no microbiological diagnosis is found in a third of patients the need for mechanical ventilation is a strong predictor of a poor outcome ph, haematocrit, urine output and diffuse changes on chest radiography all predict a worse outcome authors' contributions apw prepared the database for analysis, prepared the first copy of the manuscript and coordinated all manuscript revisions; gmc performed the primary statistical analysis, helped in the writing of the manuscript and provided the tables and figures; smck assisted in preparation of the database for analysis and helped in writing and reviewing the manuscript; ph helped with the preparation of the database for analysis and helped in writing and reviewing the manuscript; acg helped in the design of the genosept project and in the writing an reviewing of the manuscript; jr helped in the design of the genosept project and in writing and reviewing the manuscript; j-dc helped in the design of the genosept project and in writing and reviewing the manuscript; fs helped in the design of the genosept project and in writing and reviewing the manuscript; csg helped in the design of the genosept project and in writing and reviewing the manuscript; cjh helped in the design of the genosept project and in writing and reviewing the manuscript. all authors read and approved the final manuscript. andrew p walden and geraldine m clarke contributed equally as joint first authors. key: cord- -x ddi wg authors: li, wanli; an, xinjiang; fu, mingyu; li, chunli title: emergency treatment and nursing of children with severe pneumonia complicated by heart failure and respiratory failure: case reports date: - - journal: exp ther med doi: . /etm. . sha: doc_id: cord_uid: x ddi wg pneumonia refers to lung inflammation caused by different pathogens or other factors, and is a common pediatric disease occurring in infants and young children. it is closely related to the anatomical and physiological characteristics of infants and young children and is more frequent during winter and spring, or sudden changes in temperature. pneumonia is a serious disease that poses a threat to children's health and its morbidity and mortality rank first, accounting for . – . % of pediatric inpatients. due to juvenile age, severe illness and rapid changes, children often suffer acute heart failure, respiratory failure and even toxic encephalopathy at the same time. the concurrence in different stages of the process of emergency treatment tends to relapse, which directly places the lives of these children at risk. severe pneumonia constitutes one of the main causes of infant mortality. in the process of nursing children with severe pneumonia, intensive care was provided, including condition assessment and diagnosis, close observation of disease, keeping the airway unblocked, rational oxygen therapy, prevention and treatment of respiratory and circulatory failure, support of vital organs, complications, and health education. the inflammatory response was proactively controlled, to prevent suffocation and reduce mortality. in summary, positive and effective nursing can promote the rehabilitation of children patients, which can be reinforced with adequate communication with the parents and/or caretakers. severe pneumonia is a common life-threatening disease, particularly for children, and is more common in infants and young children ( , ) . the estimated worldwide incidence of severe pneumonia in children less than years of age is - per , person-years and the mortality is . - . per , person-years ( ) ( ) ( ) . it often occurs in the winter and spring, with acute onset, complex clinical manifestations and fast-changing condition, which usually involves the circulation, nervous and digestive systems ( ) ( ) ( ) . as a result, severe pneumonia produces corresponding clinical symptoms, such as respiratory failure, heart failure, toxic encephalopathy and intestinal paralysis, which endanger the lives of children in the short term, and is the first cause of death of pediatric inpatients ( , ) . it is listed as the one of the four diseases requiring prevention and treatment in children by the ministry of health ( , ) . pneumonia may occur at any point of the year, but is more common in the months of winter and spring or during the time of climate variability. the disease may be the primary illness, or secondary one after an acute infectious disease, such as bronchitis and measles, upper respiratory tract infections and whooping cough, and has a high morbidity and mortality rate in china ( ) . as reported by several studies, the number of annual pneumonia patients in china is ≤ million individuals, and among the children with pneumonia, - % cases are of severe pneumonia, which ranks fifth amongst various diseases leading to death ( ) ( ) ( ) ( ) ( ) ( ) . the aim of this study was to conduct a retrospective analysis of the clinical data of cases of children presenting with severe pneumonia at the xuzhou children's hospital between january and june , retrieve and review the literature, and summarize emergency treatment and nursing experience. general information. ten cases of children diagnosed with severe pneumonia according to the guidelines of the world health organization were included in the present study ( ) . in this group, there were males and females, aged month to years. the children patients were hospitalized due to fever, cough, asthma, dry and wet rales could be heard in lung auscultation, and a chest x-ray showed thickened lung markings with visible punctate and flake-like shadows. the children patients refused any intake of dairy products or food. there was one case of concurrent toxic encephalopathy, case of gastrointestinal bleeding, and case of toxic intestinal paralysis. the physical examination showed a body temperature of . - ˚c, pulse rate of - beats/min, breathing of - beats/min, and dry and wet rales could be heard in lung. the x-ray examination showed thickened lung markings, and flake-like shadows, and abdominal palpation showed enlarged liver and spleen. respiratory or circulatory functions were at different levels of exhaustion. treatment. after admission, comprehensive emergency treatment measures were taken, such as improving the ventilation function, oxygen uptake, maintaining airway patency, administration of cardiac and diuretic drugs, reduction of intracranial pressure, and conducting anti-infection and symptomatic support measures. nine cases were successfully treated, case succumbed to respiratory failure, and the success rate was %. condition assessment and diagnosis. a detailed inquiry of the medical history was carried out to determine whether the children patients had a history of recurrent respiratory tract infection, and whether they had measles, whooping cough and other respiratory diseases prior to onset, as well as whether the children patients had a full-term birth or asphyxia, and if the growth and development of children patients after birth was normal ( ) ( ) ( ) . conditions such as fever, cough, extent of choked asthma, the presence or absence of breathing, faster heart rate, pulmonary rales, orthopnea, nose flap, three depression signs and cyanosis, as well as clinical manifestations of infected circulatory, digestive, and nervous system were assessed. evaluation of blood routine examination, chest x-ray, and etiology test results were conducted. psychological and social conditions of children and parents were also assessed ( ) . common condition assessment and diagnostic criteria included: i) air exchange impairment, related to lung inflammation; ii) invalid airway clearance, related to excessive and sticky respiratory secretions, frail children patients, and inability to expectorate; iii) hyperthermia, related to lung infection; and iv) malnutrition, related to inadequate intake, increased consumption. close observation of the condition. attention was paid to changes in total peripheral resistance blood pressure (bp), the presence of double suction, such as nodding-like breathing, apnea and other conditions indicating respiratory failure. the presence of exacerbation of dyspnea, dysphoria, quickening heart rate, and an enlarged liver in a short period of time suggested heart failure ( ) ( ) ( ) . children patients with severe wheezing often suffered respiratory acidosis due to retention of carbon dioxide. drowsiness, convulsions or coma in sick children suggested an occurrence of toxic encephalopathy. medicines such as antibiotics, antiasthmatic drugs, cardiac drugs and other medications were administered to patients at the appropriate dosage, the appropriate time and in an accurate manner. side effects of various drugs were observed, and if children patients showed signs of dysphoria, quickening heart rate, worsening asthma, or enlarged liver in a short period of time, indicating heart failure, then the infusion rate was reduced. in our study, close observation of consciousness, pupil changes and muscle tension was carried out, and if manifestations of intracranial pressure such as drowsiness, convulsions, irregular breathing and increased muscle tension appeared, immediate rescue steps were taken. abdominal distension and decreased or disappearing bowel sounds were observed, in order to detect toxic intestinal paralysis in time ( ) . emergency treatment and therapy for respiratory failure. type i respiratory failure refers to the lone presence of hypoxemia and absence of hypercapnia, featuring ventilation dysfunctions, blood change of pao ≤ mmhg, and paco which can be maintained at normal level or reduced ( ) . type i respiratory failure also refers to the coexistence of hypoxemia and hypercapnia, impairment of ventilatory function and gas exchange functions, severe lung lesion, obstruction of trachea and bronchia caused by sticky secretions, blood change of pao < mmhg, and paco > mmhg. main clinical manifestations of children patients with type i pneumonia with respiratory failure include, poor mental state or dysphoria, polypnea, cyanosis of lips, dyspnea, nasal flaring and three depression signs. these symptoms are difficult to distinguish from type ii respiratory failure, and can be observed only by blood gas analysis, which shows a marked difference between type i and ii. type ii respiratory failure shows symptoms of type i respiratory failure and in addition more often than not, it also has symptoms such as shallow breathing, irregular rhythm, slow breathing, drowsiness or coma and even jaw breathing in some patients ( , ) . in our study, changes in condition and changes in blood were closely observed, and where required, a tracheal intubation ventilator was employed to improve the cure rate. oxygen uptake. nasal catheter is easily blocked by secretions, leading to failure of effective uptake oxygen, and is therefore difficult to utilize. we employed the conventional oxygen mask inhalation method, which is comfortable and without stimulation, and was easily accepted by children ( , ) . after using oxygen, once hypoxia was improved, timely adjustment of the oxygen flow or deactivation of oxygen was needed, because inadequate oxygen concentration such as excessive or overtime concentration may lead to changes such as lung tissue edema atelectasis and proliferation of alveolar capillary. the humidification bottle contained % alcohol, aiming to reduce the alveolar surface tension and help improve ventilation. maintenance of airway patency and administration of various aerosol inhalation therapies. i) this group of children patients took appropriate clinostatism according to the state of disease. under normal conditions, the children were placed in a horizontal position, with neck raised high, in order that airways would be unblocked. for children patients with severe heart failure and wheezing, semi-recumbent position was taken to reduce the burden on the heart and lungs ( , ) . for children patients with increased respiratory secretions, the lateral position was useful to expectorate and prevent aspiration. ii) patients received daily routine aerosol inhalation, turning drugs into aerialfog-like fine particles, inhaled to the bronchioles or alveoli, which diluted secretions and had an anti-inflammatory and anti-allergic function and reduced local inflammatory exudates, in addition to reducing airway resistance and improving ventilation. aerosol inhalation time was - min and afterwards sputum suction was carried out immediately in order to clear respiratory secretions. commonly used drugs included: , units of gentamicin, - mg of α-chymotrypsin, - mg of dexamethasone, and ml of saline. the dual role of the aforementioned ultrasonic aerosol inhalation was useful to reinforce treatment of children with severe pneumonia. according to the disease, aerosol inhalation was conducted every or h, with sputum suction ensuing, which produced a good suction effect for children patients with many sticky respiratory secretions. it should be noted that if patients had excessive sputum, the sputum suction was also carried out once prior to using ultrasound aerosol inhalation therapy, followed by thorough sputum suction. in the whole process of aerosol inhalation, providing oxygen inhalation or enhancing oxygen concentration effectively prevented the occurrence of hypoxemia ( ) . iii) hypocalcemia may produce laryngospasm and tongue tenesmus, which causes obstruction and sudden suffocation. first, the child patient's tongue tip was pulled outside the mouth, and then artificial respiration was conducted, with most patients being able to relieve themselves spontaneously. pressurized oxygen was given, endotracheal intubation was carried out if necessary and calcium was replenished immediately. iv) when nasal obstruction caused breathing difficulties, % ephedrine drops were used after clearing away nasal secretions with a cotton swab, to maintain the airway patency. v) the diet of children with severe pneumonia was affected due to high fever, vomiting, diarrhea and other factors. when children patients were unable to eat, it was appropriate to supplement water, electrolytes, vitamins, and give them high-calorie, high protein, digestible food, that was eaten in small amounts but frequently, to prevent satiety from interfering with respiratory function. vi) previous findings showed that, ambroxol and low-dose heparin coupled with aerosol inhalation significantly shortened the time of the disappearance of all the clinical signs and improved overall efficiency of the clinical treatment ( , ) . there was no bleeding and other adverse reactions involved in the clinical treatment process, which had advantages such as easy administration, safe use, inexpensiveness, a significant effect, and less adverse effects in the treatment of children with severe pneumonia, and was thus worthy of wider application. since the application of mechanical ventilation is prone to cause ventilator-related complications, to improve the successful rescue rate of children with severe pneumonia and shorten the course of treatment, the use of ncpap may be considered a priority. for severe pneumonia, ncpap can support cardiopulmonary function, and its early application can improve timely oxygenation, stabilize disease, prevent disease progression, reduce ventilator application and average hospitalization time in pediatric intensive care unit as well as avoid the adverse effects caused by intubation ( ) . however, two points are important when applying it to infants: i) grasp indications and standard of respiratory failure that the children comply with; ii) over ncpap application process, nasal and oral secretions should be immediately removed, raising the neck high, to strengthen expectoration in order to maintain airway patency. operating parameters of using the ncpap oxygen therapy machine in our department: i) oxygen concentration inhaled (fio ) or %; ii) the oxygen flow - l/min; and iii) the beginning pressure was maintained at - cm h o. after observing - h, the adjustment of pressure was determined by clinical symptoms, signs and blood gas analysis. ncpap is an effective treatment for severe pneumonia with respiratory and heart failure, and can quickly correct hypoxemia, reduce endotracheal intubation and mechanical ventilation demand ( ) . it has a positive effect and can reduce the stimulation of children, thus decreasing any occurrence of ventilator pneumonia. in addition, parents readily accepted this treatment ( ) . short-term intubation ventilator was utilized when there was: i) respiratory arrest or respiratory and cardiac arrest; ii) sputum congestion, dyspnea, serious cyanosis; and iii) blood paco > mmhg, in order to accelerate the discharge of paco and reduce respiratory acidosis. emergency treatment and treatment of heart failure. when children patients coughed, their body temperature increased to ≤ ˚c and was accompanied by dyspnea, dysphoria, pale face, cyanosis, drowsiness, fixed moist rales in lung, respiratory rate of ≥ beats/min, heart rate of ≥ - beats/min in quiet state, low and blunt heart sound, gallop rate, and a progressively enlarged liver, cm beyond the ribs or an increase of ≥ . cm in a short period of time. the above, as well as cold extremities and weak pulse, constituted clinical manifestations of a heart failure. in this situation, oxygen inhalation and tranquilizers were given immediately, . µg/(kg x min) and milrinone, a second generation of phosphodiesterase Ⅲ (pde-Ⅲ) inhibitor, which acts on the β receptor accessory pathway, was administered to maintain heart function. by selectively inhibiting pde-Ⅲ in myocardial cells, increasing intracellular cyclic adenosine monophosphate levels, promoting calcium influx, enhancing calcium concentration in myocardial cytolymph, milrinone strengthens myocardial contractility, increases cardiac output, and exerts direct relaxation on vascular smooth muscle, dilates blood vessels and reduces cardiac preload and afterload. milrinone demonstrates a better clinical efficacy in the treatment of children with pneumonia complicated by heart failure than catecholamines ( ) . emergency treatment and treatment of toxic encephalopathy and intestinal paralysis. when children show signs of exhaustion, dysphoria, coma, increased muscle tension, irregular breathing, and increased cerebrospinal fluid pressure, toxic encephalopathy should be considered. timely administration of sedatives, oxygen and mannitol are important in order to ease cerebral edema. decrease in intracranial pressure. in the present study, any changes including children' pupillary light reflex, headache, vomiting, consciousness, breathing, pulse, temperature, and fluctuation in bp were closely observed. the head was raised - ˚ to facilitate the intracranial venous return, leading to a corresponding reduction in intracranial pressure. precaution was taken when using dehydrating agent as well as to prevent it from leaking from blood vessels, and to prevent tissue necrosis. when administering furosemide, its effect and adverse drug reactions were observed. if intracranial pressure increased, lumbar puncture was avoided as much as possible, but if it had to be carried out then rescue preparation was performed. cerebrospinal fluid flow is not be in excess to avoid the formation of hernia. control of fever and convulsions. an increase in body temperature leads to an increased oxygen consumption in brain tissue, and aggravating cerebral hypoxia, which causes cerebral edema and nerve cell damage ( ) . immediate measures must be taken to control hyperpyrexia by combining the use of artificial hibernation to decrease the temperature, lower metabolism and protect the central nervous system. the drug of choice is the application of cold compressors of chlorpromazine to the head, wearing ice caps and decreasing the temperature with drugs, thereby reducing hyperpyrexia-induced damage to brain tissue. for children patients in the study, when intracranial pressure increased, the motor cortex wass stimulated, causing convulsions and exacerbating cerebral hypoxia and brain edema, thereby aggravating intracranial pressure. the ward was kept quiet and visitors were avoided. treatment and care were performed simultaneously, as much as possible, and operating and any negative stimuli were reduced to prevent convulsions. seizures required the focus of intensive nursing, a dental pad was placed between the upper and lower teeth to prevent tongue bite, and extra gear was added to prevent any fall damage during sleep. drugs, such as phenobarbital and diazepam were administered to control fear. restless children were guarded by specially-assigned persons to prevent scratches and fall damage. heads of children in a coma were leaned to one side, to facilitate the discharge of secretions. should be given. for any child having a seizure, it was considered improper for the fright-checking agent to forcibly press the child's body, in order not to cause fractures. treatment of abdominal distention. abdominal distention often occurs in children with pneumonia, leading to children feeling uncomfortable and unrest, which hinders normal breathing. a hot water bag was placed on the abdomen, to facilitate venting. anal venting was also used and children became quiet after venting. if abdominal distention did not improve, it suggested the presence of toxic intestinal paralysis, and a poor prognosis. anti-infective therapy. after a clear diagnosis of severe pneumonia, collecting blood for bacterial culture and sensitivity test was performed prior to the use of antibiotics when conditions permitted. a separate venous access for antibiotics was established as much as possible, to rationalize the use of antimicrobial drugs and avoid side effects. in principle, the application of antimicrobial drugs was carried out on the basis of evidence-based medicine, but a majority of them are classified as empirical treatment. as the therapeutic window of children with severe pneumonia was very small, the initial drugs for treatment covered an antibacterial spectrum as broad as possible, as well as all the pathogens. there were sufficient data displaying that the inappropriate choice of anti-infective drugs for initial therapy and untimely appropriate treatment (effective therapy of antigens) had adverse consequences on the prognosis. the main indicators of efficacy were a decrease in body temperature, improvement in poisoning symptoms and ability to drink water or conduct breast-feeding or food intake. in general, - days after the body temperature became stable, the drug dosage was reduced and gradually discontinued. intravenous injection of immune globulin is considered a safe and effective method for the treatment of children with severe pneumonia. it can rapidly improve immunoglobulin g (igg) levels in the patient's blood; enhance the body's resistance to infection and immune function. by passively accepting lgg, the body acquires resistance to a variety of microbial infections. high-dose intravenous igg infusion can increase the igg concentration -to -fold in circulating blood compared to a normal person. therefore it has the ability to prevent infections. using igg infusion concurrently with antibiotics can be used to treat bacterial infections and has a broad anti-bacterial and viral spectrum, as well as a dual function of anti-bacterial antigens and viral antigens. in summary, positive and effective nursing can promote the rehabilitation of children patients, reduce the incidence of complications and children's mortality, thus play an important role in the rehabilitation of children with severe pneumonia. psychological counseling may also be strengthened. children feel a sense of inadaptation with regard to unfamiliar environment by instinct. they cried and even refused infusion in the face of examination and treatment by strangers. therefore, nurses must be careful and gentle to children patients. for older children, they were able to explain to them the importance of the infusion and blood tests for the treatment of the disease, and increase their sense of trust in the medical staff. health education is also essential, nurses should communicate more with parents and acquaint them with relevant knowledge and inform them regarding the prevention and treatment of diseases, such as that children usually need to do exercise, enhance nutrition, bask more in sunshine and engage in outdoor activities, ensure adequate sleep, pay attention to personal health, and get vaccinated for the prevention of pneumonia and influenza if necessary. global burden of childhood pneumonia and diarrhoea epidemiology and etiology of childhood pneumonia in : estimates of incidence, severe morbidity, mortality, underlying risk factors and causative pathogens for countries epidemiology and etiology of childhood pneumonia incidence and risk factors of childhood pneumonia-like episodes in biliran island, philippines -a 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community-acquired pneumonia: a survey on the attitudes of physicians in iberia and south america severe pneumonia due to cytomegalovirus in chronic obstructive pulmonary disease poor adherence to the world health organization guidelines of treatment of severe pneumonia in children at khartoum, sudan childhood anemia at high altitude: risk factors for poor outcomes in severe pneumonia zinc as an adjunct to the treatment of severe pneumonia in ecuadorian children: a randomized controlled trial severe pneumonia in intensive care: cause, diagnosis, treatment and management: a review of the literature levels of serum brain natriuretic peptide in children with congestive heart failure or with severe pneumonia two severe cases of h n pneumonia patients with immunoneuroendocrine axis dysfunction and vitamin d insufficiency severe forms of acute pneumonia with a protracted course accompanied by liver and intestinal dysfunction and dysbacteriosis in young children association of bacterial pneumonia and respiratory failure in children with community-acquired influenza infection chronic obstructive pulmonary disease severity is associated with severe pneumonia childhood very severe pneumonia and meningitis-related hospitalization and death in yemen, before and after introduction of h. influenzae type b (hib) vaccine severe pneumonia mortality in elderly patients is associated with downregulation of toll-like receptors and on monocytes hospital-acquired pneumonia is an independent predictor of poor global outcome in severe traumatic brain injury up to years after discharge the management of severe community acquired pneumonia in the intensive care unit effect of semirecumbent sleep position on severity of obstructive sleep apnea in patients with heart failure oxygen-driving and atomized mucosolvan inhalation combined with holistic nursing in the treatment of children severe bronchial pneumonia atomization inhalation of ambroxol as an auxiliary therapy for severe pneumonia in neonates high incidence of pulmonary tuberculosis in children admitted with severe pneumonia in uganda moderate-dose glucocorticoids as salvage therapy for severe pneumonia in renal transplant recipients: a single-center feasibility study noninvasive ventilation in children: a review milrinone combined with dopamine for child pneumonia complicated with heart failure experimental study of relation of fever to cerebral edema key: cord- - fu uswq authors: feldman, c.; richards, g. a. title: falciparum malaria date: - - journal: infectious diseases in critical care doi: . / - - - - _ sha: doc_id: cord_uid: fu uswq malaria is one of the most common infectious diseases in the world today, being the most important parasitic infection, and plasmodium falciparum is the organism responsible for most of the mortality [ ]. it has been estimated that approximately – million people contract malaria every year, with approximately – million deaths, most of these occurring in children [ – ]. plasmodium falciparum, mycobacterium tuberculosis and measles currently compete for the title of the single most important pathogen causing human morbidity and mortality [ , ]. infection with plasmodium falciparum has a wide variety of potential clinical consequences [ , , ]. malaria is one of the most common infectious diseases in the world today, being the most important parasitic infection, and plasmodium falciparum is the organism responsible for most of the mortality [ ] . it has been estimated that approximately - million people contract malaria every year, with approximately - million deaths, most of these occurring in children [ - ] . plasmodium falciparum, mycobacterium tuberculosis and measles currently compete for the title of the single most important pathogen causing human morbidity and mortality [ , ] . infection with plasmodium falciparum has a wide variety of potential clinical consequences [ , , ] . factors that may influence presentation include the age of the patient, their degree of immunity to the parasite and the duration of infection [ ] . in holo-or hyperendemic areas, most adults and older children are partially immune and the disease burden is mainly in children in the first few years of life [ , ] . the greatest mortality is between the ages of and years [ ] . parasitization may be almost universal in this age group and effects range from an asymptomatic infection, to a febrile illness, or even life-threatening disease [ ] . in areas of low endemicity, severe malaria occurs in both adults and children and non-immune travelers and migrant workers are also vulnerable [ ] . in adults infected with falciparum malaria for the first time, the range of clinical syndromes is wide and may include specific and multi-organ failure [ ] . malaria is a protozoal disease caused by several species of plasmodium which are spread by mosquitoes of the genus anopheles [ ] . plasmodium falciparum is one of the four species of plasmodium causing human infection. the period from inoculation to the appearance of parasitemia (prepatent period) is usually - days for p. falciparum, but may be longer, particularly in those who have been on ineffective prophylaxis [ ] . during feeding the female mosquito injects saliva, within which malaria sporozoites are carried, into the skin. within minutes the sporozoites penetrate into hepatocytes and produce tissue schizonts (also known as meronts). after - days, each tissue schizont has produced , daughter merozoites that enter the circulation, invade erythrocytes and form ring trophozoites [ ] . using hemoglobin as energy, development occurs after h into late trophozoites (larger and lacier in appearance), early blood schizonts (division begins) and then mature schizonts [ ] . the erythrocytes then lyse and merozoites are released into the circulation to invade other red blood cells [ ] . in non-immune individuals, the process is amplified -fold with each cycle. when parasitemia reaches - trophozoites per microliter, it is detectable on thick blood films, ending the prepatent period [ ] . after several cycles, some trophozoites differentiate into sex cells or gametocytes, which are infectious to the mosquito. the gametocyte of p. falciparum is characteristically banana-shaped. if male and female gametocytes are taken up by the mosquito, they mate, migrate through the midgut wall and form an oocyst which eventually leads to the release of about , sporozoites after - days [ ] . these invade the salivary glands of the mosquito to complete the cycle at the next blood meal [ ] . the time from first inoculation to first symptoms is the incubation period [ ] . its length depends on the patient's immune status and is usually - days longer than the prepatent period. in non-immune patients, symptoms may occur even before parasitemia is present. the other extreme is premunition where partial immunity is associated with asymptomatic parasitemia [ ] . malaria persists in those parts of the world where the population of anopheline mosquitoes as well as the infected human population remain above the critical density required for sustained transmission. approxi-mately % of the world's population is at risk of acquiring malaria, resulting in those - million cases annually [ ] . in africa alone approximately million cases occur every year, with a mortality of about one million. in rural africa one in children die from malaria before the age of years [ ] . compounding the problem of malaria control in the developing world is the presence of drug resistance, and resistance of mosquitoes to insecticides [ ] . malaria also afflicts individuals in southeast asia, latin america and south america. in approximately % of malarial infections were acquired in africa, % in asia, % in central america and the caribbean and % each in south america, north america, and oceania [ ] . in developed countries such as the united states, malaria largely occurs as a result of importation from other countries in the blood of immigrants, visitors, military personnel and occasionally also from importation of infected mosquitoes [ ] . recently there has been renewed interest in epidemiological aspects of falciparum malaria [ ] . it is recognized that there are clear-cut distinctions between severe and non-severe disease and between the different forms of severe disease [ ] . an important question is why plasmodium falciparum causes severe infection in some, but not all, patients. this relates partly to the age of the patient, history of prior exposure to the parasite and various aspects of the hosts' immune response [ - ] . most interestingly, it is now recognized that many of the differences in response may relate to diversity or polymorphisms in both the host and the parasite, which may impact on disease pathogenesis and be the major determinants of the outcome of a malarial infection [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . for example, in the host, possession of hemoglobin as genotype influences the risk of both cerebral malaria and severe anemia [ ] , whereas possession of certain hla genotypes (e.g., hla-b ) may be linked to resistance to these two complications [ , ] . susceptibility to both these complications is linked to polymorphism in the promoter sequence for the tumor necrosis factor (tnf) gene [ , ] , whereas susceptibility to each of these complications individually is influenced by mutations at other sites [ ] . also, different strains of p. falciparum have been shown to vary in their ability to induce production of host tnf, and this may determine the clinical severity of the infection [ ] . in addition, clonal phenotypic variation with antigenic switching is linked to, and may alter, adhesive properties of the parasite and this may be an important mechanism of immune evasion [ ] . finally, specific adhesive and linked antigenic types may be associated with severe infection [ , ] . one of the best studied genetic markers associated with p. falciparum virulence is the erythrocyte membraneprotein- family that is responsible for antigenic variation and cytoadherence of parasitized erythrocytes to endothelial and placental syncytiotrophoblast cells. parasites causing severe malaria express a small subset of these proteins that differ from those expressed by parasites causing uncomplicated infection [ ] . the salient manifestations of severe p. falciparum infection are shown in table . [ ] . there has been some debate in the literature about the definition of both severe malaria and cerebral malaria. a definition of cerebral malaria proposed by the world health organization (who) reads as follows "a clinical syndrome characterized by coma (inability to localize a painful stimulus) at least one hour after termination of a seizure or correction of hypoglycemia, detection of asexual forms of p. falciparum malaria parasites on peripheral blood smears, and exclusion of other causes of encephalopathy" [ , ] . more recently, severe malaria has been recognized to be a complex multi-system disorder and to have many of the features in common with severe sepsis or a severe inflammatory response syndrome [ , ] . studies of outcome of patients with falciparum malaria in the intensive care unit (icu) commonly report that markers of severity of illness (such as the saps or apache ii score), shock, acidosis, coma, pulmonary edema and coagulation disorders are indicators of poor outcome [ ] . the level of parasitemia has not consistently been shown to be a good predictor of outcome. metabolic acidosis has long been recognized as a major predictor of, as well as a significant contributor to, death [ , ] . whereas lactate has been considered to be the major contributor to the acidosis, unidentified anions other than lactate have been shown, more recently, to be even more important [ ] . attempts to assess severity of infection objectively have included other markers, such as the procalcitonin level, which has shown some promise [ ] . one other consideration with regard to severe infection, at least in certain areas of the world, is the potential interaction between malaria and human immunodeficiency virus (hiv) infection in those patients who are co-infected [ - ] . on the one hand it has been suggested that this may be associated with increased hiv viral replication, viral genotypic heterogeneity and cd t-lymphocyte loss leading to accelerated decline in immune function, reduced survival and increased hiv transmission [ ] . on the other hand studies have suggested that hiv infection may be significantly associated with the development of severe and complicated malaria [ ] , being associated with a high parasite burden with the associated risk that this may potentially lead to poor malaria control and a greater chance for the development of resistance to anti-malarial agents [ ] . the adhesive properties that the parasite confers on the host's erythrocytes appear to play a central role in malaria pathogenicity [ - ] . as the parasite grows in the red blood cells it induces the expression of surface ligands, as well as various endothelial receptors, that mediate adhesion to the endothelium of post-capillary venules, which results in sequestration of the parasite within the peripheral circulation [ , ] . in addition, some isolates induce expression of receptors on noninfected red cells, leading to rosette formation, and still others induce expression of adhesion molecules on other parasitized cells causing auto-agglutination [ ] . these phenomena can lead to reduced microcirculatory flow or even obstruction to local blood flow and/or cause local metabolic disturbances, such as the production of lactic acid, which may manifest as organ-specific dysfunction [ - ] . multiple endothelial receptors have been recognized (reviewed elsewhere [ , ] ), and it has now been demonstrated that endothelial activation and leukocyte sequestration in the brain appear to be a feature of fatal malaria [ , ] . a number of theories have been forwarded to more fully explain the mechanisms of cerebral malaria [ , - , - ] . initially it was assumed that it was simply a mechanical effect related to sludging of parasitized red blood cells within the vasculature, causing decreased cerebral perfusion with hypoxia [ ] . other theories have included altered microvascular permeability, secondary to malarial "toxins" or mediators such as kinins, causing cerebral edema, but this has largely been discounted [ ] . immunological mechanisms were considered following the detection of immune complexes and complement in affected brains [ , ] and still others have investigated the possibility that disseminated intravascular coagulation (dic) [ ] or endotoxemia [ ] may be involved. clarke and co-workers proposed the cytokine theory of human cerebral malaria [ - ] . they recognized that cytokines such as tnf and interleukin (il)-l when overproduced could themselves cause clinical syndromes such as those seen in human malaria [ - ] . many of these may simply be manifestations of a severe systemic inflammatory response syndrome. products of schizogony have been shown to trigger release of tnf and il-l and serum levels of these cytokines correlate with the severity of malaria infection, including the presence of cerebral symptoms [ - ] . the cytokine theory is also consistent with the concept of sequestration of parasites in the cerebral circulation in that schizogony could cause higher local levels of cytokines and their products [ ] . sequestration, however, may not be essential to the development of cerebral dysfunction according to the cytokine theory [ ] . cytokines themselves may alter cerebral function through the local generation of nitric oxide (no), which may act as a vasodilator to increase intracranial pressure and which may also function as a false neurotransmitter [ , ] . cytokines, especially ifn * , tnf, and lymphotoxins, and chemokine receptors are also said to be responsible for both blood-brain barrier alterations and biochemical changes that may also lead to parenchymal brain lesions [ ] . it is also important to consider that septic encephalopathy may be a feature of any severe critical illness and may be indistinguishable clinically from cerebral malaria [ ] . for the laboratory diagnosis of malaria, thick and thin blood smears should be made according to standard procedures [ , ] . thick smears are - times more sensitive and should be used for screening. thin smears fixed with methanol, to preserve erythrocyte morphology, and giemsa stained allow speciation as well as determination of the level of parasitemia [ , ] . various clues to distinguish falciparum malaria on thin smear include the finding of small tight rings, appliqué forms and banana-shaped gametocytes. parasitemia should be quantified and counted on thin smears as parasites per , red blood cells corrected to percentage [ , ] . after initiation of treatment, parasitemia should be followed regularly until resolution to confirm therapeutic efficacy. the time from initiation of treatment until thick smears are repeatedly negative is called the parasite clearance time. a number of newer techniques have been developed for the diagnosis of malaria including quantitative buffy coat methods, antigen detection, enzyme linked immunosorbent assay, polymerase chain reaction (pcr), including real time pcr, and indirect fluorescent antibody tests [ , - ] . if possible, patients with severe or cerebral malaria should be treated in an intensive care unit (icu) [ ] . treatment should be initiated as rapidly as possible, and should not necessarily await parasitological confirmation of the diagnosis if this is likely to be delayed [ ] . patients should be weighed in order to determine drug dosages accurately [ ] . careful fluid management is essential and may be aided by the placement of a central venous catheter and a urinary catheter. the importance of hypovolemia in severe malaria is well recognized, particularly in children, and early recognition and treatment may be associated with an improved outcome [ , ] . however, care should be taken to avoid fluid overload with the possibility of precipitating pulmonary edema. in patients with severe hemodynamic instability, non-invasive cardiac output monitoring may be of value. a recent meta-analysis of exchange transfusion as adjunctive management of severe malaria concluded that there was no evidence of an increase in survival with its use [ ] . however, the authors indicated that there were substantial problems with the comparability of the two treatment groups and suggested that only a randomized controlled trial would give definitive answers. hyperpyrexia > . °c should be treated with tepid sponging, fanning and a cooling blanket [ ] . the drug treatment of severe or complicated malaria is shown in table . [ , - , , ] . has also been reported for the quinidine salt [ ] chloroquine is only used in areas where the infection is definitively known to be sensitive to this agent. parenteral antimalarial drugs are recommended initially in most cases, at least until there is clear evidence of clinical improvement and oral medication is able to be tolerated [ , , , ] . parenteral quinine is the drug of choice in most of the tropical world. in some countries, such as in the united states, quinidine may be the drug of choice. an intravenous loading dose of either agent is recommended to achieve therapeutic levels rapidly since most deaths occur within the first h [ , , , - ] . a total of days medication is required, which may be completed with oral quinine or quinidine. artemisinin and its derivatives, although not yet licensed in many areas, appear to be exciting new agents for the treatment of severe and multidrug-resistant malaria [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . this group of drugs is being used more commonly and it has been suggested that these agents may be the drugs of choice for severe malaria because of their efficacy and safety [ - ] . artesunate is water soluble and may be given intravenously or intramuscularly, while artemeter is oil-based and is given intramuscularly [ ] . both preparations come in suppository form and may be given rectally, which has also been shown to be effective in the treatment even of severe malaria [ - ] . however, there is growing concern about the development of resistance to these agents, which is already beginning to emerge [ - ] . this, together with the fact that these agents have a short halflife, has led to the recommendation that they always be given with another agent such as mefloquine, doxycycline or clindamycin, which may be associated not only with a better and/or more rapid cure of the infection, but also limit the development of resistance [ , , , ] . combination therapy with various drugs with different modes of action is increasingly being recommended for the treatment of malaria and a number of new combinations are at different stages of development [ , ] . additional drugs that are sometimes recommended as combination therapy with quinine include antibiotics such as the macrolides (e.g., azithromycin), doxycycline or clindamycin, which should be added particularly in areas of intense quinine resistance or where prolonged treatment with quinine would otherwise be necessary [ , ] . tetracyclines are contraindicated in pregnancy and childhood. where initial response to therapy is poor, halofantrine or mefloquine have been recommended [ , ] . other agents that have been used in treatment of malaria, but which are now no longer recommended, include dexamethasone, mannitol, heparin and dextran [ , , , ] . in many parts of the world cerebral malaria is said to be the most common clinical presentation of severe malaria in man, with a mortality in the region of %, or greater, and accounting for % of deaths [ , ] . however, in some parts of africa, anemia is a more common severe manifestation particularly in children. patients fulfilling the criteria for cerebral malaria [ , , , ] manifest features of a diffuse symmetrical encephalopathy. in adults cerebral malaria tends to develop after several days of fever and other non-specific symptoms [ , ] . in children it tends to be more acute in onset, usually after less than days [ , ] . it may start dramatically with a generalized convulsion followed by persistent unconsciousness. post-ictal coma should resolve within min, but coma due to cerebral malaria usually persists more than - h. the most common neurological picture of cerebral malaria is that of a bilateral symmetrical upper motor neuron lesion with increased muscle tone and reflexes [ ] . various forms of posturing may be observed, including decerebrate and decorticate rigidity [ , ] . these may occur in hypoglycemic and normoglycemic patients. while neck rigidity and photophobia do not tend to occur, mild neck stiffness is not uncommon [ , , ] . neck retraction and opisthotonus may, however, occur in both adults and children [ ] . corneal and eyelash reflexes and papillary responses are usually nor-mal in adults, but disorders of conjugate gaze are common [ , ] . the gag reflex is usually maintained but abdominal reflexes are invariably absent. this latter may be a valuable clinical sign [ , ] . papilledema is not a feature of cerebral malaria, probably reflecting the fact that raised intracranial pressure is not found in the majority of patients early in the course of the illness [ , , ] . forcible jaw closure and bruxism are common [ , ] . studies in children recovering from cerebral malaria have shown neurological sequelae in approximately % or more of cases and these occur especially with infections that were complicated by hypoglycemia [ , ] . hemiplegia, cortical blindness, behavior disturbances, cranial nerve lesions, extrapyramidal tremor, polyneuropathy, mononeuritis multiplex, guillain-barre syndrome, and prolonged coma have all been described as neurological sequelae of patients with cerebral malaria [ , ] . the apache ii severity of illness scoring index has been used to predict the mortality of patients with cerebral malaria. in one study, a cut-off of stratified patient mortality with an accuracy of > %. in that study, high apache ii score, deep unconsciousness, acute renal failure and acidemia were identified as poor prognostic factors [ ] . convulsions may occur in as many as % of cases of cerebral malaria and are more common in children [ ] . in children it may be difficult to differentiate febrile convulsions from those due to cerebral malaria and the possibility that they may be related to hypoglycemia should also always be considered [ ] . convulsions are usually generalized, but other types, including focal seizures, may occur [ ] . generalized convulsions appear to impact negatively on the outcome. the role of prophylactic anticonvulsants in patients with cerebral malaria is under investigation. once seizures occur they should be managed in the usual way [ , ] . treatment is initially with lorazepam . mg/kg or midazolam . mg/kg intravenously, together with maintenance of the airway and appropriate cooling of the patient [ , ] . studies of generalized convulsive status epilepticus have suggested that . mg/kg lorazepam, or mg/kg phenobarbital, or diazepam . mg/ kg followed by mg/kg phenytoin, are all acceptable initial treatment regimens; however most people would treat status with lorazepam initially followed by midazolam ( . mg/kg then infusion at . mg/kg/h) or propofol ( mg/kg then infusion at µg/kg/min) if not successful [ ] . the occurrence of anemia is invariable in patients with severe falciparum malaria [ , ] . it is due to hemolysis of parasitized red blood cells, shortened survival of unparasitized cells and bone marrow dysfunction [ ] . certain red cell enzyme defects, such as g pd deficiency, may increase susceptibility to antimalarial induced oxidant-mediated hemolysis. coombs positive hemolytic anemia and microangiopathic hemolytic anemia also occur [ ] . it is recommended that patients should be transfused if the hematocrit falls below % [ ] . specific clotting factors should be administered as needed [ ] . transfusions should be carefully monitored to prevent fluid overload with its associated complications and in some patients low dose loop diuretics (e.g., furosemide mg) may be administered during the transfusion to prevent its occurrence [ ] . this was previously described as the occurrence of unusually severe intravascular hemolysis with other severe manifestations of falciparum malaria, including renal failure, hypotension and coma, despite relatively low levels of parasitemia [ , ] . the condition was attributed to some form of immunological response to quinine or one of the other anti-malarial agents, but it is also possible that it may have represented unrecognized g pd deficiency [ , ] . no special treatment of hemoglobinuria is currently recommended, although alkalinization of the urine may be desirable [ ] . some degree of renal dysfunction, as manifested by a raised serum creatinine, is common in patients with severe falciparum malaria [ , ] . this may be related to hypovolemia and blackwater fever, but more commonly occurs in association with severe malaria in which the mechanism is said to be a reduction in renal capillary blood flow [ ] . a variety of glomerular lesions have been described; however the clinical course of all three forms of renal failure is usually that of acute tubular necrosis [ , , ] . the management is similar to that of renal failure in other critical care settings [ , ] . attention should be given to fluid status, electrolytes and acid-base balance. if there is anuria or oliguria after fluid replacement, increasing intravenous doses of furosemide should be given in an attempt to increase urine output [ ] . whereas the absolute indications for dialysis are similar to those of other situations and include severe hyperkalemia, fluid overload, metabolic acidosis and uremia, continuous dialysis should be initiated early, prior to the development of fluid overload, and not be dictated by an arbitrary metabolic parameter such as creatinine [ , , ] . it has been recommended that the doses of antimalarials should be reduced in patients with renal failure [ ] . this was based on the observation of high plasma concentrations of quinine in patients with renal failure. however, this was probably due to impaired hepatic clearance as a consequence of severe infection, rather than impaired renal clearance, which has not been documented to occur even in patients with moderately elevated serum creatinine levels [ ] . a suggested dosing regimen is as follows: initial dose mg/kg of intravenous quinine dihydrochloride (salt) over h followed by mg/kg every h [ ] . the infusion volume may be reduced to - ml of % dextrose. after the second day the dose should be reduced to mg/kg eight hourly [ ] . hemodialysis removes quinine and in this situation the dose should remain mg/kg every h [ ] . there does not appear to be a need to alter the dose of chloroquine in patients in renal failure, even in those on hemodialysis [ ] . this is a particularly serious consequence of severe falciparum malaria and is often fatal [ , ] . it is similar in most respects to the acute respiratory distress syndrome (ards), and hyperparasitemia (> %) and pregnancy are important predisposing factors [ ] . the pathogenesis is not entirely clear, but as it is associated with a normal/low pulmonary capillary wedge pressure it is most likely due to an increase in pulmonary capillary permeability, as occurs with ards [ , ] . the first indication of the onset of this condition is an increase in the respiratory rate, which precedes the development of any of the other chest signs [ , ] . careful fluid management is the cornerstone of the prevention and management [ ] . hemodynamic monitoring by means of a central venous catheter or a non-invasive cardiac output monitor may aid in management of fluid status. metabolic acidosis is common in patients with pulmonary edema, although it may also occur in its absence [ ] . the mechanism is not entirely certain and it may . metabolic acidosis occur even in the absence of significant hypoxia or hypoperfusion [ ] . it appears to be due to tissue hypoxia as a consequence of stagnant flow of parasitized red blood cells through capillary beds [ ] . other factors may include impaired hepatic blood flow (a site of lactate disposal) and high cytokine levels (tnf leads to lactate production) [ ] . in addition, the malaria parasite itself produces large amounts of lactate as a by-product of glycolysis. the possible important role of unidentified anions other than lactate has been described above [ ] . the acidosis of severe bacterial sepsis appears to involve peroxynitrite induced mitochondrial dysfunction and it is possible that a similar mechanism is involved with severe malaria [ ] . therapy should include correction of hypotension and hypoxemia, if present [ ] . whether correction of an acidosis improves outcome is not known. it should only be corrected if the ph falls below . , if at all, since infusion may worsen pulmonary edema due to the large sodium load [ ] . the prognosis of patients with severe lactic acidosis is poor [ ] . progressively increasing body temperature may be associated with convulsions, delirium and coma [ ] . high fever increases metabolic demand, which may further compromise tissues damaged by stagnant capillary blood flow [ ] . heat stroke may be associated with permanent neurological sequelae [ ] . patients with cerebral malaria often improve with a decrease in the temperature, which may be achieved by tepid sponging, fanning and cooling blankets [ ] . while thrombocytopenia is very common in severe falciparum malaria, in most cases it does not appear to be an indicator of disseminated intravascular coagulation as it usually occurs in the setting of normal coagulation and without evidence of bleeding [ , ] . it appears that the previous concerns of an important pathogenic role for dic in severe falciparum malaria were exaggerated and that full blown dic with bleeding probably only occurs in % or less of patients with severe malaria [ , ] . patients with a blood parasitemia of > % were previously said to be at increased risk for the complications of severe malaria, which was said to be proportionate to the degree of parasitemia [ ] . it was further recommended that patients with hyperparasitemia should have an exchange blood transfusion [ , ] . this recommendation was not based on large prospective randomized studies, but followed individual case reports and is confounded in some cases by patients with malaria with higher levels of parasitemia who recover without transfusion and in others by the fact that total parasite burden may not be reflected in the peripheral smear [ , , ] . in addition, as described above, a recent meta-analysis of exchange transfusion in the literature, while acknowledging problems with the studies reviewed, concluded that there was no evidence, in general, of its benefit [ ] . it has also been noted that facilities for full exchange transfusions ( - units of blood) are often not widely available, although in these situations it was previously suggested that partial exchange transfusion (e.g., units) could be undertaken [ , ] . while abnormalities in "liver function tests" are quite common in patients with severe malaria, true hepatic dysfunction is uncommon and if present is mild [ ] . raised bilirubin levels are often noted and are mostly due to hemolysis [ ] . raised serum levels of aspartate aminotransaminase may also be associated with hemolysis. occasional patients with severe falciparum malaria do, however, have marked jaundice with raised serum levels of both aspartate and alanine aminotransferases in addition to prolonged prothrombin time [ ] . these patients may have true hepatic dysfunction contributed to by hemolysis and dic [ ] . hypoglycemia is a commonly reported complication in severe malaria [ , ] . it occurs in two situations in particular [ , , ] . firstly it may occur in pregnant women, where in addition to neurological sequelae it may also cause fetal distress [ , ] . unless it has been prolonged and very severe it is associated with a good prognosis and responds well to glucose administration. secondly, hypoglycemia may occur in severely ill patients and be associated with severe anemia, jaundice, hyperparasitemia, lactic acidosis and coma [ , ] . quinine-induced stimulation of insulin release may be an important mechanism, but other factors, including glucose consumption by the parasite, may be contributory [ , , ] . other mechanisms that have been considered as possible causes of hypoglycemia include depleted hepatic glycogen stores and inhibition of hepatic gluco-neogenesis [ ] . many of the usual clinical features of hypoglycemia are absent, or are masked by, or interpreted as the symptoms of malaria, but whenever the level of consciousness deteriorates in patients with malaria, hypoglycemia should be suspected [ , , ] . glucose requirements may be high and infusions of % glucose followed by - % glucose, preferably through a central venous catheter, may be required to maintain adequate blood levels [ ] . gram-negative microorganisms are frequently cultured from the blood of patients with severe malaria [ , ] . while there is often no apparent source for these organisms it is possible that they may arise via translocation through ischemic bowel. in addition these patients often have central venous and urinary catheters in place [ , ] . the manifestations of bacteremia vary from asymptomatic to severe sepsis with shock [ ] . one study has shown a high incidence of bacterial infection in patients with falciparum malaria presenting in shock [ ] . the previously described "algid malaria" is very reminiscent of gram-negative sepsis and it has been suggested that they may represent one and the same condition [ , ] . many authorities recommend both conventional antibiotics and antimalarial agents in the initial therapy of patients with severe malaria [ ] . this complication has been noted particularly in patients who have been given high-dose corticosteroids, and is thought to be due to gastric erosion [ , , ] . it should be treated in the usual way together with the infusion of fresh blood [ ] . this complication may occur in any patient with a decreased level of consciousness and is particularly common in severe malaria since these patients often vomit [ ] . the latter may be associated with convulsions or be due to anti-malarial agents. antiemetics may be given but their efficacy has not been consistently demonstrated [ ] . falciparum malaria is a particularly dangerous disease in pregnancy, especially during the second and third trimester [ ] . the mortality of cerebral malaria is approximately % in pregnant women and both mother and fetus may die despite aggressive treatment [ , ] . pregnant women are at particular risk of hypoglycemia and pulmonary edema [ , ] . the exact mechanism by which pregnancy enhances the susceptibility to, and the risk of, complicated disease is not certain. however, red cells containing mature forms of the asexual parasites are found in the placenta, being a key feature of maternal infection with p. falciparum, and are associated with significant compromise of placental function [ , ] . placental parasites express different surface ligands that facilitate immune evasion and their adhesion to specific placental molecules [ ] . treatment should start immediately and the potential teratogenic or abortifacient properties of quinine and chloroquine in this severe situation should be ignored and in any case are considered by many authorities to have been largely exaggerated [ , , , ] . blood glucose levels should be measured frequently and, where possible, fetal monitoring should be undertaken [ ] . some clinicians favor cesarian section or induction of labor if the fetus is viable [ ] . children tend to have a shorter disease course and progress much more rapidly than adults to severe malaria [ ] . hypoglycemia, seizures, severe anemia and sudden death are more common, whereas renal failure, pulmonary edema and jaundice are less likely than in adults [ , , ] . although respiratory distress does not appear in the original who definition of severe malaria, it is recognized by clinicians treating children with malaria as an important sign which is not usually due to pulmonary edema or ards [ - ] . it has also been termed the malaria hyperpneic syndrome [ ] . possible causes include cardiac failure, coexistent pneumonia, direct sequestration of parasites in the lungs, or a sign of cerebral malaria [ ] . it is important to remember that the clinical features of pneumonia and malaria, both common causes of childhood morbidity and mortality in the developing world, overlap considerably and many children fulfilling the who criteria for pneumonia may actually have malaria [ , ] . the majority of cases of respiratory distress in children are associated with lactic acidosis and this is well documented as a poor prognostic factor [ , ] . after cerebral malaria, - % of children may have neurologic sequelae of . special considerations in children which half will resolve completely [ , , ] . hypoglycemic children are at greater risk of neurologic sequelae and/or death [ ] . it is important to remember in the treatment of children with malaria that drug dosages need to be modified [ ] . the enormous cost in lives, as well as the cost of treatment, makes malaria a considerable socioeconomic burden. control of the disease through control of parasite and insect vectors has become largely ineffective due to mosquito and parasite mutation with subsequent development of resistance, together with a change in the social behavior of the host [ ] . the need for effective control measures has never been greater [ - ] . measures to achieve this should include prevention, such as insecticide impregnated bednets and mosquito repellents as well as targeted chemoprophylaxis, and provision of easy access to early treatment once infection occurs [ , , ] . measures for the future include the possibility of a vaccine which could be anti-parasite or even anti-disease, a variety of which are currently being tested [ - ] . however, a recent cochrane meta-analysis [ ] of some of the studies of currently available vaccine candidates suggested that they were not yet optimal. the possible substantial socioeconomic savings that would occur with effective vaccine use underlies the need for emphasis on immunoprophylaxis. mortality by cause for eight regions of the world: 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severe falciparum malaria: pathophysiology and electron-microscopic pathology clinical and molecular aspects of severe malaria high-level cerebellar expression of cytokines and adhesion molecules in fatal, paediatric, cerebral malaria malaria pathogenesis the pathophysiology of severe falciparum malaria function of the blood-cerebrospinal fluid barrier in human cerebral malaria: rejection of the permeability hypothesis factors contributing to the development of cerebral malaria ii. endotoxin cerebral malaria: mediators, mechanical obstruction or more? roles of tumour necrosis factor in the illness and pathology of malaria proposed link between cytokines, nitric oxide and human cerebral malaria the cytokine theory of human cerebral malaria cerebral malaria -a neurovascular pathology with many riddles still to be solved neurological complications of critical medical illnesses and transplantation the laboratory diagnosis of malaria development of a real-time pcr assay for detection of plasmodium falciparum, plasmodium vivax, and plasmodium ovale for routine clinical diagnosis evaluation of the binax now ict test versus polymerase chain reaction and microscopy for the detection of malaria in returned travelers performance of the now malaria rapid diagnostic test with returned travellers: a -year retrospective study in a french teaching hospital fluid management of severe falciparum malaria in african children pre-transfusion management of children with severe malarial anaemia: a randomized controlled trial of intravascular volume expansion exchange transfusion as an adjunct therapy in severe plasmodium falciparum malaria: a meta-analysis diagnosis and treatment of malaria in britain current concepts: the treatment of malaria high first dose quinine regimen for treating severe malaria. the cochrane database of systematic reviews averting a malaria disaster artemether in severe malaria -still too many deaths a controlled trial of artemether or quinine in vietnamese adults with severe falciparum malaria q -arteether for the treatment of complicated falciparum malaria a trial of artemether or quinine in children with cerebral malaria randomized comparison of artemether-benflumetol and artesunate-mefloquine in treatment of multidrug-resistant falciparum malaria descriptive study on the efficacy and safety of artesunate suppository in combination with other antimalarials in the treatment of severe malaria in sudan rectal artemether versus intravenous quinine for the treatment of cerebral malaria in children in uganda: randomized clinical trial artemisinin-based combinations treating severe malaria south east asian quinine artesunate malaria trial (sea-quamat) group ( ) artesunate versus quinine for treatment of severe falciparum malaria: a randomized trial artemisinin derivatives for treating severe malaria. the cochrane database of systematic reviews a comparative clinical trial of artemether and quinine in children with severe malaria comparison of intramuscular artemether and intravenous quinine in the treatment of sudanese children with severe falciparum malaria randomized comparison of artesunate and quinine in the treatment of severe falciparum malaria clinical experience with intravenous quinine, intramuscular artemeter and intravenous artesunate for the treatment of severe malaria in thailand randomized control trial of quinine and artesunate in complicated malaria artemisinins association of failures of seven-day courses of artesunate in a nonimmune population in bangui, central african republic with decreased sensitivity of plasmodium falciparum resistance of plasmodium falciparum field isolates to in-vitro artemether and point mutations of the serca-type pfatpase are we losing artemisinin combination therapy already? falciparum malaria: current therapeutic challenges dexamethasone proves deleterious in cerebral malaria. a double-blind trial in comatose patients high dose dexamethasone in quininetreated patients with cerebral malaria: a double-blind, placebo-controlled trial do patients with cerebral malaria have cerebral oedema? a computed tomography study apache ii scoring for predicting outcome in cerebral malaria the management of status epilepticus acute renal failure in patients with severe falciparum malaria treatment of malarial acute renal failure by hemodialysis fluid overload in multiple organ dysfunction syndrome: prediction of survival effects of different doses in continuous veno-venous hemofiltration on outcomes of acute renal failure: a prospective randomized trial association between mitochondrial dysfunction and severity and outcome of septic shock editorial response: exchange blood transfusion in severe falciparum malaria -the debate goes on severe hypoglycemia and hyperinsulinemia in falciparum malaria shock complicating severe falciparum malaria in european adults the immunology and pathogenesis of malaria during pregnancy quinine therapy in severe plasmodium falciparum malaria during pregnancy in sudan pathophysiology of fatal falciparum malaria in african children indicators of life-threatening malaria in african children the pathogenesis of severe malaria in african children overlap in the clinical features of pneumonia and malaria in african children severe malaria in children in papua new guinea neurological sequelae of cerebral malaria in children what's new in malaria control? current status of malaria control chemoprophylaxis of malaria the malaria vaccine: anti-parasite or anti-disease? immunizing against toxic malarial antigens malaria vaccines development of a malaria vaccine evaluation of spf malaria vaccine efficacy in brazil for the rts, s malaria vaccine evaluation group ( ) a preliminary evaluation of a recombinant circumsporozoite protein vaccine against plasmodium falciparum malaria malaria vaccines evaluation and implementation a novel method for development of malaria vaccines using full-length cdna libraries malaria vaccine developments vaccines for preventing malaria. the cochrane database of systematic reviews key: cord- - bk t authors: gülsen, askin; wedi, bettina; jappe, uta title: hypersensitivity reactions to biologics (part i): allergy as an important differential diagnosis in complex immune-derived adverse events* date: - - journal: allergo j doi: . /s - - - sha: doc_id: cord_uid: bk t purpose: biotechnological substances (bss) are strongly relied upon to prevent rejection of transplanted organs, and to treat oncological, allergological, and other inflammatory diseases. allergic reactions to partly foreign biologics can occur due to their potential immunogenicity. the severity of an immune response to a biological drug may range from no clinical significance to a severe, life-threatening anaphylactic reaction. methods: detailed searches were performed on pubmed, web of science, and google scholar to include all available publications. in addition, the food and drug administration, the european medicines agency, and british columbia cancer agency drug manual databases were screened for hypersensitivity reaction (hsr), infusion reaction, injection site reaction, urticaria, and anaphylaxis for individual bss. results: treatment with bss can cause various types of hsr. these are mentioned in the literature with definitions such as allergic reactions, anaphylactoid reactions, anaphylaxis, hsr, infusion reactions, injection site reactions, cytokine release syndrome, and urticaria. due to the overlap in signs and symptoms in the reported descriptions, it is not always possible to differentiate these reactions properly according to their pathomechanism. similarly, many data reported as anaphylaxis actually describe severe anaphylactic reactions (grades iii or iv). conclusion: there is an urgent need for a simpler symptom- or system-based classification and scoring system to create an awareness for hsrs to bss. a better understanding of the pathophysiology of hsrs and increased clinical experience in the treatment of side effects will provide timely control of unexpected reactions. as a result, immunotherapy with bss will become safer in the future. cite this as gülsen a, wedi b, jappe u. hypersensitivity reactions to biologics (part i): allergy as an important differential diagnosis in complex immune-derived adverse events. allergo j int ; : - https://doi.org/ . /s - - - biotechnological substances (bss), also known as biological agents, biologicals or biologics, are pro-duced by living organisms or are synthesized from a product made by a living organism. some bss are directed against specific cytokines or cell surface signal receptors. in recent years, the bs industry has experienced rapid growth and yielded products for treatment of a broad spectrum of diseases and dis- orders. biologics are strongly relied upon to prevent rejection after organ transplantation and to treat diseases related to allergology, pneumology, rheumatology, oncology, and dermatology. bss have revolutionized the treatment of diseases such as rheumatoid arthritis, psoriatic arthritis, inflammatory bowel disease, and organ-specific cancers (lung, stomach, colon, cervix, breast, etc.). other conditions commonly treated or managed with biologics include several types of asthma, plaque psoriasis, chronic urticaria, and vasculitis. some new indications and less studied applications include interstitial lung disease, unstable angina pectoris during cardiac catheterization, paroxysmal nocturnal hemoglobinuria, neonatal bronchopulmonary dysplasia, and multiple sclerosis (tab. ). allergic reactions to partly foreign biologics can occur due to their potential immunogenicity. adverse drug reactions (adrs), however, can also result from an agent's direct biological function. for instance, neutralization of an off-target cytokine's activity can cause adrs [ ] . the severity of an immune response to a biological drug may range from no clinical significance to a severe, life-threatening anaphylactic reaction [ ] . thus it became evident that the nomenclature used to describe and document immunological adverse events was not always precise enough, or the cases had not been investigated in enough detail to clearly differentiate between truly allergic and other reactions. this review will evaluate reports of allergic and substance-specific infusion reactions (ir), injection-site reactions (isr), hypersensitivity reactions (hsr), urticaria, and anaphylaxis caused by bss. the most common indications for the use of biologics in lung diseases are allergic and severe uncontrolled asthma. biologics are used more rarely in interstitial lung diseases (tab. ). their use in lung cancer will be mentioned in the section oncology. benralizumab, lebrikizumab, mepolizumab, reslizumab, and omalizumab are used in asthmatic patients. the primary indication for nintedanib and pirfenidone is idiopathic pulmonary fibrosis. benralizumab is the newest anti-asthmatic and anti-inflammatory drug in the monoclonal antibody (mab) family developed for the treatment of severe eosinophilic and allergic asthma. it exerts its effects by binding to the α subunit of the interleukin (il)- receptor on eosinophils and basophils. in a phase iib study conducted between and , the isr rate was % and anaphylaxis was % [ ] . fda labels show a % incidence of hsrs (ana-phylaxis, urticaria, rash, and angioedema) in the treatment group receiving benralizumab and placebo [ ] . the frequency of isr (erythema, local pain, papule, or pruritus) was . % in patients treated with benralizumab [ ] . these reactions usually occurred within the first hours after s. c. administration of the drug. the most reported adr was upper respiratory tract infection and mild to moderate nasopharyngitis [ ] . in a recent study in which patients were treated, hsr and anaphylaxis were not reported; only slight fever with chills was observed in two patients [ ] . in the meta-analysis of eight randomized controlled trials, the overall risk of adrs and severe side effects was lower and the risk of headache and pyrexia higher in patients treated with benralizumab compared to placebo [ ] . in addition, an increased incidence of isrs, hypersensitivity, and death was not observed compared to placebo. the isrs were reported in . % of patients treated with mg benralizumab. post-marketing side effect recording is ongoing. lebrikizumab is a novel humanized mab that specifically inhibits the activity of il- . clinical stud- ies for the treatment of asthma that is uncontrolled by inhaled corticosteroids, chronic obstructive pulmonary disease (copd), atopic dermatitis, and idiopathic pulmonary fibrosis (ipf) are ongoing. in one placebo-controlled study using higher doses, isrs (pain, erythema) were found to be more prevalent, . % and . % in groups receiving and mg doses of lebrikizumab, . % in a group receiving . mg, and % in the group given placebo [ ] . in the phase iii study (lavolta) of lebrikizumab for asthma, isrs occurred in % of participants receiving . mg s.c. injections, % of participants receiving mg s.c. injections, and % of participants receiving a placebo once every weeks [ ] . in the same study, anaphylaxis was reported to be below %. korenblat et al. reported anaphylaxis in one patient ( %) treated with lebrikizumab, and this reaction was attributed to a known peanut allergy [ ] . in another study, lebrikizumab was well tolerated, and death and anaphylactic reactions were not reported [ ] . isrs rarely occurred ( . % in the lebrikizumab and . % in the placebo group). mepolizumab is a comparatively new anti-inflammatory, anti-asthmatic bs consisting of a mab that binds to il . therefore, it was also approved by the fda in december for use in the treatment of churg-strauss syndrome (eosinophilic granulomatosis with polyangiitis). in a multicenter study covering the years - (dream study), the most commonly reported drug-related adverse events were non-allergic, infusion-related reactions [ ] . infusion-related reactions occurred in % of participants receiving mg of mepolizumab, % of participants receiving mg, % of participants receiving mg, and % of participants receiving placebo. hsrs thought to be associated with the study drug occurred in % of participants receiving mg mepolizumab, < % of participants receiving mg, % of participants receiving mg, and % of participants receiving placebo. no anaphylactic reaction was reported. anaphylaxis has not been reported on fda labels [ ] . however, in one case, type-iv delayed hsr was observed in the th month of treatment, days after subcutaneous (sc) administration of mg mepolizumab. local isrs ( % after mg application, % after mg application, % after placebo), and only few allergic reactions (≤ %) have been reported. lugogo et al. [ ] conducted a study on sc administration of mepolizumab in severe eosinophilic asthma patients. in this study, hsrs (type iv) occurred in ≤ % of participants, local isrs occurred in %, and < % of participants experienced injection-related reactions (non-allergic). there was no report of mepolizumab-related anaphylaxis. finally, in a recent study of severe eosinophilic asthma patients, local isrs ( %), allergic/hsrs ( %), and non-allergic systemic reactions (< %) were reported [ ] . anaphylaxis was not observed. reslizumab is a biologically active humanized mab used for adjunctive therapy of severe asthma with an eosinophilic phenotype. this drug binds and inhibits the cytokine il , which is responsible for growth, differentiation, recruitment, activation, and survival of eosinophils. in a study conducted by castro et al. [ ] in , isrs (hematoma, rash, and local pain) were reported to be less than %, and an anaphylactic reaction was reported in two patients. anti-drug antibodies (ada) for reslizumab were found to be negative in these patients, and therefore no further information could be obtained. in fda labels, treatment-related anaphylaxis was reported in three patients in the reslizumab mg/kg treatment group [ ] . the same label states that this drug is produced in the murine cell line, which introduces the galactose α- , -galactose (α-gal) carbohydrate sequence as a post-translational modification into the carbohydrate side chains of the mab. this explains why this drug is immunogenic for humans, but it is unclear how α-gal acts and which role it plays in these reactions. ada were positive in approximately % of patients treated with reslizumab, but there was no correlation with hsr and allergic reactions. murphy and colleagues [ ] investigated the longterm effects of reslizumab and reported two hsrs (< %), two drug eruptions (< %), and very rare local infusion-related aes (e.g., pain at injection site) (< %). they did not report anaphylaxis during the follow-up period. in a recent study, one case of toxicoderma (the th dose of the drug h after administration) was reported [ ] . the patient developed a symmetrical rash on the trunk and proximal limbs, and the symptoms improved after administration of systemic corticosteroids and antihistamines. omalizumab is a humanized mab commonly used as an adjunctive and second-line treatment for severe allergic asthma and chronic spontaneous urticaria. the effect is due to selective binding to immuno globulin e (ige). the task force report published by the american allergy academy in states that the rate of ana-phylaxis associated with omalizumab treatment is . % [ ] . of the reactions observed to date, % occurred within h after one of the first three doses, and % occurred within min after the fourth or a later dose. omalizumab is produced in the ovarian cells of a chinese hamster and does not express the α- , -galactosyl-transferase, and therefore does not contain α-gal. as a consequence, pre-existing ige-antibodies as have been described in patients with anaphylaxis to cetuximab were not detectable in humans [ ] . in a large-scale real life study conducted between and , isrs, immediate local reactions, urticaria, and anaphylaxis were reported to be . %, . %, . %, and %, respectively [ ] . in , a sequel of the task force report of the american allergy academy written in was published on the basis of post-marketing safety data provided by genentech/novartis [ ] . in this additional report, a total of of the patients ( . %) that developed anaphylaxis against omalizumab reported that these reactions occurred within the recommended hospital waiting period (especially within the first h after the first three injections). in fda labels, anaphylaxis with symptoms such as angioedema, bronchospasm, urticaria, hypotension, and syncope has been reported in . % of patients [ ] . this can be seen after the first dose, as well as year later. isrs of any severity (pain, burning, induration, warmth, redness, bruising, itching, granuloma formation, etc.) were observed in % of patients compared with placebo ( %). more severe reactions have been reported in % of patients receiving omalizumab and % in placebo. most of these reactions occur h after injection and last for less than days. nintedanib is an antiproliferative and antitumoral oral bs of the multikinase receptor inhibitor group usually used in the treatment of idiopathic pulmonary fibrosis and non-small cell lung cancer. the australian public assessment report and the fda label did not observe an increase in the incidence of severe immunological and anaphylactic reactions related to the use of nintedanib [ , ] . both reports documented hepatotoxicity, bleeding and thromboembolic events, gastrointestinal toxicity (diarrhea, nausea, and vomiting) and perforation, myocardial infarction, and hypertension as side effects. in post-marketing studies, it is reported that pruritus and rash may be seen, but the rate is not clear (referenced in [ ] ). nintedanib capsules contain the following excipients; (i) capsule content: triglycerides, hard fat, lecithin (soya); (ii) capsule shell: gelatin, glycerol, titanium dioxide, iron oxide red and yellow; (iii) printing ink: shellac glaze, iron oxide black, and propylene glycol [ ] . in addition, the summary of product characterics indicate that those patients with soy and peanut allergy should be treated with caution, but more detailed information as to the reason for legume allergy to be considered as a risk is lacking. insufficient data were found in our literature review to assess the prevalence of allergic reactions, hsr, anaphylaxis, and urticaria due to the use of this bs. this is probably due to the fact that other side effects were considered as having higher priority. pirfenidone is an oral bs with antifibrotic and antiinflammatory properties. its only indication is the treatment of mild to moderate idiopathic pulmonary fibrosis. it exerts its effect by inhibiting transforming growth factor (tgf)-β . skin rash was reported in % of patients treated with pirfenidone and in % of patients treated with placebo [ ] . in addition, phototoxic burn-like skin rashes on sun-exposed body areas and erythematous (edematous or non-edematous) lesions were reported in % of patients and in % with placebo. in newly published fda labels, photosensitivity and rash were reported at a rate of %, but hsr and anaphylaxis were not mentioned in this report [ ] . indications for which bss are developed in dermatology include moderate to severe psoriasis, chronic urticaria, and atopic dermatitis (tab. ). currently prescribed bss include alefacept, efalizumab, ixekizumab, secukinumab, ustekinumab, dupilumab, quilizumab, ligelizumab, and omalizumab. tnf-α inhibitors such as etanercept, infliximab, and adalimumab have also been approved by the fda for treatment of moderate to severe psoriasis and psoriatic arthritis [ ] . off-label indications for tnf-α inhibitors include autoimmune bullous disease, pemphigus vulgaris, and pyoderma gangrenosum [ ] . rarer indications include connective tissue disorders such as scleroderma, dermatomyositis, systemic lupus erythematosus, sweet's syndrome, sarcoidosis, granuloma annulare, toxic epidermal necrolysis, pityriasis rubra pilaris, and behcet's disease [ ] . bss used in the treatment of psoriatic arthritis will be mentioned in the section rheumatology. alefacept is a fully human recombinant lymphocyte function-associated antigen- (lfa- ) immunoglobulin g fusion protein with a dual action mechanism that targets t cells, and can be administered intramuscularly or intravenously on a weekly basis. its primary function is to interact with cd in the membrane of cd + and cd + t cells, inhibiting activation and thus regulating cd /lfa interaction. a secondary mechanism of action is the induction of apoptosis in memoryeffect t lymphocytes. according to fda labels, four out of , patients ( . %) reported angioedema in clinical trials: two of these patients were hospitalized and treated [ ] . however, urticaria was seen in six patients (< %) during the -week period. in one patient, therapy needed to be terminated due to allergic reactions. isrs were reported in % of patients receiving alefacept by intramuscular administration, compared with % of patients treated with placebo. repeated doses did not change the rate of incidence. isrs were usually mild and were reported to manifest as pain ( %), inflammation ( %), bleeding ( %), edema ( %), local granuloma ( %), and nonspecific reaction or skin hypersensitivity (< . %). it has been reported that approximately % of patients developed low titer antibodies to the fusion protein, but a long-term effect was not known. fda approval was withdrawn in september after a decision was made by the manufacturer to stop production of the drug, and the drug was never approved for the european market. there is no information about the frequency of anaphylaxis in the literature. efalizumab is a humanized mab that binds to the cd a subunit of lymphocyte function-associated antigen (lfa- ) on the surface of lymphocytes. it is used to treat adult patients with moderate to severe psoriasis. in a randomized controlled study involving patients conducted by gordon et al., % of patients developed antibodies to efalizumab, but no anaphylaxis was observed [ ] . in controlled clinical trials involving , patients, % experienced at least one hsr (e.g., asthma, dyspnea, angioedema, urticaria, maculopapular rash) and % experienced urticaria. other reported side effects include angioedema, laryngospasm, erythema multiforme, serum sickness-like reaction and allergic drug eruption. a low concentration of antibodies to protein components of the drug have been reported in . % of patients (referenced in [ ] ). in a retrospective cohort study conducted by brunasso et al., isrs (local erythema, itching, burn-ing, pain, edema, or urticaria) were observed in % of participants [ ] . ixekizumab is an immunosuppressive and anti-inflammatory mab used to treat moderate to severe plaque psoriasis and can be administered subcutaneously. its mechanism of action involves binding of the cytokine il- a, an important factor in keratinocyte activation and proliferation. according to fda labels, isrs were reported in % of patients. the most common type of isrs were erythema and pain [ ] . less frequently, serious hsrs such as angioedema and urticaria (≤ . % each) have also occurred. in the post-marketing period, rare events of anaphylaxis requiring hospitalization have been reported. in a long-term clinical investigation by strober and colleagues [ ] following , patients, no anaphylactic reactions were reported. in this study, isr, local erythema, and drug hypersensitivity were reported in . %, . %, and . % of patient-years, respectively. secukinumab is a mab that is administered subcutaneously to treat plaque psoriasis, ankylosing spondylitis, and psoriatic arthritis. its anti-inflammatory and immunomodulatory properties result from its binding to and inactivation of il- a. in the assessment report of the ema in , no cases of anaphylaxis or angioedema were reported [ ] . urticaria was rarely (< . %) observed. injection-site pain was reported in . % of cases, and general hsr occurred in . - . %. the most common hsrs in any secukinumab group were reported to be dermatitis ( . %), eczema ( . %), and rash ( . %). less than % of patients developed antibodies during sekukinumab treatment for up to weeks. in one study, erythema and itching at the injection site were reported in . % of participants and anaphylaxis in . % [ ] . according to the fda label, urticaria was reported in . - . % of patients depending on the dose, and less than % of patients developed ada [ ] . although this report states that anaphylactic and allergic reactions are rarely seen, the rate was not given. in a recent study, hsr was reported to be . % per patient-years and isr in . - . %; in addition, adas were reported in < % of patients receiving treatment for weeks [ ] . ustekinumab is a mab that acts as an interleukin inhibitor and was approved for administration at month intervals as a second-line treatment for patients with moderate to severe plaque psoriasis. it has immunosuppressant and anti-inflammatory effects and neutralizes il- and il- . in the ema evaluation report for , isrs were reported to occur in . %. intravenous infusion was not associated with anaphylaxis, ir, or serum sickness-like reactions [ ] . fda labels show a - % incidence of isrs (bruising, hemorrhage, induration, irritation, pain, pruritus, and swelling) [ ] . in addition, one patient ( . %) experienced signs and symptoms (flushing, tightness of the throat, shortness of breath) consistent with anaphylaxis after initial sc injection, and one patient ( . %) experienced signs and symptoms consistent with or related to an hsr ( urticaria, flushing, chest discomfort, and increased body temperature) after initial intravenous injection. in addition, approximately - . % of patients developed adas, which were generally of low titer (referenced in [ ] ). a recent study examined the side effects of ustekinumab in psoriasis, psoriatic arthritis, and crohn's disease [ ] . no serum sickness-like reactions or serious anaphylactic events were reported. two patients (< . %) had temporary signs of treatment-associated hypersensitivity. the patients developed flushing, shortness of breath, and throat tightness after the first sc administration. in addition, fever, flushing, chest discomfort, and urticaria were observed after the initial intravenous administration. long-term safety and side effect studies are ongoing (c z [psolar] and cnto pso [nordic database initiative]). dupilumab is a relatively new bs in the mab group, with anti-inflammatory and selective immunosuppressive properties. it is used as second-line therapy to treat moderate to severe atopic dermatitis. it exerts its effect by binding the alpha subunit of the il- receptor, eliminating the biological effects of il- and il- . since additional indications are severe uncontrolled asthma with type inflammation and chronic rhinosinusitis with nasal polyposis. in the fda drug report, hypersensitivity reactions such as serum sickness, serum sickness-like reaction, and generalized urticaria occurred with a frequency of < %, and isrs were observed in % of cases [ ] . ada were detected in - % of treated patients, and two subjects developed serum sickness or serum sickness-like reactions. in these patients, a high antibody titer for dupilumab was reported during treatment. there is no information about anaphylaxis. in a meta-analysis conducted by ou et al., isrs were reported in . % of cases [ ] . there is no mention of anaphylaxis, hsr, or urticaria, but it is stated that the high incidence of isrs needs to be explained. the ema report shows an hsr in . - . % of adult patients and isrs in . - . % (referenced in [ ] ). severe isrs were reported in . - . %. most isrs were mild to moderate. the other most commonly reported adverse events were urticaria ( . - . %) and rash ( . - . %), depending on the dose, but with a similar incidence to placebo. anaphylactic reaction or angioedema was reported in three patients ( . %), hypersensitivity in two patients ( . %), drug hypersensitivity in two patients ( . %), and anaphylactic shock in one patient (< . %). ligelizumab is a newly developed humanized mab against human ige and belongs to the igg /κ isotype subclass. this bs has -fold greater affinity to human ige compared with omalizumab. a recent phase ii clinical trial demonstrated that a higher percentage of patients had complete control of symptoms of chronic spontaneous urticaria with ligelizumab therapy of mg or mg than with omalizumab or placebo [ ] . in the -mg treatment group in this study, mild injection site erythema was observed in . % and mild to moderate isrs in . % of patients, while in the -mg-treated group, these were reported in . % and . %, respectively. in addition, serious side effects were reported in % of patients in both treatment groups, but no mortality or anaphylaxis occurred. phase iii studies are ongoing and expected to be finalized by . quilizumab is a newly developed humanized mab against the m -prime segment of membrane-expressed ige. it leads to depletion of ige-producing and memory b cells, thus blocking ige production. its primary indications are chronic spontaneous urticaria and uncontrolled allergic asthma. in one study assessing efficacy and safety, . % of patients reported isrs (most often local pain) [ ] . further studies were discontinued due to the lack of efficacy in adults. the use of bss in oncology is quite extensive. the literature shows that these drugs are used for the treatment of bladder, breast, cervical, head and neck, gastrointestinal (stomach, bowel, colorectal), lung, and kidney cancers, as well as leukemia and lymphoma. details of the reactions are given in tab. . aflibercept is a human recombinant fusion protein targeting vascular endothelial growth factor (vegf- ) receptors used in colorectal carcinoma with metastasis, as well as retinal diseases such as macular degeneration. in the ema assessment report and fda label, it was stated that hsrs were observed at a rate of . % (placebo . %) [ , ] . severe hsrs such as anaphylaxis, angioedema, bronchospasm, and dyspnea have been described as uncommon (≥ / to < / ) [ ] . however, more detailed information has not been provided. in addition, ada have been reported to develop in . - . % of patients, and the clinical significance of their neutrali zation has not been assessed due to limited data [ ] . maculopapular rash may develop after intravitrael administration of this drug in retinal diseases [ ] . alemtuzumab is a monoclonal igg kappa antibody and a cytolytic agent used intravenously to treat leukemia and multiple sclerosis. this drug binds the glycoprotein cd on the surface of lymphocytes and other immune cells and leads to cell death. in the ema assessment report, allergic side effects were reported with relatively high frequencies (rash in %, urticaria in %, and pruritus in . % of patients) [ ] . serious and severe urticaria was seen in . % of cases. severe infusion-associated reactions, including pyrexia, urticaria, atrial fibrillation, nausea, chest discomfort, hypotension, and grade iv anaphylaxis were reported in . - . % of cases. the british columbia cancer agency (bcca) drug manual reported anaphylactic reactions and angioedema in < %, isrs in . % (severe in %) of intravenous users and isrs in % (severe in %) of sc users. reactions occurred between h after the first dose and the administration of the fourth dose. in addition, sc injection site pain was reported in . % (severe in . %). pruritus and urticaria were reported in - % ( - % of these reactions were severe; % of the reactions was associated with sc use) (referenced in [ ] ). in the last published fda label, irs and cytokine release syndrome were reported in % of treated patients, but epinephrine or atropine was administered in . % of these patients [ ] . in some patients, infusion reactions occurred h after infusion. symptoms include fatigue, dyspepsia, pruritus, flushing, urticaria, dyspnea, pain, nausea, dizziness, and pulmonary infiltrates. severe reactions occurred in approximately % of patients. two patients experienced anaphylaxis (including anaphylactic shock). symptoms include headache, rash, chest pain, angioedema, bradycardia, tachycardia (including atrial fibrillation), bronchospasm, hypotension, hypertension, transient neurological symptoms, and pyrexia. the most common side effects were rash ( %), urti-caria ( %), pruritus ( %), and flushing ( %). antialemtuzumab antibodies were detected in %, %, %, and % of the patients at , , , and months of treatment, respectively. in a recent study that evaluated efficacy and safety, . % of patients had irs such as fever, shivers, rash, and headache [ ] . atezolizumab is an igg class humanized antibody that acts by binding to programmed death ligand (pd-l ) used to treat bladder, breast, and lung cancer. pd-l is the immune checkpoint protein, which is expressed by tumor cells and inhibits the antitumor function of the t cell. inhibition of the ligand's interaction with its receptor by bs enhances the anti-tumoral activity of t cells and improves the immune system of the patients. according to the fda label, severe irs were observed in . - . % of patients [ ] . these reactions include the following symptoms: back or neck pain, chills or shaking, dizziness, feeling like passing out, fever, flushing, itching or rash, dyspnea or wheezing, and swelling of face or lips. however, immune-related adverse reactions (such as colitis, hepatitis, hypophysitis, meningoencephalitis, pancreatitis, and pneumonitis) have been reported that affect various organs. in the ema assessment report, hsr such as anaphylaxis and irs is reported to be seen in up to % of patients [ ] . in addition, itching of the skin and rash are reported to be seen in more than % of patients. according to the recent bcca drug manual, it was reported that hsr including anaphylaxis can develop in ≤ % (severe < %), irs in % (severe ≤ %), and immune-mediated rash in - % ( severe ≤ %) of patients [ ] . bevacizumab is a mab used to treat various cancers including kidney, stomach, colon, breast, and lung cancer. the antibody inhibits binding of vegf to its receptors, reducing angiogenesis (blood vessel formation) and tumor growth. according to the bcca drug manual, irs/hsrs with symptoms (dyspnea, redness, rash, hypo-or hypertension, oxygen desaturation, chest pain, tremor, and nausea/vomiting) were reported in up to % of patients (referenced in [ ] ). serious hypersensitivity reactions involving anaphylactic and anaphylactoid responses have occurred, but no rates have been reported. in addition, rash was reported in %, rhinorrhoea in %, and flushing in % of patients. in the ema assessment report, irs occurred in . - . % of cases, and the most commonly reported [ ] ). in this report, hsrs and anaphylaxis were described as treatment-emergent adverse events (teae) and were reported to be any grade of teaes in . %, grade or higher teaes in . %, and serious teaes in . % of cases. further information on reactions is not available. ir and symptoms (chest pain, chills, grade hyper sensitivity, diaphoresis, headache, hypertensive crises associated with hypertension, hypoxia, neurological signs and symptoms, and wheezing) have been reported in clinical trials and post-marketing studies, as indicated on the fda label [ ] . any grade of irs are reported to be rare (< %), and serious irs have occurred in . % of the patients. symptoms include chest pain, diaphoresis, hypertensive crises, hypoxia, rigors, and wheezing. although urticaria and anaphylaxis are not mentioned in this label, exfoliative rash was seen in % with the administration of this bs in combination with the chemotherapy protocol. blinatumomab is a bispecific mab used for cancer immunotherapy and specific forms of acute lymphoblastic leukemia (all). it exerts its effect in part by simultaneously binding cd on b cells and cd on t cells. this bs can lead to cytokine release syndrome (crs) in - % of patients, with % of these cases being severe, to hypersensitivity in % of patients, and infusion reactions in % of patients, according to the bcca's report (referenced in [ ] ). symptoms of this syndrome include asthenia, nausea, pyrexia, headache, hypotension, and increased liver enzymes. it was reported that crs peaked within the first days of treatment and was not easily differen-tiated from ir. although anaphylaxis and urticaria were not mentioned in this report, the incidence of rash was reported to be - % (severe - %). according to the ema assessment report, crs occurred in . - . % of patients, rash occurred in . %, and irs occurred in . % (grade ≥ in . %). in total, severe irs were very rarely seen ( . %) [ ] . fatal irs were not reported. additionally, maculopapular rash and flushing have been reported in . % of patients. according to the recent fda label, this bs caused crs in - % of patients ( % of these instances were severe), skin rash in % of patients (< % of these instances were severe) and irs in % of patients ( % of these instances were severe) [ ] . symptoms of crs included asthenia, headache, nausea, fever, hypotension, increased liver enzymes, increased total bilirubin, and diffuse intravascular coagulation. irs usually occurred during the first h of infusion and lasted less than days. clinical findings included hypotension, hypertension, rash, generalized pruritus, pyrexia, periorbital edema, and cytokine release syndrome. events defined as rash included maculopapular rash, erythema, contact dermatitis, and eczema. cetuximab is an intravenously administered epidermal growth factor receptor (egfr) inhibitor used to treat advanced colorectal cancer, head and neck squamous cell carcinoma, pancreatic cancer, and less frequently, non-small cell lung cancer. this antibody can reduce cancer cell survival, neovascularization, cell migration, and metastasis. cetuximab was the first anti-egfr mab and bs approved for cancer treatment; therefore, its immunological and allergic side effects are well studied. in a review of acute infusion reactions induced by mabs, acute irs with cetuximab were in the range of . - % (referenced in [ ] ). it is noted in that particular review that the pathogenic mechanism of mab-related irs has not yet been fully defined, with the exception of several suggested mechanisms by w. j. pichler [ ] . hsrs have been reported in . % of participants in a safety study with patients, with . % of reported hsrs being of grade severity, and . % being of grade anaphylactic reactions [ ] . the majority of reactions occurred after the first dose, and all grade reactions occurred within minutes following the first infusion. a review reported hsr to be in the . - . % range, and infusion reactions in the . - . % range in patients treated with cetuximab (referenced in [ ] ). according to the fda label, mild irs (fever, chills, shortness of breath, bronchospasm, angioedema, urticaria, hypertension, and hypotension) occurred in . % of patients, and severe irs occurred in . % of patients, with % of events occurring after the first infusion [ ] . in addition, % of patients developed acneiform rash during treatment, . % of which were severe. anaphylactic reactions during treatment have been reported to be increased in association with tick bites by amblyomma americanum, a history of red meat allergy, all associated with ige-antibodies against α gal [ , , ] . in , reactions during the first infusion of cetuximab and delayed type anaphylactic reactions to red meat were investigated in detail [ ] . in both diseases, patients were shown to have increased ige antibodies specific for α-gal, a mammalian disaccharide, not expressed in humans but on the tissue of mammalians. in most patients with an hsr and anaphylaxis, ige antibodies to cetuximab were also present in serum before treatment, which prompted the investigations into the sensitization route [ ] . it was shown that α-gal is also the main cause of delayed anaphylaxis against red meat and that this was associated with the occurrence of the american tick amblyomma americanum [ ] . presently, tick bites are thought to be the predominant sensitization route for antiα-gal ige induction. α-gal, however, has another implication for drug-induced hypersensitivity reactions: it is present in mammalian gelatine and, therefore, may be part of vaccines, gelatine-containing infusion solutions, ovula, capsules and pills, suppositories, snake venom antisera, and animal-derived heart valves (referenced in [ ] ). however, in a study involving patients, ( . %) reported hsr, eight of these ( . %) reported severe hsr [ ] . anti-cetuximab ige levels were detected in seven of the eight patients with severe hsr. durvalumab is a pd-l binding and blocking mab used in non-small cell lung carcinoma and metastatic urothelial carcinoma. in the fda labels, irs were reported to occur in . % (grade in . %), and immune-mediated rash in - . % (grade - in . - . %) of patients [ ] . ada developed in . % of patients, but their clinical significance has not been established. hsrs and anaphylaxis are not mentioned in these labels. in the ema assessment report, any grade of ir has been reported to occur in . % (grade in . %), and skin rash in . % (grade in . %) of patients [ ] . symptoms of ir include: chills or shaking, dizziness, dyspnea or wheezing, fever, flushing, and itching or rash. hsrs and anaphylaxis are not mentioned in these labels. according to the bcca's drug report, irs can be observed in . - . % (severe in < %) of patients, and skin rash in - . % (severe in < %) of patients [ ] . gemtuzumab (-ozogamicin) is an antibody against glycoprotein cd , which is administered intravenously to treat acute myeloid leukemia (aml). this antibody combined with a cell-toxic substance induces apoptosis of the targeted cancer cells. in a prospective observational study involving patients, as described in the fda report, irs were observed in . % of patients, and fatal reactions in . % [ ] . in addition, local reactions to the application occurred in . %. pruritus was described in . % and rash in . % of patients. infusion-related reactions usually developed within h after infusion. signs and symptoms were similar to anaphylactic reactions. these included chills, hypoxia, fever, tachycardia, bronchospasm, and respiratory failure. according to the ema assessment report, irs, including anaphylaxis, were reported in . % of patients, and severe reactions in . % of patients [ ] . symptoms included fever, chills, injection-site urticaria, and less frequently, hypotension, tachycardia, and respiratory symptoms. all symptoms occurred during the first h after drug administration. in addition, rash was reported in . % ( s evere in . %), local erythema in . % (severe in . %), and pruritus in . % (severe in . %) of patients. the literature contains one case report of hsr mortality, which was associated with platelet transfusion after gemtuzumab treatment [ ] . ibritumomab (-tiuxetan) is used in non-hodgkin's lymphoma patients or for cancers that are refractory to rituximab. it acts by binding to the antigen cd , which is present on the surface of malignant and normal b lymphocytes. in the first fda report, allergic reactions were reported in % of patients, urticaria in %, and severe life-threatening reactions such as angioedema, lung edema, tachycardia, subdural hematoma, and pulmonary embolism in < % of patients [ ] . in the ema evaluation report, allergic (hypersensitivity) reactions and infusion reactions were described [ ] . common symptoms (up to one in patients) include skin reactions, difficulty in breathing, swelling, itching, redness, chills, and dizziness (a potential marker for hypotension). severe hypersensitivity reactions involving anaphylaxis occurred in < % of patients. according to the bcca's drug report, isrs (dermatitis, desquamation, and ulcer following extravasation) occurred in . % of patients, angioedema in . % (severe angioedema in . %), pruritus in . %, rash in . %, and mucocutaneous reactions in < % of patients (referenced in [ ] ). that particular monograph states that rituximab is an essential component of ibritumomab-based treatment regimens. infusion reactions may occur frequently after pretreatment with rituximab, or during or after ibritumomab administration. asthenia, cough, dizziness, nausea, pruritus, pyrexia, rash, rigors, tachycardia, and vomiting have been reported among the symptoms of irs. in a recent case report, patients were shown to develop human anti-murine antibodies (hama) after exposure to murine antibodies [ ] . the low incidence of hypersensitivity reactions in patients with high hama titers is particularly promising for ibritumomab. imatinib is an active ingredient in capsules or tablets containing a group of kinase inhibitors used to treat chronic myeloid leukemia. other and rarer indications include acute lymphoblastic leukemia, skin tumors, some gastrointestinal tumors, hypereosinophilic syndrome, atypical myelodysplastic or myeloproliferative disorders, as well as systemic mastocytosis. the first fda report does not include information on hypersensitivity and anaphylactic reactions. this report includes only skin rash in - % of patients (severe skin rash in - %), and pruritus in - % (severe in . - . %) [ ] . bcca drug manuals describe cutaneous reactions in < %, rash in - % (severe rash in - %), and pruritus in - % (with severe pruritus in - %) of patients. this manual does not include information on hypersensitivity and anaphylactic reactions [ ] . ipilimumab is a fully-human mab used primarily for the treatment of metastatic melanoma, metastatic colorectal carcinoma, and renal cell carcinoma. this bs shows its effect by binding and inhibiting human cytotoxic t lymphocyte antigen (ctla- ). as a result, t cell infiltration increases in tumor cells and tissues, thereby leading to their destruction. in addition, studies are ongoing for its use in the treatment of lung cancer, bladder cancer, and metastatic prostate cancer. in the fda labels, irs were reported to occur in . - . % of patients and urticaria in . % of patients [ ] . symptoms of ir include: chills or shaking, dizziness, dyspnea, feeling like passing out, fever, flushing, and itching or rash. in the ema assessment report, hsrs are reported as uncommon (≥ / -< / ), urticaria as common (≥ / to < / ), and anaphylactic reaction as very rare (< / , ) [ ] . irs were observed at a rate of . - . % depending on the dose of the drug, but grade - irs were not reported. according to the bcca's drug report, irs can be observed in . - . % (severe in < %) of patients, pruritus in - . % of patients, and skin rash in - . % (severe in < %) of patients [ ] . additionally, a case that developed a delayed type of hsr during melanoma treatment has been reported [ ] . necitumumab is a human mab used in chemotherapy combinations in the treatment of non-small cell lung cancer, and shows its effect by binding to the egfr. egfr plays an important role in the uncontrolled growth and proliferation of tumor cells and tissue as a result of apoptosis inhibition. in the fda labels, irs of any severity have been reported to occur in . % (grade in . %) of patients [ ] . these reactions have been frequently observed after the first two doses. the same data are also reported in the ema assessment report [ ] . in addition, it is stated that skin reactions can be seen in . % (grade in . %) of patients. symptoms of ir include: chills, dyspnea, or fever. urticaria and anaphylaxis are not mentioned in that report. in addition, sudden death or cardiorespiratory arrest was reported in . % of patients, which may also be a symptom of anaphylaxis. nivolumab is a human mab that is used in the treatment of advanced renal cell carcinoma, hepato-cellular carcinoma, hodgkin's lymphoma, melanoma, non-small cell lung cancer, squamous cell cancer of the head and neck, as well as urothelial cancer and acts by binding to the programmed cell death protein (pd)- -receptor. in the fda labels, irs were reported to occur in . % of patients [ ] . the ir causing permanent discontinuation or withholding of the bs was reported in . - . % of patients within h after infusion. anaphylaxis and hypersensitivity have not been reported, but it has been reported that the incidence of symptoms such as abdominal pain, arthralgia, asthenia, back pain, constipation, cough, decreased appetite, diarrhea, dyspnea, fatigue, headache, musculoskeletal pain, nausea, pruritus, pyrexia, rash, upper respiratory tract infection, and vomiting is more than %. some of these symptoms can be seen in anaphylaxis. in treatment with this bs, immune-mediated organopathies such as colitis, encephalitis, hepatitis, pneumonitis, endocrinopathies, and nephritis can also be observed. the clinical significance of adas for these side effects is still unknown. in the ema assessment report, irs and hsrs are stated as common (≥ / -< / ), rash and prutitus as very common (≥ / ), and anaphylactic reaction as rare (≥ / , -< / ) [ ] . according to the bcca's drug report, irs can be observed in . - . % of patients, pruritus in - . % of patients, skin rash in - . % (severe in . %), and urticaria in . % of patients [ ] . allergic symptoms (eyelid angioedema, flushing, and hives on the neck and face) were reported in one patient during treatment with nivolumab [ ] . however, a case with cytokine release syndrome has been reported [ ] . in a recent case report, a patient who had recurrent infusion reactions despite premedication against nivolumab was successfully treated with pembrolizumab, another pd-l /pd-l inhibitor [ ] . panitumumab is a mab egfr inhibitor delivered intravenously to treat metastatic colorectal cancer. it acts by binding and inhibiting egfr. panitumab appears to be a safe option for treatment and potential side effects in patients with anti-α-gal ige antibodies [ ] . this is due to the fact that α-galactosylated glycans are absent in purely human mabs and that the system in which it is expressed does not seem to induce posttranslational modifications like allergenic glycosylation. the chimeric mab cetuximab is not suitable in this sensitive patient group due to its high α gal content. in the fda report, irs were reported in % of patients (severe reactions in %) [ ] . severe irs included anaphylactic reactions, bronchospasm, and hypotension. angioedema and fatal reactions have also been documented in post-marketing reports. in the bcca drug monograph, hsr was reported in % of patients (occurring within h), while irs were reported in - % (severe irs were seen in %, occurring within h) (referenced in [ ] ). angioedema was seen very rarely. some cases were fatal, with possible late onset more than h post infusion. the symptoms of most irs were mild and included chills, fever, or dyspnea. in addition, pruritus was reported in - % (severe in - %) and rash in - % (severe in - %). at the annual congress of the european academy of allergy and clinical immunology (eaaci) in , a case of a successful desensitization was presented in a patient that had developed anaphylaxis against panitumumab [ ] . pembrolizumab is a monoclonal antibody that is used to treat non-small cell lung cancer, lymphoma, melanoma, and urothelial carcinoma and which binds to the pd- -receptor and inhibits its interaction with its ligands. in the ema assessment report, irs are stated as common (≥ / -< / ), rash and pruritus as very common (≥ / ), and gastrointestinal symptoms (such as abdominal pain, constipation, diarrhea, nausea, vomiting) as very common (≥ / ) [ ] . according to the bcca's drug report, irs can be observed in < . % of patients, pruritus in - . % of patients, and skin rash in . - . % (severe in . %) of patients [ ] . in the fda labels, severe or life-threatening anaphylaxis, hsrs or irs were reported in . % of patients [ ] . symptoms of ir include: chills, dizziness, fever, flushing, hypotension, hypoxemia, pruritus, rash, rigors, and wheezing. however, the incidence of adas against pembrolizumab was . %, and it was reported that ada had no clinical relevance [ ] . pertuzumab is a humanized mab that acts by blocking the human epidermal growth factor receptor protein (her ), and is used in combination with trastuzumab and docetaxel in the treatment of breast cancer. her plays an important role in the growth and differentiation of tumor cells, and its blocking results in apoptosis. according to the bcca's drug report, hsrs can be observed in . % (severe in - %) of patients, irs in [ ] [ ] [ ] [ ] [ ] [ ] [ ] . %, rash in - % (severe in %), and pruritus in % of patients [ ] . symptoms of ir include: asthenia, chills, fatigue, fever, hypersensitivity, and vomiting. in the ema assessment report, irs and rash are stated as very common (≥ / ), hsrs as common (≥ / to < / ), anaphylaxis as uncommon (≥ / -< / ), and crs as rare (≥ / , -< / ) [ ] . the most common irs have been reported as asthenia, chills, fatigue, headache, hypersensitivity, pyrexia, and vomiting. in the fda labels, irs have been reported to occur in - . % (severe less than . %), and hypersensitivity/anaphylaxis in . - . % of patients [ ] . it is stated that crs may also develop. ramucirumab is a human mab used in the treatment of gastric, breast, lung, and colorectal cancers, showing its effect by selectively inhibiting vegf receptor . vegf plays an important role in angiogenesis, cell division, and migration in vascular endothelium. according to the bcca's drug report, irs can be observed in . - . %, and rash in . % of patients [ ] . symptoms of ir include: back pain, chest pain, chills, dyspnea, flushing, paresthesias, rigors, and wheezing. some cases have been observed with more severe symptoms such as bronchospasm, hypotension, and supraventricular tachycardia. in the fda labels, irs have been reported to occur in . - . % (severe in < . %) of patients [ ] . symptoms of ir include: back pain/spasms, chills, chest pain and/or tightness, dyspnea, flushing, hypoxia, paresthesia, rigors/tremors, and wheezing. in the ema assessment report, irs can be observed in up to . % of patients [ ] . symptoms of ir include: back or chest pain, chest tightness, chills, dyspnea, feeling of tingling or numbness in hands or feet, flushing, increased muscle tension, and wheezing. some cases have been observed with more severe symptoms such as breathing distress, feeling faint, and tachycardia. anaphylaxis and hypersensitivity are not mentioned in that report. trastuzumab is an anti-her -antibody combined with or conjugated to a cytotoxic drug that is used to treat local or advanced inoperable breast and gastric cancer. in the bcca drug assessment report (referenced in [ ] ), irs occurred in - % of cases, with severe reactions in %. allergic reactions were reported in % of cases. irs occurred in % of patients with their first infusion. mild to moderate symptoms included headache, dizziness, rash, cough, nausea, vomiting, pain, chills, and asthenia. severe reactions and symptoms were most often respiratory distress, bronchospasm, wheezing, low oxygen saturation, and hypo-or hypertension. [ ] ) reported an hsr rate of . - . % and an infusion-related reaction rate of %. according to the ema assessment report , hsr was reported in . - . %, irs in . - . %, and anaphylaxis in - . % of patients [ ] . according to the fda's report, irs occur in . - . % of patients and include symptoms such as fever, chills, redness, shortness of breath, bronchospasm, wheezing, tachycardia, and hypotension. in the same report, pruritus was seen in . % and drug hypersensitivity was found to occur in . - . % of patients. in preliminary studies, anaphylaxis was seen in one patient [ ] . bss are often used to treat rheumatoid arthritis, psoriatic arthritis, ankylosing spondylitis, inflammatory bowel diseases (ulcerative colitis, crohn's disease), still's disease, and vasculitis in rheumatology. details on the reactions are given in tab. . adalimumab is a recombinant human igg that binds to tnf α with high affinity. it acts by inhibiting the binding of tnf α to p and p surface receptors on target cells. in a study on patients conducted by puxeddu et al. [ ] , hypersensitivity reactions occurred in seven patients ( . %), local reactions occurred in three patients ( . %), and urticaria and angioedema occurred in three patients ( . %). anaphylaxis was not observed in this study, but has been reported very rarely in the post-marketing period. another study conducted by tarkiainen et al. [ ] reported hsr frequencies of . %, isrs frequencies of . %, and allergic symptoms in . % of the patients. fda labeling showed that local isrs are more common in patients treated with the tnf α inhibitor than in those treated with placebo [ ] . the incidence of isrs including erythema, itching, hemorrhage, and pain or swelling is . - . % in patients treated with adalimumab vs % in patients treated with placebo. anaphylaxis and angioedema have rarely been reported following the application of this bs, and in the first weeks of treatment, approximately % of patients had non-severe local allergic hypersensitivity reactions and allergic rashes. anakinra is a genetically engineered and recombinant human il receptor antagonist. its main effect is by neutralizing the biological activity of il- α and il- β, a proinflammatory cytokine. as a result, it contributes to the reduction of synovial inflammation by reducing many cellular responses. however, anakinra has the following therapeutic indications: rheumatoid arthritis (ra), cryopyrinassociated periodic syndrome (caps), and still's disease. in the ema summary of product characteristics (smpc) report, isrs are generally reported to occur in the first weeks of treatment (for ra and still patients) and resolve within - weeks [ ] . in that particular report, allergic reactions such as anaphylactic reactions, angioedema, urticaria and pruritus are expected to range from / to / , . according to the fda's label, the most frequently reported adverse reactions were isrs ( . %), which were mild to moderate in the majority of patients [ ] . only . % of these reactions were classified as severe, with the majority includ-ing ecchymosis, erythema, inflammation, and local pain. reactions usually occurred during the first month. this label states that anaphylaxis, angioedema, urticaria, and rash may occur, but no specific rate or detailed information is provided. in the literature, it has been reported that anakinra treatment may cause anaphylaxis, cutaneous drug reactions, erythematous plaques, pruritic rash, and severe hsrs [ ] . in a pediatric case, a patient with anakinra-induced anaphylaxis was successfully treated with canakinumab, an alternative anti-il agent [ ] . however, a desensitization protocol is also available [ ] . belimumab is a recombinant human igg mab that binds and inhibits the biological activity of the sol- in the ema assessment report, any event of isrs (local pain, erythema, hematoma, induration, and pruritus) were reported in . % of patients, hsrs in < . % of patients, and post-injection anaphylactic reactions were reported in . - . % of patients (serious reactions, . %) [ ] . hsr and anaphylactic reaction are mentioned to occur mostly during the first two infusions. in the newly published fda report, intravenous administration was evaluated [ ] . hsrs were reported in . % (in the placebo group %), anaphylaxis in . % (placebo . %) of patients. symptoms included angioedema, dyspnea, hypotension, pruritus, rash, or urticaria. irs were observed in % of patients and severe irs in . %, with placebo in % and . %, respectively. it is stated that it is not easy to differentiate between hsrs and irs due to the overlap in signs and symptoms. however, it has been reported that there is insufficient evidence as to how the premedication of some patients affects the frequency or severity of reactions. canakinumab is a human mab against il- beta and indicated in periodic fever syndromes, caps, tumor necrosis factor receptor-associated periodic syndrome (traps), familial mediterranean fever (fmf), still's disease, and gouty arthritis. according to the fda's label, the most frequent adverse drug reactions were infections predominantly of the upper respiratory tract, and mild to moderate isrs equal or greater than . % [ ] . no anaphylactoid or anaphylactic reactions were observed in more than , patients whose clinical development was tested for this biologic, but the risk of severe hypersensitivity reactions was mentioned. no cases of canakinumab-related anaphylaxis have been reported in the current literature [ ] . in addition, patients with anaphylaxis to anakinra have been shown to tolerate canakinumab very well [ ] . etanercept is a subcutaneously administered selective immunosuppressive and anti-inflammatory bs belonging to the group of tnf α inhibitors used to treat rheumatoid arthritis and other rheumatic diseases. it exerts its effect by inhibiting the activity of tnf α. puxeddu et al. reported a . % rate of hsrs, a . % rate of anaphylaxis, a . % rate of angioedema or urticaria, and a . % rate of local reactions in a study involving patients treated with etanercept [ ] . in another study, the rate of hsrs was . %, that of isrs was . %, and reactions at sites other than the injection site occurred at a rate of . % [ ] . the fda drug label published in reports that allergic reactions associated with this treatment occur in < % of patients, and mild to moderate isrs (erythema, itching, pain, swelling, bleeding, bruising) occur in - % of patients during the first months of treatment [ ] . isrs were usually seen in the first month, and with decreased frequency thereafter. golimumab is a selective immunosuppressive and anti-inflammatory tnf-α-inhibitor applied subcutaneously to treat rheumatoid and psoriatic arthritis, as well as ankylosing spondylitis. it is a mab that inhibits the activity of soluble and membrane-bound forms of the proinflammatory cytokine tnf α by binding tnf α and inhibiting its r eceptor binding. in a -year safety study, the frequency of isrs was . - . %, depending on the drug dose. no anaphylactic reactions or serum sickness-like reactions have been reported to date during the follow-up period of up to weeks [ ] . the fda drug label reports an isr frequency of . - . % [ ] . the reactions were often mild and included erythema, urticaria, induration, and pain. no anaphylactic reactions were observed in phase ii and iii studies with the drug. infliximab is an intravenously administered selective immunosuppressive and anti-inflammatory tnf α inhibitor used to treat rheumatoid and psoriatic arthritis, crohn's disease, and ankylosing spondylitis. this bs is a human-murine chimeric monoclonal igg κ antibody that binds tnf α and inhibits its effects. in a review by maggi et al. summarizing nine studies, infusion reactions occurred in - % of patients depending on the disease being treated ( referenced in [ ] ). in a study on patients, puxeddu and colleagues reported hypersensitivity reactions in . %, anaphylaxis in . %, urticaria or angioedema in . %, and local reactions in % of patients [ ] . another study reported a . % rate of infusion-related hsrs, and a . % rate of anaphylaxis [ ] . the fda drug label reports irs in % of patients during or within h after the infusion [ ] . a total of % of all patients given infliximab infusions experienced nonspecific symptoms (fever or tremor), while % experienced cardiopulmonary reactions (chest pain, hypotension, hypertension, or dyspnea). symptoms related to pruritus, urticaria, and cardiopulmonary reactions were detected in % of patients. serious infusion reactions such as anaphylaxis, convulsions, erythematous rash, and hypotension have occurred in < % of patients. delayed type hsr was detected in approximately % of patients. this usually occurred within weeks of re-infusion, and was a combination of fever and/or rash symptoms with arthralgia and/or myalgia. these findings have been classified and reported as serum sickness. in a study conducted by vultaggio et al., it was reported that ige or igm antibodies against infliximab could be detected and may play a role in igeand non-ige-mediated anaphylaxis [ ] . skin testing and infliximab-specific antibody detection have been shown to be useful in predicting and preventing severe drug-related reactions [ ] . certolizumab is a subcutaneously administered selective immunosuppressive and anti-inflammatory tnf-α-inhibitor used to treat rheumatoid and psoriatic arthritis, spondyloarthritis, and crohn's disease. it is a mab that binds to and blocks the activity of the proinflammatory cytokine tnf α. in the ema assessment report, mild to moderate isrs were reported in . % of patients, injection-related acute systemic hsr (pre-syncope) was reported in . %, and delayed hsr was seen in . % of patients [ ] . recently published fda labels have reported that hsrs may rarely occur and include allergic dermatitis, rash, angioedema, dizziness, pyrexia, flush, isrs, malaise, serum sickness, hypotension, dyspnea, and vasovagal syncope [ ] . it is noted that some of these reactions were observed after the first administration. isrs were reported in . - . % of patients. although the literature contains no clear information on rates of hypersensitivity and anaphylaxis, it is stated that no single-drug related anaphylactic shock was observed in the one study conducted in [ ] . rituximab is an intravenously delivered mab that selectively binds to the cd antigen of b lymphocytes. indications for its use include non-hodgkin's lymphoma, rheumatoid arthritis, and chronic lymphocytic leukemia. maggi et al. have reported acute infusion reaction frequencies of - % during the first infusion (referenced in [ ] ). the fda drug label for subcutaneous use [ ] reports a - % rate of local isrs (pain, swelling, induction, hemorrhage, rash, pru-ritus, and erythema). incidence rates reportedly decrease subsequent to the initial injection. the label also states that severe cytokine release syndrome and its symptoms (fever, tremor, urticaria, angioedema, bronchospasm, and severe dyspnea) may be seen within - h of infusion. however, the ratio is not specified in this label. the recent fda drug label for intravenous use [ ] reports a ≥ . % rate of irs that typically ( %) occurred at the first infusion with the time onset between - min. this rate decreased after each ongoing infusion. severe irs may include symptoms such as acute respiratory distress syndrome, anaphylactic/anaphylactoid events, angioedema, bronchospasm, cardiogenic shock, hypoxia, hypotension, myocardial infarction, pulmonary infiltrates, urticaria, ventricular fibrillation, and death. according to the bcca drug assessment report (referenced in [ ] ) hsrs occurred in - . % of patients, irs in - . % (severe, - . %), isrs in . %, and urticaria in . % (severe, . %) of patients. isrs include local erythema, hemorrhage, induration, pain, pruritus, rash, and swelling at the injection site. in a study conducted in , the presence of serum anti-rituximab antibodies was reported in some patients in association with a less favourable treatment outcome [ ] . vultaggio and colleagues have shown that rituximab plays a role in specific t-helper cell and ige responses in acute infusion reactions in addition to cytokine release mechanisms [ ] . a less frequent use of bss is to minimize the rejection of transplanted organs (mostly kidneys, hearts, and livers). basiliximab, muromonab, and daclizumab have been developed for this purpose. today, only basiliximab is approved for this application. the other two drugs have been withdrawn from the market for a variety of reasons. details about the reactions are given in tab. . basiliximab is an immunosuppressive intravenously administered chimeric ( % human, % murine) mab. it is used in combination with other immunosuppressive drugs to prevent acute graft rejection following renal transplantation. it acts by binding the alpha chain of the il receptor on the surface of t lymphocytes. the fda drug label reports a > . % rate of side effects including nausea, vomiting, fever, dyspnea, and hypertension, and . - . % of side effects include angina pectoris, cardiac failure, tachycardia, dizziness, bronchospasm, pulmonary edema, rhinitis, rash, and pruritus [ ] . more detailed informa-tion is not provided. in a study conducted in , infusion-related isrs (pain, swelling, erythema) were found in . % of patients [ ] . in post-marketing surveys, hsrs have been reported in patients undergoing basiliximab therapy, including one case of ige-mediated anaphylaxis [ ] . data on this bs are relatively limited, but according to the reports it seems to be mainly well tolerated by the patients [ ] . allergic reactions and anaphylaxis do not seem to be easily identified, since the patients experience intense immuno suppressive effects due to the concomitant medication. it does not induce crs like muromonab-cd , and its side effects are reported to be similar to placebo. belatacept is a human ctla- /human igg fusion protein that selectively inhibits t cell activity by binding to cd and cd of the antigen presenting cells. according to the fda labels [ ] , no patient had developed anaphylaxis or hsrs at the year follow-up. however, milder irs ( %) were reported, similar to placebo, within h of infusion. in post-marketing experiences, a case of anaphylaxis has been reported. in the ema assessment report [ ] , acute infusion events (flushing, headache, hypotension, hypertension) were reported in . - . % of patients. symptoms of diarrhea and pyrexia (≥ %), as well as hypertension, peripheral edema, nausea, and vomiting (≥ %), which may be "possible peri-infusional events," were reported in year follow-up of the patients. there was no association found between these reactions and belatacept adas. anaphylactic events have been observed in the post-marketing period, but no incidence was given. muromonab is an immunosuppressive mab given to reduce acute rejection in patients with kidney, liver, or heart transplants. it exerts its effect by targeting the cd receptor, a membrane protein expressed on the surface of t cells. developments and progress of medical treatment in transplant medicine led to its withdrawal in . the literature contains information on cytokine release syndrome and anaphylaxis. in a book on this subject, symptoms of anaphylaxis such as pyrexia ( %; . °c or higher in % of cases), chills ( %), dyspnea ( %), nausea and vomiting ( %), chest pain ( %), diarrhea ( %), tremor ( %), bronchospasm ( %), headache ( %), tachycardia ( %), stiffness ( %), and hypertension ( %) were reported [ ] . pilot studies of this drug have been performed orally in patients with moderate to severe ulcerative colitis [ ] . in this study, no severe side effects or crs have been reported. daclizumab is a subcutaneously administered, immunosuppressive mab used to treat relapsingremitting multiple sclerosis and to prevent rejection of transplanted kidneys. the antibody reduces the number of circulating activated t cells by binding to the alpha subunit of the il- receptor on t lymphocytes and inhibiting interaction with il- . in , the drug was withdrawn from the market owing to reports of severe hepatotoxic side effects and encephalopathies. other indications for the use of biologicals include multiple sclerosis, paroxysmal nocturnal hemoglobinuria, instable angina pectoris during insertion of cardiac catheters, congenital bronchopulmonary dysplasia, and congenital heart disease. abciximab is an intravenously administered chimeric mab fragment that inhibits binding of platelet adhesion proteins (such as fibrinogen, fibronectin, von-willebrand-factor) to glycoprotein iib/iiia receptors on the surface of platelets. it is frequently used to prevent ischemic complications associated with invasive heart procedures, as well as to prolong cardiac infarct prophylaxis in patients with unstable angina pectoris due to its antithrombotic effect. the registry study of the drug reported hypersensitivity rates of . - . %, an urticaria rate of . %, and no cases of anaphylaxis or angioedema [ ] . in the same study, human anti-chimeric antibodies were found to be associated with thrombocytopenia. the literature contains a very small number of case reports. one developed an anaphylaxis - h after treatment [ ] . the fda label reported bronchospasm ( . %), injection site pain ( . - . %), and pruritus ( . %) [ ] . no incidence of anaphylaxis has been reported in any of the phase iii studies (clinical trials). eculizumab is the humanized mab used primarily for the treatment of paroxysmal nocturnal hemo-globinuria and atypical haemolytic uremic syndrome. it can also be used in the treatment of generalized myasthenia gravis and neuromyelitis optica spectrum disorder with antibody positivity. this drug acts by binding to the c component of the complement system, inhibiting its activation, and consequently contributes to reducing erythrocyte destruction and hemolysis. fda labels reported that irs, hrs, and anaphylaxis may develop during treatment, but irs requiring discontinuation of the drug have not been reported [ ] . in the ema assessment report, hsrs, irs, urticaria, and anaphylaxis are stated as uncommon (≥ / -< / ) [ ] . lanadelumab is a human mab (class igg kappa) that specifically inhibits the activity of plasma kallikrein, used to reduce attacks and representing a prophylaxis measure in patients with hereditary angioedema [ ] . in a study published in , . % of patients reported isrs ( . % in the placebo group) with symptoms such as bruising, dizziness, erythema, and local pain [ ] . only one patient ( . %) developed mild-to-moderate hsrs, which included transient symptoms of oral tingling and pruritus, and resolved spontaneously within day after onset without need for medication. the ema evaluation report [ ] provides more detailed data on the study conducted by banerji et al. [ ] . in this report, isrs are given as follows; pain ( . %), erythema ( . %), bruising ( . %), discomfort ( . %), hemorrhage ( . %), pruritus ( . %), swelling ( . %), hematoma ( . %), induration ( . %), paresthesia ( . %), warmth ( . %), edema ( . %), and rash ( . %). in addition, no events of anaphylactoid reaction or anaphylaxis were reported (referenced in [ ] ). natalizumab is an intravenously administered, immunosuppressive recombinant humanized igg antibody that is produced in mouse cells. it acts by binding integrin a and is used to treat relapsingremitting multiple sclerosis. maggi and colleagues (referenced in [ ] ) reported irs in - % of the patients in their review. according to the ema evaluation report [ ] , . % of patients experienced infusion-related reactions, hsrs occurred in up to % of patients, and anaphylactic/anaphylactoid reactions in less than %. hsrs usually occurred during infusion or within h after completion. hypersensitivity symptoms included rash and urticaria, hypotension, hypertension, chest pain, chest complaints, dyspnea, and angioedema. the fda's label reported infusion-related reactions in - % of patients (placebo - . %), acute urticaria in - %, acute hsrs in . %, pruritus in %, and serious irs and anaphylaxis in < % [ ] . symptoms frequently associated with these reactions were fever, chills, rash, urticaria, pruritus, dizziness, nausea, flush, hypotension, dyspnea, and chest pain. the reactions occurred mostly within h after infusion. palivizumab is an intramuscularly administered humanized mab ( % human and % murine) that is used in pediatric patients to prevent respiratory syncytial virus (rsv) infections. the antibody binds to coat proteins on the viral surface, thereby inhibiting its entry into host cells. it is used in patients with bronchopulmonary dysplasia, cystic fibrosis, and neonatal heart disease that are considered to be at high risk of severe rsv-associated disease. serious acute hsrs were rare among the , patients treated to date (> million doses) [ in cr e a si n g r is k fo r se v e re r e a c ti o n s bss are increasingly used today and play an important role in the treatment of allergic, pulmonary, dermatological, oncological, rheumatological, as well as various other diseases. these drugs can be used as first-line or second-line treatment, as well as for prevention purposes. observed hsrs may be mild, moderate, but may also be life-threatening by severely affecting vital parameters. all organ systems can be affected during reactions, so clinical symptoms are quite diverse. reactions may occur due to allergic, non-allergic, or immunological side effects. the risk of allergic reactions decreases proportionally with the increase in human homology of the bs ( fig. ; [ ] ). vultaggio and co-authors explain the reactions to bss with two possible main mechanisms: (i) loss of immune tolerance due to side effect of the bss; and (ii) triggering of immune responses and reactions to non-self epitopes in bss [ ] . this comprehensive review reveals a hypersensitivity prevalence against various bss and an inconsistency and lack of precision regarding the nomenclature used for immune reactions to bss (hypersensitivity, anaphylaxis, anaphylactoid, and infusion reactions) in the literature. theoretically defined hypersensitivity and the hypersensitivity that is observed and classified in clinical practice seem to be different. similarly, many data reported as anaphylaxis actually describe severe anaphylactic reactions (grades iii or iv). it became evident that these definitions describe many common symptoms. due to the overlap in signs and symptoms in the reported descriptions, it is not always possible to differentiate these reactions properly according to their pathomechanism. the common points of all these reactions are indicated in tab. [ , , ] . in addition, reactions such as pruritus, flush, and urticaria are also included in the definitions of ir, hsr, anaphylaxis, and crs. a summary view of the reactions according to severity is shown in fig. . fig. shows a synopsis of the incidences of hsrs (fig. a) , acute irs (fig. b) , and anaphylaxis to bss (fig. c) as extracted by the authors from the comprehensive review of the databases. there is an urgent need to define and classify these reactions more precisely in the future. furthermore, the development of a scoring system, including symptom-or system-based approaches, can make it easier for us to identify the reactions of our patients and treat them adequately. this scoring can be evaluated according to which organ system (cardiac, gastrointestinal, hematologic, muscular, neurological, renal, respiratory, skin, vascular) is affected and the severity of symptoms. one of the important points that may have been overlooked is the understanding of whether some symptoms (such as nasopharyngitis, conjunctivitis, rhinitis, thrombocytopenia, anemia, or gastrointestinal symptoms, etc.) have an immunological origin. the role of adas in these symptoms is not yet known. in addition, the risk of patients at each injection/infusion should be defined according to real life. in general, the literature gives percentages by calculation and only follow-up reactions in the treatment process. the long-term evaluations seem to be rather small in number. crs, which is a form of severe ir, can also be observed during alemtuzumab, anti-cd monoclonal antibody tgn , anti-thymocyte globulin, blinatumomab, brentuximab, dacetuzumab, muro monab-cd , nivolumab, rituximab, and obinutuzumab treatments (referenced in [ ] ). a large majority of patients may have sepsis/septic shock fragments, as well as symptoms that may also be confused with tumor lysis syndrome. il- plays an important role in the development of crs; therefore siltuximab or tocilizumab, a chimeric and a humanized mab against il- and the il- receptor, respectively, are used for the treatment of this condition [ ] . in cases where crs developed during treatment, the relationship between the drug itself and ada is so far unknown. bss are important immunotherapeutic tools used in the treatment of many diseases today. various side effects and reactions can be observed during c treatment, but there are deficiencies in their identification and classification. during this comprehensive research, it became evident that a more precise nomenclature should be applied. in order to achieve a harmonization in this regard, a simpler symptomor system-based classification and scoring system is needed. a better understanding of the pathophysiology of hypersensitivity reactions and increased clinical experience in the treatment of side effects will provide timely control of unexpected reactions. as a result, immunotherapy with bss will become safer in the future. adverse drug reactions to biologics clinical 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management and preparedness for infusion and hypersensitivity reactions guideline for acute therapy and management of anaphylaxis. s guideline of dgaki cytokine release syndrome a. gülsen: data collection, design of tables and figures, analysis and interpretation of data, writing and drafting of the manuscript; b. wedi: major role in revising the manuscript; u. jappe: concept of the manuscript, acquisition of data, design of figure , writing and revising the manuscript. a. gülsen, b. wedi, and u. jappe declare that they have no competing interests. this article is licensed under a creative commons attribution . international license, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons licence, and indicate if changes were made. the images or other third party material in this article are included in the article's creative commons licence, unless indicated otherwise in a credit line to the material. if material is not included in the article's creative commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. to view a copy of this licence, visit http://creativecommons.org/licenses/by/ . /. gülsen a, wedi b, jappe u. hypersensitivity reactions to biologics (part i): allergy as an important differential diagnosis in complex immune-derived adverse events. allergo j int ; : - https://doi.org/ . /s - - - key: cord- -d qj b authors: vincent, jean-louis; abraham, edward; annane, djillali; bernard, gordon; rivers, emanuel; van den berghe, greet title: reducing mortality in sepsis: new directions date: - - journal: crit care doi: . /cc sha: doc_id: cord_uid: d qj b considerable progress has been made in the past few years in the development of therapeutic interventions that can reduce mortality in sepsis. however, encouraging physicians to put the results of new studies into practice is not always simple. a roundtable was thus convened to provide guidance for clinicians on the integration and implementation of new interventions into the intensive care unit (icu). five topics were selected that have been shown in randomized, controlled trials to reduce mortality: limiting the tidal volume in acute lung injury or acute respiratory distress syndrome, early goal-directed therapy, use of drotrecogin alfa (activated), use of moderate doses of steroids, and tight control of blood sugar. one of the principal investigators for each study was invited to participate in the roundtable. the discussions and questions that followed the presentation of data by each panel member enabled a consensus recommendation to be derived regarding when each intervention should be used. each new intervention has a place in the management of patients with sepsis. furthermore, and importantly, the therapies are not mutually exclusive; many patients will need a combination of several approaches – an 'icu package'. the present article provides guidelines from experts in the field on optimal patient selection and timing for each intervention, and provides advice on how to integrate new therapies into icu practice, including protocol development, so that mortality rates from this disease process can be reduced. sepsis is the tenth most common cause of death in the us [ ] . a recent us study reported that severe sepsis accounts for in excess of , deaths annually from a total population of approximately , patients-a mortality rate of approximately % (with published studies quoting a range of - %) [ ] . this persistent, high mortality rate is clearly unacceptable, given that it ranks sepsis above some of the higher profile causes of in-hospital death, including stroke ( - % risk of death in the first days) and acute myocardial infarction (ami) ( % risk of death in the first days) [ ] . moreover, the actual number of deaths associated with the condition may be even higher than current estimates suggest. many sepsis patients have at least one comorbidity and deaths are often attributed to these conditions rather than to sepsis [ ] [ ] [ ] . unfamiliarity with the signs and symptoms of sepsis may further hinder accurate diagnosis. there are many possible reasons for this high mortality. sepsis is certainly a complex disease state; the pathophysiology is only now beginning to be unraveled, and it is complicated by heterogeneous presentation (possible signs of sepsis are presented in table ). while none of these signs alone is specific for sepsis, the otherwise unexplained presence of these signs should signal the possibility of a septic response. many cases of sepsis are recognized late, and patients are often inappropriately treated before entering the intensive care unit (icu) by physicians unfamiliar with the signs and symptoms of the condition. furthermore, treatment may be initiated by any of a number of physicians (anesthetists, hematologists, intensivists, infectious disease specialists, pulmonologists, and emergency physicians). there are presently various defined supportive strategies for treating patients with sepsis, but improvements are needed to reduce the unacceptably high mortality rate. moreover, as with other areas of medicine, the application and integration of new but proven strategies for reducing morbidity and mortality into clinical practice has been slow. encouraging new data have recently been presented on new approaches to the management of patients with sepsis. many of these approaches attempt to modulate or interrupt the sepsis cascade and to address the cause of multiorgan dysfunction. although many of these approaches are in early phases of development (e.g. antibodies to tumor necrosis factor [tnf] alpha, bactericidal permeability increasing protein, high-flow hemofiltration to remove circulating inflam-matory mediators, platelet-activating factor acetyl hydrolase, and antielastases), other approaches are more advanced and are already beginning to impact on outcomes in the icu. at a roundtable discussion in london in june , professor jean-louis vincent brought together five experts to discuss more effective implementation of five exciting new interventions in the icu setting to decrease the unacceptable burden of mortality in patients with severe sepsis. each of the roundtable panelists is a highly respected physician in the world of sepsis and critical care medicine. the interventions discussed encompassed low tidal volume in patients with acute lung injury (ali)/acute respiratory distress syndrome (ards) (edward abraham), early goal-directed therapy (egdt) (emanuel rivers), drotrecogin alfa (activated) (gordon bernard), moderate-dose corticosteroids (djillali annane), and tight control of blood sugar (greet van den berghe). the purpose of the roundtable discussion was to provide guidance for clinicians on the integration of new interventions into the icu to reduce the mortality in sepsis, on appropriate patient selection for these interventions, and on appropriate timing of these interventions. the present review reports the discussions and recommendations of the panel. the overall -day mortality in the icu is typically ~ % [ ] . the -day mortality in the population with severe sepsis, defined as sepsis with organ dysfunction, is - %. it is clear from this figure that severe sepsis contributes disproportionately to the overall -day mortality in the icu and compares unfavorably with some of the higher profile acute killers in hospital (e.g. stroke and ami) [ ] . despite the general improvements in medicine overall, this mortality rate has remained essentially unchanged for the past years. this has contributed to a feeling of pessimism among table possible signs of sepsis (adapted from [ intensivists and other medical professionals regarding treatment prospects for severe sepsis, and a reluctance to rapidly incorporate new interventions into clinical practice [ ] . although the sepsis mortality rates are unacceptable, they camouflage some significant developments that are and have been occurring for hospital patients, for the general icu population and, particularly, for those with severe sepsis. direct comparison of mortality rates among patients with identical acute pysiology and chronic health evaluation (apache) scores in the placebo arm of anti-tnf or anti-endotoxin studies published - years ago [ ] [ ] [ ] with more recent studies [ , ] , demonstrates that the mortality rate is much lower in more recent studies. interestingly, this decrease was apparent even before the five interventions discussed in the present article were published, reflecting improvements in the general supportive care of sepsis patients. indeed, the panel contends that mortality from septic shock has already been reduced. some patients who in the recent past would have died from severe sepsis or septic shock do not reach the icu now because they are well managed on the wards, in the emergency department, and even during preoperative and postoperative care. for example, those sepsis patients that receive prompt antibiotic therapy have a - % lower mortality rate than those who receive antibiotic therapy later in their care [ ] . progress is also being made in diagnosing sepsis: more patients are being tested to identify the source of infection and the pathogens involved, supportive care measures have been improved (e.g. hemodynamic support), and other measures have been put in place to reduce the incidence of nosocomial infections (e.g. reducing the need for pulmonary artery catheters by using echo techniques to assess cardiac function). there has also been a realization of the importance of specially trained intensive care physicians in the icu. it has been internationally recognized that changing the icu from an 'open format', whereby patients are cared for by their admitting physician, to a 'closed format', whereby patients are managed by appointed intensivists, reduces mortality rates [ ] . although the mortality rate is beginning to decline, it still remains unacceptably high. furthermore, the number of patients with severe sepsis and septic shock is increasing; people are living longer, and there has been a rise in the number of immunocompromised patients due to aggressive cancer therapy and the increased prevalence of hiv. in-hospital ami-associated mortality rates averaged approximately - % in the s [ ] . this clearly unacceptable mortality rate was addressed by the development of a number of new pharmacological and mechanical interventions together with improvements in supportive care. in the landmark second international study of infarct survival trial, published in , , suspected ami patients were treated with either streptokinase or aspirin, with both drugs, or with neither. the mortality rate in the combination group of this trial was %, compared with . % in those patients given neither streptokinase nor aspirin [ ] . cardiologists have effectively implemented multiple pharmacologic and supportive care interventions to reduce mortality in ami from - % to % and lower. not satisfied with this already remarkable figure, they are trying to reduce it further. physicians treating patients with sepsis are clearly faced with a very different situation to those treating patients with ami, and so direct comparisons are not possible. however, several factors have contributed to the success of ami therapy and possibly to the lack of such success in sepsis (table ) . sepsis is undoubtedly complicated. however, many of the lessons that have been learned through effective application of therapies in other disease states can be applied to severe sepsis. furthermore, the encouraging data that are beginning to appear in the literature indicate that sepsis may not be as intractable to treat as once thought. the following sections provide salient information on five interventions that have shown a significant positive impact on mortality rates in sepsis, severe sepsis, septic shock, or sepsis-related diseases in recent clinical trials. the interventions were presented at the roundtable by one of the principal investigators of the key trial of the intervention. each section concludes with recommendations for the integration of the particular intervention into clinical practice. the traditional approach in patients with ali/ards is to ventilate using tidal volumes between and ml/kg body weight, almost twice the average tidal volume at rest ( - ml/kg body weight), and to maintain a low positive endexpiratory pressure (peep). the purpose of this approach is to achieve normal values for the ph and partial pressure of arterial carbon dioxide. however, this method leads to high inspiratory airway pressures and to excessive stretch of the aerated lung. in , tremblay et al. examined the effect of ventilation strategy on lung inflammatory mediators in the presence and absence of a pre-existing inflammatory stimulus in sprague-dawley rats [ ] . in both stimulated and nonstimulated groups, the presence of inflammatory mediators (tnf-α, il- β, il- , il- , macrophage inflammatory protein , and ifn-γ) was highest in those rats ventilated with a large tidal volume and zero peep. furthermore, in a study by ranieri et al. in [ ] , the concentration of inflammatory mediators hours after randomization of the groups was significantly lower in the lung-protective strategy group (tidal volume, . ± . ml/kg) than in the control group (tidal volume, . ± . ml/kg) (p < . ). following on from the positive results in the tremblay et al. trial [ ] , a small study ( patients) was carried out by amato et al. in brazil [ ] . the mortality rate was % in patients given 'protective' ventilation (peep above the lower inflection point on the static pressure-volume curve, tidal volume < ml/kg ideal body weight, driving pressures < cmh o above the peep value, permissive hypercapnia, and preferential use of pressurelimited ventilatory modes) compared with % in patients on conventional ventilation (p < . ). this impressive reduction in mortality was tempered by the higher than normal mortality level in the control group, prompting the national institutes of health-funded acute respiratory distress syndrome network to set up a similar, larger ( patients), prospective, multicenter, randomized trial in the us [ ] . for a summary of the protocol used in this study, see appendix . the trial was stopped after the fourth interim analysis because the use of lower tidal volumes was found to be associated with a significantly reduced mortality (p = . for the difference in mortality between groups). the primary endpoints were mortality prior to hospital discharge with unassisted breathing and ventilator-free days (days alive, off mechanical ventilation, between enrollment and day ). both of these endpoints were achieved (figs - ). in addition, patients receiving a tidal volume of ml/kg ideal body weight had increased organ failure free days and lower il- levels. ali is seen in - % of patients with sepsis [ ] . although the approach has only been tested in patients with ali/ards, a tidal volume of ml/kg ideal body weight is at the lower end of the range of physiologic ventilation. hence, this approach should be suitable for most patients in the icu setting. furthermore, as many patients with severe sepsis or septic shock progress to frank ali/ards, the panel believes that low tidal volume therapy is a valid option in these patients, and an option that may indeed prevent the development of ali/ards. although patient selection in the clinical trial specified both blood gas and lung infiltrate criteria, at least % of patients in the general icu setting meet the criteria for blood gas but table a comparison of acute myocardial infarction (ami) and sepsis market issues significant publicity surrounding and general awareness lack of understanding among physicians and the of the condition; large trials general public diagnosis a relatively straightforward and relatively common complicated by a long list of signs and symptoms diagnosis (electrocardiogram, enzymes, troponin), and and few objective tools for validation one that can be made by generalists, not just cardiology specialists generally single organ disease (notable exception when often chronic or acute comorbidities complicated by cardiogenic shock) generalists have been taught to recognize the signs sepsis patients often come 'second hand' from a and symptoms of ami; initial treatment is usually specialist who may not be appropriately trained to provided by emergency physicians, who are trained diagnose, manage, and refer patients with sepsis to treat these patients mortality prior to hospital discharge in patients receiving a tidal volume of and ml/kg ideal body weight. acidosis is more likely to develop in patients with severe lung problems rather than in those exhibiting milder disease when tidal volumes are kept low. however, acidosis is seldom a clinical problem and rarely requires administration of bicarbonates. one of the issues with low tidal volume therapy is that the patients are often more uncomfortable, at least initially, when they are being ventilated with a tidal volume of ml/kg ideal body weight. the patients tend to exhibit tachypnea and may become more agitated. sedation is generally required, but the ventilator setting can be maintained. of more concern is that icu staff may consider a respiration rate of /min to be a sign of something more serious and may attempt to terminate the intervention. education of staff is clearly essential. the strategy assessed in this trial not only includes ventilation with a low tidal volume, but also the provision of extrinsic peep. there may be some concern that an increased respiratory rate may result in intrinsic peep and hemodynamic problems (e.g. decreased cardiac filling, decreased cardiac output, and diminished blood pressure). the panel believes that auto peep was not an issue in the acute respiratory distress syndrome network study. in addition, in the groups with low tidal volume, at least % more oxygen was required to maintain the fraction of inspired oxygen (fio ), suggesting that there was very little auto peep occurring. when mechanical ventilation is indicated for treatment of patients with ali/ards, the tidal volume should be limited tõ ml/kg ideal body weight. goal-directed therapy represents an attempt to adjust the cardiac preload, afterload, and contractility to balance systemic oxygen delivery with oxygen demand. in patients with severe sepsis and septic shock, such an approach would seem eminently reasonable as part of general supportive measures to restore and maintain adequate cellular perfusion and to prevent organ dysfunction. in the setting of the icu, however, supranormal and normal approaches have met with little or no success [ , ] . it is possible that, by the time these therapies are applied in the icu, any such intervention may have been too late. hence, the focus has shifted towards hemodynamic optimization in the early presentation of disease, such as in the emergency department. a prospective, randomized, predominantly blinded study was initiated by the early goal-directed therapy collaborative group to examine the results of hemodynamic interventions in the emergency department [ ] . in this study, patients were randomly assigned to either hours of egdt or to standard therapy prior to admission to the icu. baseline characteristics (including the adequacy and duration of antibiotic therapy) in the egdt and standard therapy groups were not significantly different. the vital signs, resuscitation endpoints, organ dysfunction scores, and coagulation-related variables were similar in these groups at baseline [ ] . however, there were some important differences between the treatment groups (see table ). available online http://ccforum.com/content/ /s /s proportion of patients alive and off the ventilator having been ventilated with a tidal volume of and ml/kg ideal body weight. median number of ventilator-free days in patients receiving a tidal volume of and ml/kg ideal body weight. patients randomized to egdt received the same therapy but, in addition, were monitored for the endpoint of central venous oxygen saturation (scvo ) > %. egdt patients were given more intravenous fluids (including blood transfusions) and more inotropic support (mostly dobutamine). for more information on the protocol used in this study, see appendix . key data are presented in table . the in-hospital mortality was . % in the group assigned to egdt and was . % in the group assigned to standard therapy (p = . ), indicating that egdt provides significant benefits in improving outcomes in patients with severe sepsis and septic shock. during the interval from to hours, patients assigned to egdt exhibited a more significant improvement in mean scvo ( . ± . % versus . ± . %), in lactate concentration ( . ± . mmol/l versus . ± . mmol/), in base deficit ( . ± . mmol/l versus . ± . mmol/l), and in ph ( . ± . versus . ± . ) than patients assigned to standard therapy (p ≤ . for all comparisons). during the same period, the mean apache ii scores were significantly lower, indicating less severe organ dysfunction, in the patients assigned to egdt than in those patients assigned to standard therapy ( . ± . versus . ± . , p < . ). the protocol was based predominantly on guidelines published in by the society of critical care medicine [ ] . however, these guidelines have not been universally followed in clinical practice since their publication. an increasing number of critically ill patients are presenting to, and being treated in, emergency departments [ , ] . this is present-ing significant resource challenges in the emergency department environment. the inability to institute egdt may thus not be a conscious decision by the clinician not to follow the society of critical care medicine guidelines. emergency medicine in general may have to develop and formulate the cost-benefit analysis to support or implement such care in this environment in order to improve outcomes. there are sufficient evidence-based data to recommend that all patients with severe sepsis or septic shock should receive early and aggressive resuscitation based on this egdt protocol (see appendix ) . it is important that the interventions are individualized to each patient. a negative or positive value indicates how the control group therapy compares with the treatment group. a p < . , b p = . , c p = . , d p = . , e p = . . egdt, early goal-directed therapy. it is possible to identify patients with profound global myocardial dysfunction who are hence at risk of impaired perfusion. these patients, almost % of those in the egdt group, received dobutamine during the first hours because myocardial suppression was diagnosed. once myocardial dysfunction is corrected (and compliance improved), these patients become more suitable for volume loading, so this group received almost . liters more fluids in the first hours than the control patients. therefore, although vasopressor use was similar in the first hours, patients in the egdt group were more aggressively weaned off these agents during this period, resulting in fewer patients in this group entering the icu on vasopressors than in the control group. the lack of aggressive volume loading in the control group led to greater use of vasopressors in patients over the subsequent hours. in spite of more volume loading, the egdt group received less mechanical ventilation over the subsequent hours than in the standard treatment group. why was cardiovascular collapse a significant cause of death in the control group? cryptic shock (shock with normal vital signs) is a frequent occurrence in early severe sepsis and septic shock. despite resuscitation to the goals for mean arterial blood pressure and cvp, almost % of control patients continued to exhibit global tissue hypoxia (decreased scvo and increased lactate levels); in these patients, there was a twofold increase in hemodynamic deterioration, requiring more mechanical ventilation, pulmonary artery catheterization, and vasopressor use in the subsequent hours. how do severe sepsis and septic shock differ hemodynamically in the early stages compared with that classically described in the icu? patients presenting with early sepsis and septic shock are characterized by hypovolemia (low cvp), normal to increased blood pressures, and decreased cardiac output (decreased central venous oxygen saturation and low cardiac index). this is in contrast to icu patients who are euvolemic, have high scvo , and have elevated cardiac indices [ ] . what are the most important ways in which egdt can improve outcomes? the key factors are early detection of high-risk patients in cryptic shock, early reversal of hemodynamic perturbations and global tissue hypoxia, prevention of acute cardiovascular collapse, and the possibility of preventing the inflammatory aspects of global tissue hypoxia that accompany the inflammation or infection. severe sepsis and septic shock patients should receive early aggressive therapy to restore and maintain oxygen availability to the cells. there should also be generous use of fluids and inotropic agents titrated by appropriate hemodynamic monitoring. background a large number of observational studies have shown that patients with sepsis have severe depletion of protein c [ , ] . a number of studies have also shown the association of protein c depletion with high mortality in sepsis [ ] [ ] [ ] . furthermore, baboon studies have demonstrated that treatment with activated protein c prevents death from live escherichia coli infusions [ , ] . activated protein c exerts a number of actions. anticoagulant action includes the inactivation of coagulation factors va and viiia, and the inhibition of the formation of thrombin. profibrinolytic action allows the activity of tissue plasminogen activator (endogenous tissue plasminogen activator), by inactivating plasminogen activator inhibitor and thrombin activatable fibrinolysis inhibitor. finally, anti-inflammatory action reduces il- (in vivo) and proinflammatory cytokines (in vitro). the specific mechanisms by which drotrecogin alfa (activated) exerts its effect on survival in patients with severe sepsis are not completely understood. the efficacy of drotrecogin alfa (activated) (recombinant human activated protein c) in reducing mortality in patients with severe sepsis was investigated in a large multicenter, blinded, placebo-controlled, randomized, phase iii clinical trial, the protein c worldwide evaluation in severe sepsis (prowess) trial [ ] . all patients in the prowess trial received standard supportive care in addition to either drotrecogin alfa (activated) or placebo. for a summary of the protocol used in the prowess study, see appendix . the overall mortality in patients treated with drotrecogin alfa (activated) was . % compared with . % in patients receiving placebo, an absolute risk reduction of . % (p = . ) (see fig. ). the absolute risk reduction in patients with high risk of death defined by an apache ii score ≥ was . % (p < . ). the absolute risk reduction in patients with high risk of death defined by multiple organ failure was . % (p = . ). no substantial differences in drotrecogin alfa (activated) treatment effects were observed in subgroups defined by gender, ethnic origin, or infectious agent. can drotrecogin alfa (activated) be used in patients on dialysis for pre-existing renal failure, a category that was specifically excluded in the prowess trial? no pharmacokinetic data were available on drotrecogin alfa (activated) in patients on chronic dialysis when the prowess trial began, so such patients were excluded from the trial. subsequent research has shown that the pharmacokinetics of drotrecogin alfa (activated) are not substantially changed in patients on chronic dialysis. the design of the prowess trial allowed a maximum of hours between the onset of first organ dysfunction and the receipt of drotrecogin alfa (activated) (a -hour window was allowed for receipt of the drug following the first confirmation of first organ dysfunction, which in turn had to have been present for no more than hours). the treatment effect of drotrecogin alfa (activated) was consistent across all time intervals from meeting the entry criteria to the receipt of the study drug. treatment with drotrecogin alfa (activated) thus does not appear to be as time critical as interventions such as tissue plasminogen activator in stroke or myocardial infarction. because most of the experience with drotrecogin alfa (activated) was based on organ failure times less than hours, treatment should not be delayed when an appropriate candidate is identified. the time window employed in the prowess trial should allow a full history to be taken and other tests to be performed to determine the bleeding risk. as with all anticoagulants, drotrecogin alfa (activated) is associated with a risk of severe bleeding. during the infusion period in the prowess trial, the bleeding rates were . % in the drotrecogin alfa (activated) group versus . % in the placebo group (p = . ). the risk of bleeding was fairly constant across most subgroups. however, severe thrombocytopenia (< , /mm ) was commonly associated with serious bleeding and intracerebral hemorrhage. patients at high risk of death in the prowess trial were most likely to benefit from drotrecogin alfa (activated). in the prowess trial, the apache ii score was the most effective predictor of risk of death and likelihood of benefit from drotrecogin alfa (activated), particularly in those patients with an apache ii score ≥ . in the prowess trial, the number of organ dysfunctions was also an important indicator that supported an association between likelihood of benefit from drotrecogin alfa (activated) and risk of death. two or more organ dysfunctions identify a population that responds well to therapy, and is a practical measurement. the panel believes that acute respiratory failure or hypotension unresponsive to fluid challenge should suggest the use of drotrecogin alfa (activated). however, coagulopathy, a platelet count < , /mm , acidosis, or low urine output alone should not suggest its use. a very large international study of , patients will be started in late to investigate the efficacy of drotrecogin alfa (activated) in patients with a single organ failure and/or apache scores < . the decision on whether to administer the drug should ultimately depend on whether the patient meets the selection criteria. a patient presenting in the emergency room with acute respiratory failure or acute cardiovascular decompensation should receive appropriate treatment there. the drawback to treatment in the emergency room is that there may not be sufficient time in which to evaluate the patient's bleeding risks. delaying treatment for a few hours will enable more tests to be performed and a fuller history to be taken, both of which will provide a better indication of whether drotrecogin alfa (activated) is appropriate. the dose is always the same ( µg/kg/hour), regardless of the type of organ failure or the degree of sepsis severity. in addition, the -hour window of treatment is always the same so that interruptions of treatment are made up at the end to maintain a total of hours of treatment. twenty-eight-day survival in patients treated with drotrecogin alfa (activated) or placebo: all-cause mortality. do patients require any laboratory testing before they receive drotrecogin alfa (activated)? no laboratory testing was carried out in the prowess trial, and subgroup analysis identified no biochemical marker that conclusively indicates treatment. for example, treatment-associated reductions in mortality were observed in patients with normal protein c levels and in those with low protein c levels. clinical criteria are recommended for the initiation of therapy. aspirin ( mg/day) was allowed in the prowess trial. patients on glycoprotein iib/iiia inhibitors were excluded because no data were available regarding drug interactions and pharmacokinetics. use of these types of agents is likely to increase the risk of bleeding with drotrecogin alfa (activated) therapy. the anticipated benefits must therefore be weighed against the potential risks. in the prowess trial, efforts were made to correct the international normalized ratio towards normal if it was greater than at any time during infusion of drotrecogin alfa (activated). approximately one-third of patients in the prowess trial received steroids at the same time as drotrecogin alfa (activated). there was no interaction with steroid use, presumably because the mechanism of action of steroids is so different from that of activated protein c. hence, steroids should be used if they are needed, and if the patient qualifies for drotrecogin alfa (activated) the two should be used together. drotrecogin alfa (activated) should be considered for use in all adult patients with recent onset severe sepsis or septic shock, and a high risk of death. the value of steroids in the treatment of patients with severe sepsis and septic shock has been fiercely debated for some time. although a number of well-designed, randomized, controlled trials failed to show any benefits of steroid therapy in terms of improved survival in patients with severe sepsis (reviewed in [ , ] ), with mortality increased in many as a result of an increased incidence of nosocomial infections, these trials were primarily investigating the efficacy of short courses of high-dose steroids. the question of whether lower doses of steroids may provide benefit in these patients has only recently been addressed. there is a relatively strong rationale for considering the use of steroids in patients with refractory septic shock. relative adrenal insufficiency is common in patients with refractory septic shock ( - % of patients) [ ] . in addition to such relative adrenal insufficiency and the blunted response to corticotrophin, a large body of evidence indicates that sepsis and refractory septic shock are characterized by peripheral tissue resistance to corticosteroids [ , ] . in septic patients, this can be evidenced in a variety of ways. first, global cortisol binding, which carries cortisol from the adrenal glands to the tissues, decreases in patients with severe sepsis [ ] . second, the number and binding affinity of glucocorticosteroid receptors may be reduced in patients with sepsis and severe sepsis [ ] , leading to a decrease in the conversion of cortisone to its active form, cortisol, particularly by il- levels in the tissues. finally, data have been published demonstrating that moderate doses of steroids may restore cell sensitivity to vasopressors [ ] . this may reduce the intensity of the inflammatory response and decrease organ dysfunction. low-dose steroid treatment is also well tolerated [ ] . this body of evidence prompted the initiation of a phase iii randomized, controlled trial performed in centers in france with patients [ ] . the aim of the trial was to determine whether moderate-dose corticosteroid therapy affected survival in patients with refractory septic shock and adrenal insufficiency. all patients had to be treated with vasopressor agents and mechanical ventilation. for a summary of the protocol used in this study, see appendix . patients were stratified according to their response to the adrenocorticotrophic hormone (acth) test. nonresponders were defined by an increment in cortisol levels < µg/dl or < nm/l after challenge with µg cosyntropin. of the patients included, there were nonresponders to the corticotropin test (placebo, patients; steroids, patients). a significant survival benefit was demonstrated among nonresponders receiving moderate-dose corticosteroids. there were deaths in the placebo group ( %) and deaths in the steroid group ( %) (hazard ratio, . ; % confidence interval, . - . ; p = . ). no beneficial effects were observed in the subset of patients who were classified as responders. hence, in this paradigm, the acth test serves as a useful prognostic measure. since a beneficial effect was observed in the total population, however, the need for an acth test can be challenged and further studies are required. if an acth test is performed, corticosteroid administration can be started before results are received. moderate-dose corticosteroids should be administered to patients with established refractory septic shock. what is the optimal dose for this intervention? hydrocortisone should be given daily at a dose of - mg. fludrocortisone should be given daily at a dose of µg. what is the optimal duration for this intervention? moderate doses of steroids should be given for days. hydrocortisone can be administered as serial boluses or as a continuous infusion. it may be that rebound phenomena at treatment discontinuation are more frequent when hydrocortisone is given as a continuous infusion. in addition, in the phase iii randomized trial, hydrocortisone was given as serial boluses. the phase iii randomized trial has shown that the combination of hydrocortisone and fludrocortisone increased survival. in addition, sepsis is more frequently associated with a mineralocorticoid deficiency than a glucocorticoid deficiency. hence, fludrocortisone should be added to hydrocortisone. administration of moderate-dose corticosteroids should be considered in cases of refractory septic shock, particularly in those with relative adrenal insufficiency. it is recommended that an acth test be carried out before starting the intervention. hyperglycemia, caused by insulin resistance in the liver and muscle, is a common finding in icu patients. it can be considered an adaptive response, providing glucose for the brain, red cells, and wound healing, and is generally only treated when blood glucose increases to > mg/dl (> mmol/l). previous studies have shown that high levels of insulin-like growth factor binding protein (a very good marker of lack of hepatic insulin effect) predict mortality [ , ] . patients with high insulin-like growth factor binding protein also tend to have the lowest insulin levels, indicating that beta cell function is impaired and, therefore, not enough insulin is being produced. these results indicate that hyperglycemia may not always be adaptive and that it should be treated to avoid the onset of specific complications. nevertheless, conventional wisdom in the icu has been that hyperglycemia is beneficial and that hypoglycemia should be avoided. the hypothesis that hyperglycemia (> mg/dl, > . mmol/l) predisposes to specific icu complications, prolonged intensive care dependency and death was tested in a prospective, randomized, controlled trial [ ] . for a summary of the protocol used in this study, see appendix . thirty-five of the patients ( . %) in the intensive insulin group died in the icu, compared with patients ( . %) in the conventional therapy group. for further mortality data on both the length of hospital stay and the cause of death, see tables and . for morbidity data, see figure . tight control of blood sugar, as outlined in appendix , requires a strict protocol for insulin administration and repeated determination of blood sugar. this is yet to be proven, and is the subject of an ongoing study. because medical patients tend to stay in the icu longer than surgical patients, the results from this study indicate that this intervention would be even more favorable to medical icu patients. however, one needs to be careful with application of the algorithm in certain disease states, especially severe hepatic dysfunction and renal failure. no, all carbohydrates are included. see appendix for guidelines on feeding. the level was chosen because it is in the physiologic range for healthy people. as well as its effect on glycemia, insulin has been shown to inhibit tnf-α and macrophage inhibitory factor (when infused concomitantly with glucose). this has led to some doubts as to whether the effect in this study was due to normalization of blood glucose levels. however, multivariate analysis of all the risk factors for mortality, including severity of illness on admission, indicated that blood glucose determines the outcome; there was a % increase in risk of death per mg/dl increase in blood glucose. it is not yet possible to determine this. although it was blood glucose levels that were measured, the effects of insulin may in fact be on free fatty acids, as they change in parallel with s blood glucose. one of the key mechanisms may be prevention of hypertriglyceridemia and high concentrations of free fatty acids. it is strongly advisable to tightly control blood sugar close to physiologic levels, especially in surgical patients. implemen-tation of this recommendation requires a well-defined icu protocol. the interventions discussed in the present article have been applied in different patient populations and at different times in the course of the disease (see table ). it is essential for physicians to understand that these therapies are not mutually exclusive. optimal patient management may require a combination of approaches: mechanical ventilation to preserve lung function, hemodynamic support to maintain adequate scvo , intensive insulin therapy to normalize blood sugar, steroids to provide adequate immunosuppression, and drotrecogin alfa (activated) to prevent the systemic coagulopathy characteristic of severe sepsis and, hence, to preserve organ function. a sound understanding of the indications and contraindications of these interventions will guide appropriate intervention. similarly, the timing of therapy needs to be closely monitored. education in the signs and symptoms of sepsis and severe sepsis should prompt early initiation of therapy. many of the interventions discussed in this article were tested at specific available online http://ccforum.com/content/ /s /s multiple organ failure, no sepsis focus multiple organ failure, with sepsis focus most important effects on morbidity [ ] . cvvh, continuous venovenous hemofiltration; icu, intensive care unit; nnt, number needed to treat; rrr, relative risk reduction. despite the wealth of data to support the approaches discussed, it is clear that uptake of these interventions into clinical practice has been slow. although there may be practical reasons for this, it would appear in many cases to involve either unfamiliarity with the data or a reluctance, or at least inertia, to change established practices (witness the necessity of proving that hypoglycemia is beneficial in icu patients despite no good evidence to the contrary). the icu has changed in the past years; there are more tools to use and more interventions to implement. despite application of new methods, however, outcomes have changed very little and certainly not in proportion to the changes that were expected based on the results from clinical trials. efficient integration of new interventions into the wider icu population is clearly essential. the panel believes that optimal use of existing therapies and the integration of proven new therapies will reduce mortality rates. further positive results from new trials with improved trial designs should encourage intensivists to incorporate new interventions into their practice. protocols are essential to ensure efficient integration of new therapies and to improve outcomes on the wards. morris predicted in a recent paper that an increase in compliance with evidence-based recommendations through the use of protocols would decrease error and would enhance patient safety [ ] . however, a complete treatment protocol is only effective when each ward (inside and outside of the icu) has the trained staff to implement it, and when a skilled intensive care physician is available to lead the team. training and education of staff is essential. all five of the interventions discussed in this article have generated convincing evidence for their use, and they hold out hope for reducing mortality in patients with sepsis, severe sepsis and septic shock. yet, despite compelling data, the application of these interventions has yet to become routine practice in most icus. it is our hope that this article will enable physicians to understand how best to apply these therapies in clinical practice; from appropriate patient selection and timing of therapy, to combining different approaches for optimal patient management. a willingness to embrace new interventions, coupled with the development and implementation of rigorous protocols to ensure appropriate use, will improve outcomes and lead to a substantial reduction in mortality in these patients. • a respiratory rate ≥ breaths/min or a partial pressure of arterial carbon dioxide ≤ mmhg, or the use of mechanical ventilation for an acute respiratory process. • a white cell count ≥ , /mm or ≤ /mm , or a differential count showing > % immature neutrophils. patients should meet at least one of the following five criteria: • pregnancy or breastfeeding. • aged younger than years or weight > kg. • platelet count < , /mm . • conditions that increase the risk of bleeding: • surgery requiring general or spinal anesthesia within hours before the infusion, the potential need for such surgery during the infusion, or evidence of active bleeding postoperatively; • a history of severe head trauma requiring hospitalization, intracranial surgery, or stroke within months before the study, or any history of intracerebral arteriovenous malformation, cerebral aneurysm, or mass lesions of the central nervous system; • a history of congenital bleeding diatheses; gastrointestinal bleeding within weeks before the study unless corrective surgery had been performed; or • trauma considered to increase the risk of bleeding. • a known hypercoagualable condition including: • resistance to activated protein c; • hereditary deficiency of protein c, protein s, or antithrombin iii; • presence of anticardiolipin antibody, antiphospholipid antibody, lupus anticoagulant, or homocysteinemia; or • recently documented (within months) or highly suspected deep-vein thrombosis or pulmonary embolism. • patient's family or physician, or both, not in favor of aggressive treatment of the patient, or the presence of an advanced directive to withhold life-sustaining treatment. • patient not expected to survive days because of an uncorrectable medical condition, such as poorly controlled neoplasm or other end-stage disease. • moribund state in which death is perceived to be imminent. • human immunodeficiency virus infection in association with a last known cd cell count ≤ /mm . • history of bone marrow, lung, liver, pancreas, or smallbowel transplantation. • chronic renal failure requiring hemodialysis or peritoneal dialysis (acute renal failure was not an exclusion criterion). • known or suspected portosystemic hypertension, chronic jaundice, cirrhosis, or chronic ascites. • acute pancreatitis with no established source of infection. • participation in an investigational study within days before treatment. • use of any of the following medications or treatment regimens: • unfractionated heparin to treat an active thrombotic event within hours before the infusion (prophylactic treatment with a dose of unfractionated heparin of up to , u/day was permitted); • low molecular weight heparin at a higher dose than recommended for prophylactic use (as specified in the package insert) within hours before the infusion; • warfarin (if used within days before study entry and if the prothrombin time exceeded the upper limit of the normal range for the institution); • acetylsalicylic acid at a dose of more than mg/day within days before the study; • thrombolytic therapy within days before the study (thrombolytic agents permitted for the treatment of thromboses within a catheter); • glycoprotein iib/iiia antagonists within days before study entry; • antithrombin iii at a dose of more than , u within hours before the study; • protein c within hours before the study. drotrecogin alfa (activated) should be given at a dose of µg/kg/hour for hours. infusion should be interrupted hour prior to any percutaneous procedure or major surgery, and should be resumed and hours later, respectively, in the absence of bleeding complications. there was an -hour time window from shock onset to check for eligibility and to perform a short acth test (blood samples before and and min after a µg intravenous bolus of tetracosactrin). patients were then randomly assigned to receive mg hydrocortisone as an intravenous bolus every hours and one µg tablet of fludrocortisone through a nasogastric tube once a day, or their respective placebos. treatments were given for days, and patients were followed up for year. on admission, patients should receive continuous intravenous glucose ( - g over hours). after hours, total parenteral, combined parenteral and enteral, or total enteral feeding should be instituted: - nonprotein kcal/kg/day with a balanced composition ( . - . g nitrogen/kg/day and - % nonprotein calories in the form of lipids). total enteral feeding should be attempted as early as possible. national vital statistics reports epidemiology of severe sepsis in the united states: analysis of incidence, outcome, and associated costs of care the task force on the management of acute myocardial infarction of the european society of cardiology epidemiology of sepsis: an update severe sepsis and septic shock. definitions, epidemiology, and clinical manifestations gram-negative sepsis: a dilemma of modern medicine sepsis definitions. lancet infect dis use of the sofa score to assess the incidence of organ dysfunction/failure in intensive care units: results of a multicenter, prospective study improved survival of patients with acute respiratory distress syndrome (ards): - [concepts in emergency and critical care treatment of gram-negative bacteremia and septic shock with ha- a human monoclonal antibody against endotoxin. a randomized, double-blind, placebo-controlled trial treatment of gram-negative bacteremia and shock with human antiserum to a mutant escherichia coli the tnfα α mab sepsis study group: monoclonal antibody to human tumor necrosis factor alpha (tnfα α mab): efficacy and safety in patients with the sepsis syndrome lenercept study group: lenercept (p -tumor necrosis factor receptor fusion protein, ro - , tenefuse) patients with severe sepsis or early septic shock. a randomized double-blind placebo-controlled multicenter phase iii trial with patients recombinant human protein c worldwide evaluation in severe sepsis (prowess) study group: efficacy and safety of recombinant human activated protein c for severe sepsis current concepts: treating patients with severe sepsis need for intensivists in intensive care units randomised trial of intravenous streptokinase, oral aspirin, both, or neither among , cases of suspected acute myocardial infarction: isis- (second international study of infarct survival) collaborative group injurious ventilatory strategies increase cytokines and c-fos m-rna expression in an isolated rat lung model effect of mechanical ventilation on inflammatory mediators in patients with acute respiratory distress syndrome: a randomized controlled trial effect of a protective-ventilation strategy on mortality in the acute respiratory distress syndrome ventilation with lower tidal volumes as compared with traditional tidal volumes for acute lung injury and the acute respiratory distress syndrome. the acute respiratory distress syndrome network airway and lung in sepsis elevation of systemic oxygen delivery in the treatment of critically ill patients a trial of goal-oriented hemodynamic therapy in critically ill patients. svo collaborative group early goal-directed therapy collaborative group: early goal-directed therapy in the treatment of severe sepsis and septic shock task force of the american college of critical care medicine, society of critical care medicine: practice parameters for hemodynamic support of sepsis in adult patients in sepsis critical care in the emergency department: a physiologic assessment and outcome evaluation critical care provided in an urban emergency department a hemodynamic comparison of early and late phase severe sepsis and septic shock low levels of protein c are associated with poor outcomes in severe sepsis protein c, protein s, c b-binding protein in severe infection and septic shock prognostic value of protein c concentrations in neutropenic patients at high risk of severe septic complications van der voort e: protein c and s deficiency in severe infectious purpura of children: a collaborative study of cases. intensive care med epidemic meningioccemia and purpura fulminans with induced protein c deficiency protein c prevents the coagulopathic and lethal effects of escherichia coli infusion in the baboon the endothelial cell protein c receptor aids in host defense against escherichia coli sepsis corticosteroid treatment for sepsis: a critical appraisal and meta-analysis of the literature steroid controversy in sepsis and septic shock: a meta-analysis a -level prognostic classification in septic shock based on cortisol levels and cortisol response to corticotropin stress doses of hydrocortisone reverse hyperdynamic septic shock: a prospective, randomized, double-blinded, singlecenter study prolonged methylprednisolone treatment suppresses systemic inflammation in patients with unresolving acute respiratory distress syndrome: evidence for inadequate endogenous glucocorticoid secretion and inflammation-induced immune cell resistance to glucocorticoids patterns of corticosteroidbinding globulin and the free cortisol index during septic shock and multitrauma adrenal insufficiency during the late stage of polymicrobial sepsis reversal of late septic shock with supraphysiologic doses of hydrocortisone effect of a treatment with low doses of hydrocortisone and fludrocortisone on mortality in patients with septic shock reactivation of pituitary hormone release and metabolic improvement by infusion of growth hormone-releasing peptide and thyrotropin-releasing hormone in patients with protracted critical illness a paradoxical gender dissociation within the growth hormone/insulin-like growth factor i axis during protracted critical illness bouillon r: intensive insulin therapy in the critically ill patients decision support and safety of clinical environments. qual saf health care the roundtable discussion was supported by an unrestricted educational grant from eli lilly and company. jlv, ea, gb and er are consultants to eli lilly and company. all authors received an honorarium/grant for participating in this meeting. inclusion criteria • partial pressure of arterial oxygen/fio ≤ mmhg. • bilateral infiltrates consistent with pulmonary edema on frontal chest radiograph.• no clinical evidence of left atrial hypertension, pulmonary artery wedge pressure ≤ mmhg if measured. • positive pressure ventilation via endotracheal tube. inclusion criteria patients must have a known infection or a suspected infection, as evidenced by one or more of the following:• white cells in a normally sterile body fluid.• perforated viscus.• radiographic evidence of pneumonia in association with the production of purulent sputum. • a syndrome associated with a high risk of infection (e.g. ascending cholangitis). patients should meet at least three of the following four criteria:• a core temperature ≥ °c ( . °f) or ≤ °c ( . °f).• a heart rate ≥ beats/min, except in patients with a medical condition known to increase the heart rate or those receiving treatment that would prevent tachycardia.available online http://ccforum.com/content/ /s /s the study included all mechanically ventilated patients entering the icu: predominantly surgical patients, with some neu-rological patients (the icu in which the trial took place also sees such patients). medical icu patients (e.g. those with chronic obstructive pulmonary disease or oncologic or hematological disorders) were not included as they are not treated in the unit where the study was conducted. however, septic patients that were initially surgical but then came back from the ward with sepsis were included.only those patients who were moribund or had do-notresuscitate status at icu admission were excluded from the trial. if blood glucose ≥ mg/dl (≥ . mmol/l), infuse with insulin to maintain normoglycemia ( - mg/dl, . - . mmol/l). do not exceed iu/hour. adjust insulin dose based on measurements of whole-blood glucose in undiluted arterial blood, performed at - hour intervals, based on the following algorithm: adjust the dose in proportion to the observed change in blood glucose level (if blood glucose decreases by % then the insulin dose should be decreased by % and checked within the next hour). appendix table provides information on the appropriate action depending on the blood glucose level. the numerical instructions provided in appendix table are a guide; insulin dosage should always be done with common sense, proportionate to the previous changes in blood glucose observed upon previous changes in dosage.available online http://ccforum.com/content/ /s /s appendix table appropriate action depending on blood glucose level key: cord- -v hh w r authors: leung, c.w.; chiu, w.k. title: clinical picture, diagnosis, treatment and outcome of severe acute respiratory syndrome (sars) in children date: - - journal: paediatr respir rev doi: . /j.prrv. . . sha: doc_id: cord_uid: v hh w r children are susceptible to infection by sars-associated coronavirus (sars-cov) but the clinical picture of sars is milder than in adults. teenagers resemble adults in presentation and disease progression and may develop severe illness requiring intensive care and assisted ventilation. fever, malaise, cough, coryza, chills or rigor, sputum production, headache, myalgia, leucopaenia, lymphopaenia, thrombocytopaenia, mildly prolonged activated partial thromboplastin times and elevated lactate dehydrogenase levels are common presenting features. radiographic findings are non-specific but high-resolution computed tomography of the thorax in clinically suspected cases may be an early diagnostic aid when initial chest radiographs appear normal. the improved reverse transcription-polymerase chain reaction (rt-pcr) assays are critical in the early diagnosis of sars, with sensitivity approaching % in the first days of illness when performed on nasopharyngeal aspirates, the preferred specimens. absence of seroconversion to sars-cov beyond days from disease onset generally excludes the diagnosis. the best treatment strategy for sars among children remains to be determined. no case fatality has been reported in children and the short- to medium-term outcome appears to be good. the importance of continued monitoring for any long-term complications due to the disease or its empiric treatment, cannot be overemphasised. severe acute respiratory syndrome (sars), a newly described infectious disease caused by the novel sarsassociated coronavirus (sars-cov), has become a major threat to public health globally. - sars is highly contagious and has been aptly coined 'the first plague of the twenty-first century'. the disease is characterised by transmission in healthcare and household settings and through intriguing superspreading events which were pivotal in its global spread. [ ] [ ] [ ] [ ] [ ] [ ] [ ] superspreading events including a major hospital outbreak, in-flight transmission on board commercial paediatric respiratory reviews ( ) summary children are susceptible to infection by sars-associated coronavirus (sars-cov) but the clinical picture of sars is milder than in adults. teenagers resemble adults in presentation and disease progression and may develop severe illness requiring intensive care and assisted ventilation. fever, malaise, cough, coryza, chills or rigor, sputum production, headache, myalgia, leucopaenia, lymphopaenia, thrombocytopaenia, mildly prolonged activated partial thromboplastin times and elevated lactate dehydrogenase levels are common presenting features. radiographic findings are non-specific but highresolution computed tomography of the thorax in clinically suspected cases may be an early diagnostic aid when initial chest radiographs appear normal. the improved reverse transcription-polymerase chain reaction (rt-pcr) assays are critical in the early diagnosis of sars, with sensitivity approaching % in the first days of illness when performed on nasopharyngeal aspirates, the preferred specimens. absence of seroconversion to sars-cov beyond days from disease onset generally excludes the diagnosis. the best treatment strategy for sars among children remains to be determined. no case fatality has been reported in children and the short-to medium-term outcome appears to be good. the importance of continued monitoring for any long-term complications due to the disease or its empiric treatment, cannot be overemphasised. ß elsevier ltd. all rights reserved. abbreviations: sars, severe acute respiratory syndrome; sars-cov, sars-associated coronavirus; rsv, respiratory syncytial virus; ards, acute respiratory distress syndrome; cxr, chest radiograph; hrct, high-resolution computed tomography; boop, bronchiolitis obliterans-organising pneumonia; npa, nasopharyngeal aspirate; rt-pcr, reverse transcription-polymerase chain reaction; ifa, immunofluorescence assay; elisa, enzyme-linked immunosorbant assay. *correspondence to: c.w. leung; e-mail: leungcw@ha.org.hk. airliners, transmission in a hotel and a large-scale community outbreak in a densely populated residential complex, primarily resulting from environmental contamination by a 'superspreader' with diarrhoea, were well described. , , [ ] [ ] [ ] [ ] the disease first started as a mysterious outbreak of atypical pneumonia in the guangdong province of southern china in november . by july , , up to countries and regions of the world had been affected by sars. a worldwide total of cases of probable sars, ( %) of these being healthcare workers and deaths ( . %) were recorded. in hong kong, the toll was affected individuals, including ( %) healthcare workers and deaths ( %). the subsequent reemergence of the first six sporadic cases of sars, two of which were probably laboratory-acquired, did not result in local transmission in singapore, taiwan and china. [ ] [ ] [ ] [ ] children appeared to be less affected by the disease, with smaller case numbers and less severe illness reported. [ ] [ ] [ ] [ ] all age groups are susceptible to sars-cov, which is new to humans. however, rapid isolation of diseased adults, whose infectivity is lower in the first few days of illness, has contributed to reduced frequency of household exposure for children. the exact number of children affected by sars worldwide is unknown as the age breakdown of reported cases was not available or incomplete for some of the affected countries (who sars surveillance team, personal communication). it is estimated that children < years of age only accounted for about % of the total affected. there was no reported mortality in children (who sars surveillance team, personal communication). a total of children aged < years were registered in the e-sars database of the hospital authority of hong kong, accounting for % of all patients notified. the crude age-specific attack rate for children in hong kong was . per persons < years of age. serologic confirmation of sars was documented in children ( . per persons < years of age). sixty-four children with clinical disease and seroconversion to sars-cov were managed in the authors' hospitals. the experience with this cohort of laboratory-confirmed patients forms the basis of the clinical information presented in this review. , most children reported worldwide were previously healthy and there was no sex predominance. thirty-five ( %) of the children managed by the authors were girls. the male to female ratio was : . . their mean and median ages were . and years, respectively. the youngest patient was a -day-old premature infant, which is the youngest case reported to date. comorbidity was only present in children ( %) but none of them were immunocompromised. an epidemiologic link was available in the vast majority of children with sars, which appeared to be the most important clue leading to diagnosis in an epidemic situation. worldwide, children were usually secondary household contacts of affected adults, some of whom were healthcare workers or international travellers returning from areas with local transmission of sars. transmission among children or from children to adult contacts was uncommon. about % of serologically confirmed children in hong kong were victims of a point source community outbreak due to environmental contamination. the actual proportion of children being secondary household contacts in the particular outbreak could not be determined given the short incubation period between exposure, either to a common environmental source or an index household member, and presentation. there is no published report on the differences in susceptibility and communicability between children and adults. any apparent difference might be related to different risks of exposure for the two age groups. sars is largely an atypical pneumonia with minimal or no extrapulmonary manifestation, apart from diarrhoea. cellular tropism of the sars-cov has been demonstrated primarily in pneumocytes and surface enterocytes of the small bowel. the clinical presentation of sars is nonspecific, with features overlapping those of atypical pneumonia caused by other respiratory pathogens such as influenza virus (including highly pathogenic avian influenza viruses), parainfluenza virus, adenovirus, respiratory syncytial virus (rsv), mycoplasma pneumoniae, chlamydia pneumoniae, chlamydia psittaci and legionella pneumophila. the clinical course of sars in adult patients is well described and appears to follow a triphasic pattern. , [ ] [ ] [ ] [ ] [ ] following an incubation period of - days (mean . days, % ci . to . ), adults present with a prodrome characterised by high fever (temperature > c), chills or rigor, malaise, headache, dizziness and myalgia. upper respiratory symptoms such as coryza and sore throat are mild and uncommon. diarrhoea is a presenting feature in - % of adult patients. , , after - days the disease progresses to involve the lower respiratory tract and a dry, non-productive cough or dyspnoea becomes prominent. in - % of cases, progression to acute respiratory distress syndrome (ards) necessitating intubation and assisted ventilation is observed. mortality results primarily from respiratory failure and a significant proportion of patients recover from pulmonary destruction over an extended period. sars appears to run a less aggressive clinical course in children compared with adults. the severity of illness varies and the extent of asymptomatic infection is unknown, although it is believed to be uncommon. children are usually hospitalised - days after the onset of symptoms. in one paediatric case series, the mean duration of fever before admission was . ae . days (median , range - ). the most common presenting clinical features in children include fever, malaise, cough, coryza, chills or rigor, sputum production, headache and myalgia (table ) . , lethargy, poor feeding or anorexia, nausea, vomiting, diarrhoea, abdominal pain, sore throat, dyspnoea and dizziness are less commonly encountered. less than % of children may pass loose to watery stools, but profuse diarrhoea is rare throughout the course of illness. blood and mucus in the stool, features suggestive of inflammatory enterocolitis, have not been reported. cough, predominantly unproductive in nature, is only found in just over half of the children at presentation. definite physical signs of consolidation are hardly evident and crepitations (crackles) on chest auscultation are unusual despite prominent radiographic evidence of pulmonary infiltrates, even in patients who develop respiratory distress, hence the description of 'atypical' pneumonia. lymphadenopathy, hepatosplenomegaly or clinical bleeding is absent. skin rash is an exceedingly rare manifestation. hypoxaemia is seldom noted at presentation and generally develops towards the end of the first week or the beginning of the second week of illness in severe cases. the youngest patient, however, presented with a cyanotic attack, dyspnoea, cough and hypothermia with subsequent development of fever. teenagers (aged > years) may resemble adults in presentation and disease progression. they tend to have more constitutional upsets and systemic symptoms of malaise, chills or rigor, headache, myalgia and dizziness are significantly more common ( table ) . they appear sicker, have a greater need for oxygen therapy and other respiratory support and may require intensive care. children years of age generally have milder symptoms and coryza is significantly more common ( table ) . they appear to run a milder and shorter clinical course. the clinical picture is sometimes indistinguishable from other viral infections of the upper respiratory tract, thus posing a diagnostic challenge. the clinical course of sars in the majority of children follows a biphasic pattern. the phase of viral replication, which lasts for a few days, is characterised by an abrupt onset of fever and constitutional symptoms in association with an increase in body viral load. the phase of immunopathologic damage is marked by the progression of pneumonia and hypoxaemia, when the body viral load declines and an exaggerated host immune response supervenes. the prodromal and pneumonic phases of the disease, however, may be less distinct in comparison with adult patients. progression to ards, or the third phase as in adults, is only seen in a very small number of children, predominantly adolescents. the natural history of untreated sars in both adults and children remains unclear. as most patients worldwide had received some form of empiric treatment in the form of antiviral agents with or without corticosteroids, the probability of spontaneous recovery could not be ascertained. nevertheless, three children with mild disease had recovered on supportive therapy alone in the authors' cohort. , anecdotal reports of extrapulmonary manifestations of sars, in the form of central nervous system dysfunction and probable viral hepatitis, have been described in adults. [ ] [ ] [ ] atypical presentation of sars, in the form of non-specific febrile illness or febrile non-pneumonic respiratory illness, have been observed in both children and adults. , , such cases are likely to evade clinical detection in the absence of a definite contact history with patients with suspected or confirmed sars. the full spectrum of clinical as well as subclinical illnesses caused by infection with sars-cov will unfold with further epidemiological studies and case reports. as sars is basically a pneumonic infection, chest radiograph (cxr) is therefore an essential diagnostic tool. the principal radiographic abnormality of sars in children is illdefined airspace shadowing, which presents as ground-glass opacities and/or unifocal, lobar or multifocal areas of consolidation. [ ] [ ] [ ] [ ] , unilateral focal opacity was reported as the most common finding in one paediatric case series and was evident in % of children at presentation (fig. ). in adults, regions of airspace disease predominate in the lower lobes but are also noted elsewhere. there appears to be no predominant distribution pattern of consolidation in children. [ ] [ ] [ ] cxr opacities are most often peripheral or mixed central and peripheral in location. the lung opacities show a tendency to progress, with increase in size or involvement of multiple areas either unilaterally or bilaterally in moderate to severe cases. rapid progression to unilateral multifocal or bilateral involvement, with reduction in lung volumes in the second week of illness, is typical in children who develop severe hypoxaemia (fig. ) . in the advanced stage of the disease, which only occurs in a very small number of children, widespread ground-glass opacities and diffuse patchy consolidations are seen, likely representing progression to ards. pneumonic changes may not be apparent at presentation as mildly symptomatic individuals may be identified early in the prodromal period through contact tracing of patients diagnosed with sars. repeat cxr examination, as guided by failure of resolution of symptoms or change in clinical condition, will clarify the picture by revealing new pulmonary infiltrates as the disease progresses. frequent monitoring of cxr changes has the additional benefit of detecting early radiographic deterioration in many patients, heralding clinical deterioration. radiographic resolution, on the other hand, generally lags behind clinical improvement. complete resolution of the airspace opacities can take more than a month in the most severely affected children. no preliminary evidence of pulmonary fibrosis, bronchial wall thickening, bronchiectasis or lung volume loss was observed on follow-up in one paediatric case series. viral pneumonias tend to show reticulo-nodularity as well as a symmetrical perihilar peribronchial pattern of infiltration which is sometimes marked by hilar adenopathy. in contrast to pneumonias caused by other respiratory pathogens, the cxr of children with sars shows no evidence of interstitial disease, hilar adenopathy, mediastinal widening, significant pleural effusion, cavitation, abscess formation, pneumatocele, pneumothorax or pneumomediastinum. [ ] [ ] [ ] [ ] , nevertheless, the radiographic features of sars in children are non-specific. radiological differentiation of sars from other commonly encountered childhood respiratory illnesses causing airspace disease can be difficult. high-resolution computed tomography (hrct) of the chest has been used as an early diagnostic tool in clinically suspected children with initial negative or equivocal chest radiographs. [ ] [ ] [ ] [ ] hrct findings may include groundglass opacification, unifocal or multifocal consolidation in subpleural, peripheral or central regions and interlobular septal and intralobular interstitial thickening (fig. ) . the characteristic peripheral alveolar opacities are reminiscent of bronchiolitis obliterans-organising pneumonia (boop). , , in general, hrct is sensitive in detecting more extensive airspace consolidation and ground-glass attenuation than cxr. the investigation is particularly useful when lung parenchymal abnormalities are minimal early in the course of illness, or being obscured by the diaphragm and the cardiac silhouette. the utility of chest hrct lies in the early confirmation of airspace disease in radiographically inapparent cases with a strong contact history and clinical features highly suspicious of sars, thus allowing prompt isolation and monitoring for clinical and radiological deterioration. the haematological and biochemical abnormalities of sars in children are neither diagnostic nor specific. like adults, the most consistent haematological finding is lymphopaenia, which is present in about % of children at presentation and about % during the course of illness. , depletion of lymphocytes may be secondary to the direct cytopathic effect of the virus, cytokine-mediated apoptosis, lymphocyte margination due to increased cortisol secretion from activation of the hypothalamic-pituitary-adrenal axis or the administration of high-dose glucocorticoids, which have a profound lympholytic effect, especially on t lymphocytes. [ ] [ ] [ ] other haematological abnormalities such as leucopaenia, thrombocytopaenia and mildly prolonged activated partial thromboplastin times are observed in about % of children. anaemia is rarely found at presentation and is only detected in < % of children. , unlike adults, a significant drop in the haemoglobin level during the course of illness that necessitates discontinuation of empiric antiviral therapy, namely ribavirin, has not been observed. [ ] [ ] [ ] reactive thrombocytosis on recovery from sars is significantly more common in children years of age. this phenomenon is sometimes observed in children recovering from systemic viral infections and is probably not related to the use of corticosteroids. despite an abnormal clotting profile with elevated d-dimer levels and the detection of lupus anticoagulants in a small number of children, bleeding events or thrombotic complications have not been reported. , the most common biochemical abnormality in children with sars is an elevated lactate dehydrogenase level, which is present in about % at presentation and about % during the course of illness. elevated alanine aminotransferase levels are seen in < % of children at presentation and < % during the course of illness. elevation of creatine kinase levels vary from % to % between case series. , teenage patients tend to have more derangement of laboratory parameters and they may take longer to resolve. similar to human infection with avian influenza a h n virus, cytokine dysregulation is believed to be pivotal in the immunopathogenesis of sars among adults and children. serial monitoring of the plasma inflammatory cytokine profile using flow cytometry in a cohort of eight paediatric patients suggests that the caspase- -dependent pathway in infected macrophages is selectively activated, as reflected by substantial elevation of circulating interleukin- b levels. conversely, interleukin- and tumour necrosis factor-a levels, which are markedly increased in human infection with avian influenza a h n virus, are not overtly elevated throughout the course of illness. , the predominant activation of the th immune response facilitates viral clearance and may explain the rapid recovery of children. as sars is a newly emerging infectious disease with unknown aetiology initially, the initial case definitions of suspected and probable sars promulgated by the world health organization were meant for surveillance and were necessarily broadly inclusive and non-specific. patients were categorised based on clinical, radiologic and epidemiologic features and after exclusion of alternative diagnoses. the original who surveillance case definitions for sars required that lower respiratory symptoms of cough, short-ness of breath or difficulty breathing were present. applying this would have missed many children who do not present with the above symptoms. the lack of sensitivity and specificity of the initial who case definitions have generated uncertainty in individual case management at the point of care. , with more understanding of the disease and identification of a novel coronavirus as the causative agent, the case definitions of sars were revised on may , . as the clinical and radiologic features were non-specific, much emphasis was placed on the identification of an epidemiologic link to suggest the diagnosis. the vast majority of patients in the last epidemic had a clear history of exposure, either to patients suspected of or diagnosed with sars, or to a setting where recent local transmission was occurring. when the epidemic was over, an epidemiologic clue became more difficult to ascertain in sporadic cases that re-emerged. the latest who case definitions in the post-outbreak period now incorporate both clinical and laboratory elements, with further emphasis on clearly defined microbiologic criteria besides exclusion of alternative diagnoses (table ) . nevertheless, careful epidemiologic history taking remains essential in the diagnostic work-up and early implementation of appropriate infection control measures in suspected patients. important questions to ask in the 'peace time' include: ( ) history of recent travel to pre-viously sars-affected areas or areas with an increased likelihood of animal to human transmission of sars-cov infection; ( ) close contact with a suspected sars patient; ( ) history of recent hospitalisation or contact with a healthcare facility; ( ) individuals who are either healthcare workers or laboratory workers with potential exposure to sars patients or live sars-cov; and ( ) link to a cluster of cases of unexplained respiratory illness in the community. microbiological investigations are the cornerstones for the confirmation of sars. the diagnostic work-up should include tests for pathogens which cause communityacquired pneumonia in children. a blood culture is also needed. for children with productive cough who are old enough to produce a reliable specimen, sputum for bacterial culture should be performed. nasopharyngeal aspirate (npa) should be saved for rapid antigen detection of influenza a and b, rsv, adenovirus and parainfluenza types , and , using direct immunofluorescence assays. urine samples may be tested for legionella pneumophila and streptococcus pneumoniae antigens. npa specimens should also be inoculated into different cell lines for isolation of respiratory viruses. serologic studies should include mycoplasma pneumoniae igm and paired acute and convalescent sera for igg against mycoplasma pneumoniae, chlamydia pneumoniae, chlamydia psittaci, legionella pneumophila, influenza a and b, rsv, adenovirus and parainfluenza types , and . specific tests for the detection of sars-cov include: ( ) molecular or nucleic acid amplification test using reverse transcription-polymer-ase chain reaction (rt-pcr); ( ) antibody tests; and ( ) cell culture. in view of the high transmissibility of sars in hospitals, laboratory confirmation of the diagnosis early in the course of illness is vital to allow for the best utilisation of the limited isolation and cohorting facilities in most hospitals. rapid diagnosis with rt-pcr tests targeting specific segments of the sars-cov genome, primarily the polymerase gene, were used extensively during the last epidemic. [ ] [ ] [ ] [ ] [ ] [ ] [ ] the method can be applied to nasopharyngeal aspirates, nose and throat swabs, saliva, sputum, endotracheal aspirates, bronchoalveolar lavage, stool, urine, plasma and serum. nasopharyngeal aspirates, combined nose and throat swabs and stool are the most commonly used. experience in hong kong and toronto suggests that the first generation conventional rt-pcr assays in use at the time of the initial outbreak lacked sufficient sensitivity to clinically rule out sars. , despite initial optimism, the test has a sensitivity of % in npa, % in combined nose and throat swabs and % in stool in the first days of illness. it only reaches a maximum sensitivity of % when performed on upper respiratory specimens collected between days to from onset of fever (government virus unit, public health laboratory centre, hong kong special administrative region. data on file), where day coincides with the maximum viral load in npa specimens as clinical picture, diagnosis, treatment and outcome of sars in children a person with a history of: fever (! c) and one or more symptoms of lower respiratory tract illness (cough, difficulty breathing, shortness of breath) and radiographic evidence of lung infiltrates consistent with pneumonia or rds or autopsy findings consistent with the pathology of pneumonia or rds without an identifiable cause and no alternative diagnosis can fully explain the illness laboratory definition of sars a person with symptoms and signs that are clinically suggestive of sars and with positive laboratory findings for sars-cov based on one or more of the following diagnostic criteria: (a) pcr positive for sars-cov using a validated method from: at least two different clinical specimens (e.g. nasopharyngeal and stool) or the same clinical specimen collected on two or more occasions during the course of the illness (e.g. sequential nasopharyngeal aspirates) or two different assays or repeat pcr using a new rna extract from the original clinical sample on each occasion of testing (b) seroconversion by elisa or ifa negative antibody test on acute serum followed by positive antibody test on convalescent phase serum tested in parallel or four-fold or greater rise in antibody titre between acute and convalescent phase sera tested in parallel (c) virus isolation isolation in cell culture of sars-cov from any specimen and pcr confirmation using a validated method measured in adult patients. the low viral load in the upper respiratory tract in the initial few days of illness poses a diagnostic challenge. the lower respiratory tract as the primary target of sars-cov infection is the probable explanation. sputum specimens appear to have a higher diagnostic yield but productive cough is uncommon in sars patients in the early phase of illness and sputum is difficult to obtain in children. the overall diagnostic yield in the second week of illness increases to > % when stool specimens are also examined, with stool yielding better results than respiratory specimens. improving rna extraction from the specimen can markedly improve the sensitivity of conventional rt-pcr assays. when a modified rna extraction protocol is combined with an optimised real-time rt-pcr assay, a sensitivity of % and specificity of % can now be achieved in the first days of illness, using npa as the preferred specimen. a recently described real-time nested pcr assay performed on throat swabs is capable of detecting < copies of viral genome per reaction and achieves a much shorter turn-around time than conventional nested rt-pcr. the technique of real-time rt-pcr has also been applied to plasma and serum samples. it has been shown that % of plasma and % of serum samples are positive for sars-cov rna during the first week of illness in adult sars patients. a detection rate of . - % obtained in the plasma of eight paediatric patients within the first week of illness similarly suggests that plasma sars-cov rna quantification is a very sensitive and potentially useful early diagnostic tool. the potential advantages of realtime rt-pcr include an increase in sensitivity, reduction in analytical time, reduction of risk of carry over contamination and availability of quantitative result for disease monitoring and prognostic purposes. interestingly, despite a milder clinical course in paediatric patients, no significant differences in plasma viral loads are observed in plasma samples taken from paediatric and adult sars patients within the first week of admission and at day after fever onset. obtaining an npa specimen has been regarded by some as a hazardous procedure posing significant risk to the operator, although it is the best specimen for the rapid diagnosis of sars and the exclusion of other pathogens in the early phase of illness. to obviate the need for the protection of healthcare workers, an ingenious method for self-obtaining nasopharyngeal specimens through conjunctiva-upper respiratory tract irrigation (curti) has been described as an alternative. the lack of serologic evidence of prior sars-cov infection in humans suggests that the virus has only recently entered the human population, presumably from an animal reservoir in southern china. , specific igm and igg antibodies appear in response to sars-cov infection, with their levels changing during the course of the infection. serum antibody testing by immunofluorescence assay (ifa) or enzymelinked immunosorbant assay (elisa) have been developed to diagnose sars. , , , the ifa test detects igm and igg antibodies and yields positive results in % and % of cases, respectively, after days of illness. both are detectable in % of ifa tests by days. an indirect immunofluorescence test for igg antibody provides a sensitivity and specificity of %. the elisa test detects a mixture of serum igm and igg antibodies, % and % respectively being positive by the second week. detection rate for both is % by week . the decay curves suggest that igm seropositivity is lost by about weeks, while igg titres peak at weeks and remain elevated until weeks. the antibody response is usually negative until days from onset of symptoms. by day , seroconversion is demonstrated in % of sars patients despite corticosteroid therapy. seroconversion from negative to positive or a !four-fold rise in igg antibody titres indicates recent infection. no detection of antibody in serum obtained > days from onset of illness indicates an absence of sars-cov infection and is the only laboratory method for excluding the diagnosis. , serologic testing appears to be the best method for confirming sars, with positive rates ranging from % to %. , , igm or other antibody assays have not been successful in closing the diagnostic window within the first week of illness. even if some patients seroconvert early, the utility of serology is confined to retrospective diagnosis given the generally long lag time to seroconversion. igg usually remains detectable after resolution of the illness but the duration of persisting protective neutralising antibodies and their boosting response remain unknown. sars-cov can be isolated from respiratory secretions, blood or stool by inoculating cell cultures and growing the virus. vero e cells and fetal rhesus monkey kidney cells are suitable to support the viral growth, with the cytopathic effect demonstrable by - and - days respectively after inoculation. [ ] [ ] [ ] the cultured virus must be identified as sars-cov with further tests, primarily rt-pcr assays. the major limitation of viral culture in sars is its very low sensitivity. in one paediatric series, the virus was only successfully isolated from npa cultures in % of children. negative cell culture results, like negative rt-pcr results, do not exclude sars infection. cell culture is also a very demanding test and primary virus isolation takes too long to be meaningful for early diagnosis. furthermore, amplification of the viable virus is associated with a potential biohazard, necessitating biosafety level three containment. culture-based diagnostic techniques are unlikely to be widely available but with the exception of animal inoculation, it is the only way to show the existence of viable sars-cov. , the usual 'gold standard' of microbiological diagnosis, namely the isolation of the pathogen, has limited application in sars. during the global outbreak of sars, it was understandable that treatment was empiric, given the explosive epidemic of a life-threatening infection in multiple countries before the viral agent was even identified. time for planning, let alone conducting, a well-designed prospective clinical trial to assess the efficacy of any treatment regimen was simply not there. a proposed regimen consisting of antibiotics, ribavirin and corticosteroids was based on initial anecdotal successes in outbreak studies in adult patients. , subsequently, a standard treatment protocol was developed by a group of physicians in hong kong, which included ( ) antibiotics for treatment of community-acquired pneumonia caused by usual and by atypical pathogens, ( ) ribavirin as a broad-spectrum antiviral agent targeting the presumed viral etiology of sars, and ( ) immunomodulating agents in the form of glucocorticoids. a similar regimen in children consisting of antibiotics and ribavirin, with or without corticosteroids, was used. , , in adult patients, the high incidence of deranged liver function, leucopaenia, severe lymphopaenia, thrombocytopaenia and progression to ards suggests severe systemic inflammatory damage induced by sars-cov. the pathogenesis of the infection is postulated as an over-exuberant immunopathological reaction or a ''cytokine storm'' resulting from unrestricted viral replication during the early stages of the disease. findings consistent with cytokine dysregulation are the radiological changes of multifocal, flitting, boop-like features with progression to ards, the histological changes of macrophage infiltration and diffuse alveolar damage and the dramatic clinical and radiologic improvement with high-dose corticosteroid therapy. , the viral load in sars followed an inverted v pattern, with progressive fall in viral shedding after day - , correlating with seroconversion. the logical approach to preventing severe disease is to restrict viral replication and to modulate inappropriate immunological responses. in principle, antiviral agent should be prescribed first during the phase of active viral replication, followed by an immunomodulator if the former fails and the patient is affected by immune hyperactivation. the use of ribavirin in adults and children has been reported by groups of investigators worldwide. , , [ ] [ ] [ ] [ ] [ ] [ ] [ ] , [ ] [ ] [ ] [ ] [ ] [ ] ribavirin was empirically chosen in sars because of its broad-spectrum of activities against dna and rna viruses. ribavirin was also known to be effective in the treatment of fulminant murine hepatitis, which is caused by an animal coronavirus. in the murine hepatitis model, ribavirin exerted an immunomodulatory effect by decreasing the release of proinflammatory cytokines from the macrophages and switching the immune response from a th to a th response. , however, it was later learnt that ribavirin demonstrated no or minimal activity against sars-cov isolates in vitro. , in vitro testing indicated that ribavirin failed to inhibit replication or cell to cell spread at low drug concentrations. although inhibitory activity was demonstrated at high drug concentrations, the resultant cytotoxic effects were undesirable. it appeared that due to the low activity of ribavirin in vitro, inhibitory concentrations might not be achieved clinically without causing significant toxicity. investigators in canada have generally used ribavirin at a higher dosage similar to that recommended for treatment of several viral haemorrhagic fever syndromes and have observed severe adverse events in adult patients. booth et al. reported that % of patients had elevated hepatic transaminase levels, % had sinus bradycardia, % had haemolysis with haemoglobin levels declining by at least g/ dl in % and that % had to discontinue treatment. knowles et al. reported that %, % and % of patients had haemolytic anemia, hypocalcemia and hypomagnesaemia, respectively. children appear to tolerate ribavirin much better than their adult counterparts. [ ] [ ] [ ] [ ] solid clinical data to demonstrate the efficacy of ribavirin is lacking. the limited data suggest that, at least in adults, dosages of about g/d might be effective while not causing severe adverse reactions. such doses should be considered for further studies. doses lower than g/d appear ineffective. the only randomised controlled trial involving the use of ribavirin in the treatment of sars was conducted in china by zhao et al. the open-label study failed to demonstrate any efficacy and led the investigators to conclude that ribavirin, given at - mg/d, was less effective than early and aggressive use of corticosteroids combined with non-invasive ventilatory support. non-randomised studies of corticosteroids have been reported in both adults and children with seemingly favourable outcomes in terms of clinical and radiologic improvements, suggesting that the combined use of ribavirin and corticosteroids might be effective. , , [ ] [ ] [ ] , , , , , , other reports on the combined regimen were inconclusive or failed to demonstrate obvious benefit. , , in the paediatric series reported by leung et al., % and % of the children with laboratory-confirmed sars were treated with ribavirin and corticosteroids respectively, without significant adverse events and all patients recovered. in the series reported by chiu et al., % and % of the children received ribavirin and corticosteroids, respectively and achieved similar outcomes. all were subsequently confirmed by seroconversion to sars-cov after the report was published. bitnun et al. reported the use of ribavirin without corticosteroids in children with probable sars but virologic confirmation was lacking. in contrast, zeng et al. treated children with chinese traditional medicine and antibiotics with good results. only of the children had an epidemiologic link to sars, however, and virologic data were not available. the use of corticosteroids in viral infections is controversial and is potentially hazardous. as an immunosuppressive agent, corticosteroids might promote viral replication, enhance infectivity and possibly cause a rebound of infection. it is known that in acute viral respiratory infections, early-response cytokines such as tumour necrosis factor, interleukin- and interleukin- mediate lung injury. the rationale for using corticosteroids is to suppress the ''cytokine storm'' which is thought to be the main factor accounting for the progression of disease. but using corticosteroids with possibly ineffective antiviral therapy in patients with viral pneumonitis can be hazardous. despite the initial success of corticosteroids in the treatment of sars, the report of an adult patient whose clinical course was complicated by fatal aspergillosis was disturbing and had even led others to recommend close laboratory monitoring for aspergillosis in sars patients receiving corticosteroids. , in retrospect, we do not think that ribavirin alone has any significant effect in halting disease progression and corticosteroids are probably unnecessary for children who do not develop moderate to severe hypoxaemia. in our experience, as with others, corticosteroids may be life saving in patients who are threatened by impending acute respiratory failure. we cannot categorically recommend this treatment strategy in view of the small number of children treated and the lack of objective evidence from a controlled trial. the place of corticosteroids in the rescue therapy of patients who have clearly experienced failure of supportive care remains to be determined. no evidence-based therapeutic approach for sars exists although more than papers have been published internationally that mention antiviral treatment. various other antiviral and immunomodulating agents have been used in adult patients with preliminary success. these include the use of lopinavir / ritonavir in combination with ribavirin and corticosteroids, interferon a plus corticosteroids and convalescent plasma from patients. [ ] [ ] [ ] [ ] their true role in the treatment of children is unknown. knowledge generated by detailed bioinformatic analysis of the sars-cov genome can be harnessed to identify possible targets for antiviral therapy, such as enzymatic proteins of the viral replicase-transcriptase complex. this approach has been reviewed by davidson and siddell who concluded that the most economical and effective way to contain the virus would be the therapeutic use of antiviral agents to block viral entry to target cells or to inhibit intracellular viral replication. in vitro studies have highlighted the antiviral potential of several compounds, including recombinant human interferon b- a, interferon b- b, glycyrrhizin, human monoclonal antibody against the spike protein of sars-cov and small interfering rna. [ ] [ ] [ ] [ ] [ ] with more understanding of the pathogenesis as well as the clinical course of the disease, treatment will evolve. the best treatment for sars in adults and children remains unknown. time is now on our side to plan for clinical trials should the disease re-emerge. with increased vigilance, rapid detection and effective infection control measures, outbreaks of sars seem less likely. it might never be possible, therefore, to recruit a sufficient number of patients to complete the trials and give us an early answer. in adults, the risk factors for severe illness are advanced age, high initial absolute neutrophil counts, low platelet counts, high initial or peak lactate dehydrogenase levels and positive rt-pcr results for npa specimens. , [ ] [ ] [ ] [ ] only one paediatric series has identified risk factors for severe illness in terms of requirements for oxygen and intensive care. these include a sore throat, a high neutrophil count at presentation, and peak neutrophilia. the finding of sore throat as an independent risk factor is intriguing but may be incidental, given the small number of patients. no association between the presence of sore throat and the detection of sars-cov by rt-pcr or culture in npa specimens, which might correlate with higher viral load, could be demonstrated. the short-term outcome of sars among children is good in comparison to adults. no case fatality has been reported. the need for intensive care and mechanical ventilation was up to . % and . % respectively in adults. chiu et al. reported that . % of children required oxygen supplementation and none required assisted ventilation. leung et al. reported an oxygen requirement in . % and assisted ventilatory support in . % of children. the figures for oxygen requirement and assisted ventilation in the two paediatric series combined are % and %, respectively. diffuse thinning and shedding of hair was observed in . % of children in one series, generally at - months after disease onset. the condition was self-limiting and spontaneous recovery occurred within - months. this is consistent with acute telogen effluvium secondary to febrile systemic illness, critical care or severe psychologic stress in life-threatening situations. li et al. examined the radiologic and pulmonary function outcomes of children, months after diagnosis and detected mild radiologic abnormalities with hrct and in pulmonary function testing in % and . % respectively. however, all children were asymptomatic and had normal clinical examination, premorbid hrct and pulmonary function test results were not available for comparison. in contrast, some adult patients have devel-oped pulmonary fibrosis despite recovery from the primary illness. the psychological impact of separation, isolation in an intimidating hospital environment, bereavement and family disintegration following the death of close adult family members in children who recovered from sars are immense. however, children appear to be more resilient than adults in psychological adjustment to sars and serious psychological sequelae were not evident months after discharge. continued monitoring for delayed onset of psychological problems in children is essential. children who have recovered from the acute illness should be monitored for the possibility of continued viral shedding and the development of pulmonary sequelae and postviral complications (e.g. chronic fatigue), as well as for any long-term complications of high-dose corticosteroid therapy. children are susceptible to infection by sars-cov. despite the milder clinical picture, the good short-to medium-term outcome and the availability of reliable early diagnostic techniques, treatment remains controversial. the long-term outcome of sars in children remains unknown. there are still enormous gaps in our knowledge about sars. much work needs to be done, urgently. sars is largely an atypical pneumonia with minimal or no extrapulmonary manifestation apart from diarrhoea. the clinical picture of sars is milder in children but teenagers may develop severe illness resembling adults. the clinical, radiologic and laboratory features of sars are non-specific. an epidemiologic link is the most important clue to diagnosis in an outbreak situation. refined rt-pcr assays can achieve a sensitivity of % in the early diagnosis of sars in the first days of illness. npa specimens are the preferred specimens for rt-pcr assays in the first week of illness. both npa and stool specimens should be tested in the second week. a negative rt-pcr result cannot exclude the diagnosis. absence of seroconversion beyond days from disease onset generally excludes the diagnosis. apart from supportive treatment, including oxygen therapy and assisted ventilation, other treatment modalities remain unproven. molecular biology of sars-cov and mechanisms of its genome expression. pathogenesis of sars-cov infection. natural history and full spectrum of sars-cov infection. improved early diagnostic techniques. 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sars with human interferons glycyrrhizin, an active component of liquorice roots, and replication of sars-associated coronavirus potent neutralization of severe acute respiratory syndrome (sars) coronavirus by a human mab to s protein that blocks receptor association inhibiting sars coronavirus by small interfering rna severe acute respiratory syndrome: clinical outcome and prognostic correlates outcomes and prognostic factors in patients with severe acute respiratory syndrome in hong kong coronavirus-positive nasopharyngeal aspirate as predictor for severe acute respiratory syndrome mortality prognostic factors for severe acute respiratory syndrome: a clinical analysis of cases radiological and pulmonary function outcome of children with sars thin-section ct in patients with severe acute respiratory syndrome following hospital discharge: preliminary experience key: cord- - l mk authors: clark, ian a.; griffiths, michael j. title: the molecular basis of paediatric malarial disease date: journal: pediatric infectious diseases revisited doi: . / - - - - _ sha: doc_id: cord_uid: l mk severe falciparum malaria is an acute systemic disease that can affect multiple organs, including those in which few parasites are found. the acute disease bears many similarities both clinically and, potentially, mechanistically, to the systemic diseases caused by bacteria, rickettsia, and viruses. traditionally the morbidity and mortality associated with severe malarial disease has been explained in terms of mechanical obstruction to vascular flow by adherence to endothelium (termed sequestration) of erythrocytes containing mature-stage parasites. however, over the past few decades an alternative ‘cytokine theory of disease’ has also evolved, where malarial pathology is explained in terms of a balance between the pro- and anti-inflammatory cytokines. the final common pathway for this pro-inflammatory imbalance is believed to be a limitation in the supply and mitochondrial utilisation of energy to cells. different patterns of ensuing energy depletion (both temporal and spatial) throughout the cells in the body present as different clinical syndromes. this chapter draws attention to the over-arching position that inflammatory cytokines are beginning to occupy in the pathogenesis of acute malaria and other acute infections. the influence of inflammatory cytokines on cellular function offers a molecular framework to explain the multiple clinical syndromes that are observed during acute malarial illness, and provides a fresh avenue of investigation for adjunct therapies to ameliorate the malarial disease process. although many species of malarial parasite exist, only plasmodium falciparum, vivax, ovale, and malariae are classically associated with human infection. the former two species are most frequently associated with malarial disease in humans, with severe malarial disease almost exclusively associated with p. falciparum infection. falciparum malaria is responsible for considerable morbidity ( - million annual clinical cases) and death across the globe, with a particular burden of mortality among children in sub-saharan africa. infection with p. vivax is rarely fatal, but is associated with considerable morbidity outside the african continent. it should also be recalled that malaria causes social and economic disruption on a uniquely large scale [ ] . severe adult malaria is a clinical syndrome originally classified using defining and supportive (often overlapping) clinical features unified by the presence of asexual malarial parasites in the peripheral blood smear [ ] . based on observations of children in coastal kenya, paediatric severe malaria has similarly been distilled into three main (again often overlapping) clinical syndromes, anaemia, respiratory distress (an indicator of an underlying metabolic acidosis) and impairment of consciousness [ ] . these clinical syndromes are discussed below. in the review mentioned above [ ] , the authors' judicious use of the term impaired consciousness, rather than cerebral malaria (cm), promoted the useful concept that the neurological features (and in-turn the underlying mechanisms) associated with severe malaria are not necessarily unique to malarial disease. indeed, over years ago, it was noted that the clinical features of malaria can resemble those exhibited in patients with fulminant bacterial or viral infections [ ] . severe malaria has been intensively studied, and there appears to be a complex interplay between host infection and disease. this is highlighted by the different clinical manifestations of severe malaria exhibited by children and adults. these differences are undoubtedly, in part, a function of patient age. however, age is just one of a series of interacting factors, e.g. geographical region, level of malaria transmission, degree of previous malaria exposure, length of illness prior to treatment and host immunity that may influence the clinical presentation of severe malaria. this variation in clinical presentation has been mirrored by a similar multitude of proposals regarding the functional mechanisms underlying pathogenesis of severe malaria. one concept of pathogenesis consistently articulated has been the 'mechanical theory'. historically, this theory was developed from two fundamental differences between p. falciparum and p. vivax infection. firstly, erythrocytes parasitised with p. vivax do not sequester. secondly, death following p. vivax infection is rare. consequently, pathogenesis is believed to be due to obstruction of micro-vascular flow by erythrocytes containing mature-stage falciparum parasites adhering to the endothelium (termed sequestration). more recently the 'cytokine theory of disease' has also gained credence. this theory can be applied to disease following both falciparum and vivax infection. the lower mortality associated with p. vivax being explained by a relatively milder degree of pro-inflammatory imbalance during the host's response to p. vivax infection. the main theme of this chapter is to examine the increased understanding of the functions of inflammatory cytokines gained over the past years, and explore how these insights are changing attitudes in malarial disease research. we also discuss how two theories (mechanical and cytokine) can, as proposed first in a recognisable form at least years ago [ ] , be complementary. the majority of the clinical cases of malaria occur in sub-saharan africa. nevertheless, malaria also accounts for considerable morbidity and mortality in other continents particularly south east asia [ ] . in malaria-endemic regions (e.g. sub-saharan africa), where the resident population have continuous exposure to malarial parasites, most of the severe cases are seen in children [ ] . in hypoendemic regions (e.g. south east asia), where parasite exposure is more intermittent, cases of severe malaria are also common in adults (tab. ). clinical features associated with malaria mortality vary between children and adults, but acidosis and coma are associated with malarial mortality in both populations [ , ] . acute renal failure (arf) and pulmonary oedema, a marker for adult respiratory distress syndrome (ards), are almost exclusively reported among adults [ , ] , whereas mortality associated with hypoglycaemia is frequently reported among children [ ] . why malarial disease displays such age-related differences in pathophysiology is unclear. however, these differences are not exclusive to malaria. ards, which is more frequently observed as a complication of trauma in adults compared with children [ ] , is believed to reflect an exaggerated pro-inflammatory response within the lung [ ] . a possible lead for future studies on these age-related differences in malaria is suggested by a report of peritoneal macrophages collected from healthy adults producing much less interleukin (il)- (an anti-inflammatory cytokine), but the same levels of pro-inflammatory cytokine, than those from healthy children, giving adults a much higher pro-inflammatory status [ , ] . the mechanism of malarial arf pathogenesis is postulated to be multifactorial, involving mechanical, haemodynamic, and immunological factors [ ] . the observation that arf is more frequently observed as a complication of trauma in adults than children [ ] suggests that age-related variations in cytokine response may again influence pathogenesis. hypoglycaemia is regarded as a more frequent complication of sepsis in paediatric populations compared with adults [ ] . hypoglycaemia in children may, in part, be associated with a higher basal metabolic rate, and lower glycolytic [ ] and gluconeogenic substrate reserves compared to adults [ ] . however, these substrates are not always limiting during acute paediatric malaria, suggesting functional impairments of glucose metabolism may also occur [ ] . such functional impairments may, in part, be influenced by increases in inflammatory cytokines as the infection progresses. once the malarial parasite was identified as the cause of disease, it quickly became apparent that illness and death were linked with parasite invasion into bloodstream and subsequent parasite growth within (and release from) the erythrocytes. by the start of the th century, two major theories, capillary blockage and toxicity of the parasites themselves, had been proposed to explain morbidity and mortality. thus, the study of malarial disease is not a settled story requiring regular updates, but one containing, from its beginning, an unresolved tension. vascular occlusion and malarial toxin (nowadays vascular occlusion and inflammatory cytokines) have been alternative approaches to understanding malarial disease as a whole, as well as the coma, for over a century, and the two have often been discussed side by side [ , , ] . the presence of hyperlactataemia, hypoglycaemia, and metabolic acidosis, all three consistent with a patient being forced to rely on anaerobic glycolysis for energy production, have provided a consensus that hypoxia is central to disease pathogenesis in falciparum malaria. as sum-marised below, the modern literature offers two main theories for cellular hypoxia during infection; insufficient oxygen delivery to cells and impaired oxygen utilization within the cells. both mechanisms may be governed by the host inflammatory cytokine response to infection. this chapter focuses on how an increased understanding of the molecular functions of cytokines during disease demonstrates a closer alignment between the pathogenesis of falciparum infection and other systemic infectious diseases. one hundred and twenty years ago, golgi (of the golgi apparatus [ ] ), noted onset of malarial fever and illness at a predictable short interval after the regular shower of new parasites were released from bursting red cells. the nature of the putative toxin so released was much discussed in the first decade of the th century [ ] . it was assumed to be directly toxic, in the manner of tetanus toxin. the proposal that malarial products were not harmful in themselves, but only through causing the infected host to harm itself through generating toxic amounts of molecules (pro-inflammatory cytokines) that, in lower concentrations, inhibit growth of malarial parasites did not arise until [ ] . indeed, acceptance of the broad applicability of this concept to infectious disease in general is now sufficient for its evolution to be a subject for research [ ] . tumour necrosis factor (tnf) is regarded as a major player, malaria being the first disease in which it was proposed to cause systemic illness and pathology [ ] . multiple tnf promoter polymorphisms have since been independently associated with severe malaria across several geographical populations [ ] . a longitudinal study in burkino faso has also demonstrated several tnf promoter polymorphisms associated with the regulation of host-parasite density [ ] . the tnf concept has since begun to dominate the sepsis literature [ ] , and the virulence of different strains of influenza, a disease that is a standard clinical misdiagnosis for imported malaria, has recently been expressed in terms of their capacity to induce tnf [ ] . the critical role of tnf in both malaria and influenza pathogenesis is consistent with the clinical similarities between the diseases. indeed, tnf infusions in tumour patients produce side effects mimicking both diseases [ ] , as discussed below. although tnf is the prototype pro-inflammatory cytokine linked with severe malaria, other cytokines (and mediators) including interferon (ifn)- [ ] , its corresponding receptors ifn-receptor- [ ] and ifn-receptor- [ ] , il- [ ] , il- [ ] and il- [ ] have all be identified through genetic association analysis to be linked with their potential regulation of malarial disease severity. all the above cytokines typically act as homeostatic agents, but can cause pathology if produced excessively. when this happens they also induce a late-onset, but long-acting cytokine termed the high mobility group box (hmgb ) protein, which prolongs and amplifies inflammation [ , ] . this molecule, normally in the cellular nucleus and previously known only for several physiological functions, now shows great promise as a therapeutic target in sepsis, in that countering it after the onset of illness protects well in experimental sepsis [ , ] . it accumulates, in proportion to degree of illness, in serum from african children infected with falciparum malaria [ ] . once neutralising anti-tnf antibodies became available for human use, they were tested for efficacy against malarial disease. unfortunately, a central tenet of the cytokine concept of infectious disease (that the proinflammatory cytokines that cause disease are the same mediators that, in lower concentrations, are responsible for the innate immunity that controls parasite growth) was not taken into consideration. tnf has been shown to inhibit a mouse malarial parasite in vivo [ ] , and p. falciparum in vitro, provided white cells to generate the next down-stream mediator, possibly nitric oxide (no) [ ] , were present [ ] . this is consistent with findings in human subjects [ ] . thus, it is not surprising that anti-tnf antibody, by removing inhibitory pressure from the pathogen, can enhance the disease in falciparum malaria [ ] , as shown years earlier in human sepsis [ ] . cytokines as a disease mechanism extends beyond malaria as noted above, the idea that excessive production of inflammatory cytokines underlies the pathology of illness is used widely, from malaria across a range of conditions, infectious or otherwise. as reviewed recently [ ] , this now includes the illnesses caused by rickettsias, protozoa other than malaria, and viruses. increased circulating levels of these cytokines have been detected in the serum very soon after onset of illness in virtually all those infectious diseases in which they have been sought. some cytokine increased, and consequences are shown in table . when rtnf was under trial in volunteers as an anti-tumour agent [ , ] nearly years ago, virtually all of the symptoms and signs they share were reproduced as side effects. this includes headache, fever and rigours, nausea and vomiting, diarrhoea, anorexia, myalgia, thrombocytopaenia, immunosuppression, and central nervous system manifestations, all of which have been shown to be caused by a mechanism involving inflammatory cytokines. the rate, timing and intensity of cytokine release vary in different disease states, and provide them with somewhat individual clinical pictures, but the fundamentals remain. nevertheless, the clinical patterns generated are remarkably close, in that, at least in some populations, clinical features cannot predict a diagnosis of malaria from other causes of fever [ ] . mature erythrocytic forms of p. falciparum are not seen in peripheral blood smears, and cause the erythrocytes they inhabit to adhere to the walls of venules and capillaries. from this observation arose the widely held view that much of the pathology following malarial infection is explained through parasite sequestration causing impairment of microvascular flow. sequestration certainly occurs, since the life cycle dictates this. however, whether the temporal and anatomical patterns of sequestration are the same in both individuals with fatal disease and in parasite tolerant individuals has not been ascertained. consequently, whether sequestration is the principal instigator of local pathology, or whether sequestration is an associated feature of all malarial infections with local pathology determined by other factors in the host response to the infection, e.g. a local imbalance of inflammatory mediators, has not been fully elucidated. erythrocyte cyto-adherence (irrespective of whether this adhesive process is directly or indirectly due to parasite sequestration) has repeatedly been shown to be mediated through a series of host-derived ligands. cd and thrombospondin were the first described endothelial receptors that bound infected red blood cells (rbcs) [ , ] , with most studied wild parasite isolates demonstrating adhesion to cd [ ] . more recently, it has been shown that p. falciparum also interacts with other host adhesion receptors, i.e. intercellular adhesion molecule- (icam- cd ), vascular cell adhesion molecule- (vcam- cd ) and e-selectin [ , ] . certain adhesive phenotypes, such as rosetting (the spontaneous tethering of infected and non-infected rbcs) and clumping (tethering of infected rbcs through platelets) have been preferentially associated with severe malarial disease [ , ] . cd is involved in both mechanisms of adhesion, and a non-sense mutation in the gene encoding for cd has also been associated with protection from severe malaria [ ] . polymorphisms in the gene encoding icam- have also been associated with susceptibility to severe disease [ ] . furthermore, icam- , together with vcam and e-selectin, are up-regulated by tnf, with circulating levels of these ligands shown to be increased in severe malaria compared to uncomplicated infection [ ] . sequestration during falciparum malaria appears to be concentrated in the brain and placenta. there is some evidence to suggest that the propensity of inflammatory cytokines to up-regulate cell adhesion molecules, secondary to local variation in the density of thrombomodulin, is potentially higher in the microvasculature of the brain and placenta compared to other tissues. as reviewed [ ] , tnf and il- increase tissue factor expression on endothelial cells, thereby initiating pathways that generate thrombin [ ] . when thrombin binds to thrombomodulin on the endothelial cell surface, protein c is activated, which in turn can lead to further downstream activation of the coagulation cascade. therefore vasculature with lowest thrombomodulin densities on the endothelial cell surface (brain least, placenta next least, and other organs more [ ] ) will have more unbound thrombin available for its other functions on activated endothelium. these other functions include up-regulation of adhesion molecules such as selectins, icam- , vcam- [ ] and monocyte chemotactic protein- (mcp- ) [ ] . therefore, up-regulation of adhesion molecules within the cerebral vessels may occur as a local endothelial response to systemic inflammation and may not necessarily be precipitated by parasite sequestration. anaemia is another obvious way in which too little oxygen reaches cells, and thus their mitochondria [ ] . as recently reviewed [ ] , critical illness associated with an inflammatory response invariably causes multifactorial anaemia. obviously a high parasite load in malaria indicates that the infected rbcs will soon burst when the next generation of erythrocytic forms escapes, but anaemia does not correlate with parasitaemia, and sometimes is extreme when very few parasites are, or have been, present. the severe anaemia in transgenic mice expressing human tnf [ ] incriminates the inflammatory response itself, so anaemia and mitochondrial dysfunction (see mitochondrial dysfunction section below), both consequences of systemic inflammation, can be expected to coexist, and both contribute to total energy depletion. the lifespan of an rbc is, in part, limited by how long it can remain flexible enough to squeeze through fenestrations in specialised vessels in the red pulp of the spleen, and thus avoid phagocytosis by adjacent macrophages. normally this loss is balanced by erythropoiesis, and haematocrit remains normal. if rbcs develop a premature loss of deformability they are removed from the circulation earlier. this loss of deformability happens to both infected and non-infected red cells in malaria, whether caused by p. vivax or p. falciparum. under physiological conditions, erythrocytes (and other cells) control the passive influx of osmotic active solutes (especially na + ) via an active, energy-dependent elimination of these solutes using na + /k + -atpase. this prevents intracellular accumulation of osmotically active solutes, preventing a subsequent influx of water, cell swelling and loss of cell integrity. during human [ ] and monkey [ ] malaria infection, intracellular na + accumulates within erythrocytes (both parasitised and non-parasitised) implying that this na + /k + pump is impaired during the disease process. parallel changes in the ionic content of erythrocytes have been documented in a sepsis model of infection [ ] . similarly, reduction in erythrocyte deformability was shown to be associated with increased no, an inhibitor of this membrane pump [ ] , in another sepsis model [ ] . since inhibition of the na + /k + pump in vitro correlates with both reduced red cell deformability and decreased red cell filterability [ ] , any factor that inhibits the na + /k + pump could potentially worsen anaemia. identification of inducible no synthase (inos) activity, as one factor influencing red cell deformability, suggests that a pro-inflammatory milieu [ ] may again govern the reduction in red cell deformability observed during malaria infection. originally observed in uraemic patients, poor red cell deformability was recognised in a small pilot study of malaria patients in [ ] . it was reported soon afterwards in sepsis [ , ] , and subsequently studied in falciparum malaria with a view to understanding both circulatory obstruction [ ] and anaemia [ ] . it seems clear that a short life (poor deformability), and a slow replacement rate (dyserythropoiesis, below) can combine to cause severe anaemia in various diseases, particularly in chronic infections such as malaria. when red cells have a shortened lifespan, e.g. secondary to reduced erythrocyte deformability, replacement by new recruits is vital to avoid anaemia. unfortunately, the same inflammatory cytokines that shorten lifespan also retard replacement. some years ago researchers began to stress the contribution of bone marrow dyserythropoiesis to the anaemia of falciparum malaria [ , ] . a group in oxford [ ] , seeking an explanation for this dyserythropoiesis through an electron microscopy study of bone marrow, observed sequestration of parasitised red cells and argued that this caused the bone marrow dysfunction in falciparum malaria by restricting blood flow and thus inducing hypoxic changes. this idea proved inadequate, however, when this same group subsequently reported dyserythropoiesis and erythrophagocytosis in vivax malaria, in which parasitised red cells do not sequester [ ] . some time ago an undefined product in macrophage supernatants [ ] , later identified as tnf [ ] , was found to inhibit the growth and differentiation of erythroid progenitor cells. when rtnf became available, the dyserythropoiesis and erythrophagocytosis seen in terminal plasmodium vinckei-infected mice was reproduced by giving a single injection early in the course of the infection [ ] . phagocytosis of erythroblasts in bone marrow, a phenomenon also reported by wickramasinghe et al [ , ] in human malaria, also occurred. decreased erythropoiesis was subsequently reported in mice receiving continuous tnf infusions via implanted osmotic pumps, and mice expressing high levels of human tnf have been shown to become markedly anaemic during malaria infections [ ] , even though parasite numbers, and therefore red cell loss post-schizogony, are considerably reduced. the past decade has seen an expansion of this line of enquiry into human malaria, and also the number of cytokines, both pro-inflammatory and anti-inflammatory [ , ] in absolute amounts and ratios [ , ] , that have been investigated in this context. investigations have been extended to include other pro-inflammatory cytokines, such as il- [ ] and fasl [ ] , and examined the role in anaemia of the persistence of cytokine production during malaria infection [ ] . another inflammatory cytokine, macrophage inhibitory factor (mif) that is increased in malaria, and induced by tnf, has been shown to cause dyserythropoiesis in in vitro studies on bone marrow cells [ , ] . thus, inflammatory cytokines generated during malaria are a major determinant of the degree to which anaemia influences the amount of oxygen that reaches tissues in malaria. mitochondria are vital to energy (atp) generation through cellular respiration. cellular respiration requires oxygen and pyruvate, as well as multiple cofactors and active transport molecules. within the matrix of the mitochondrion organelle, pyruvate is catabolised via the krebs cycle and oxidative phosphorylation (involving nadh and fadh ) to generate atp. when this series of reactions are % efficient (unlikely in vivo), molecule of glucose generates molecules of pyruvate, which are further catabolised to water and carbon dioxide with the concomitant generation of molecules of atp. in comparison, during anaerobic glucose catabolism, pyruvate is converted to lactate with the concomitant generation of molecules of atp, a process that also facilitates regeneration of nadh and fadh . evidence is accumulating that inflammatory cytokines, as released in malaria, sepsis, and viral diseases, induce mitochondrial dysfunction and dysregulate cellular respiration, resulting in the incomplete catabolism of pyruvate. the process, termed 'cytopathic hypoxia' [ ] , mimics cellular hypoxia, in that it results in the incomplete catabolism of pyruvate and accumulation of lactate. awareness of this mechanism began with oxygen tension being shown to be increased in septic rats [ ] and patients [ ] . a cytokine model of mitochondrial dysfunction has since been developed in which impairment of cellular respiration occurs following induction of sepsis (or exposure to pro-inflammatory cytokines), despite sufficient oxygen supply [ , , ] . more recently, impairment of enzyme activity associated with the mitochondrial complexes has been demonstrated in muscle biopsies retrieved from rodent models of sepsis [ ] and septic patients [ , ] . the observation that the inflammatory cytokines implicated in mitochondrial shutdown are prominent in both sepsis and malaria [ , ] supports such organelle dysfunction being equally plausible in malaria. researchers are also becoming aware that, beyond energy production, mitochondria also play a vital role in cell homeostasis through generation and detoxification of reactive oxygen species [ ] . the accelerated oxidative damage that accompanies sepsis could be both a cause and a consequence of cytokine-induced mitochondrial dysfunction. interestingly, the ultrastructural damage reported to accompany mitochondrial dysfunction in sepsis [ ] reflects maegraith's observations in monkey malaria [ ] [ ] [ ] decades ago. metabolic acidosis, often associated with hyperlactataemia, has been described in african children with severe falciparum malaria [ , ] . it is not unique to this disease, being seen in viral, rickettsial and bacterial infections [ ] as well as acute gastroenteritis, where its prevalence is higher than in malaria [ ] . the terms hyperlactataemia and lactic acidosis are often mistakenly used interchangeably in the malaria literature. as often reviewed in the basic literature [ ] [ ] [ ] [ ] , protons (h + , the basis of acidosis) are not formed when atp and lactate are generated during glycolysis, but on the subsequent hydrolysis of atp in tissues. every time a molecule of atp undergoes hydrolysis, a proton is released. if this occurs under aero-bic conditions, these protons are consumed within atp regeneration from adp, and ph remains normal, i.e. acidosis does not occur. in contrast, if the mitochondria are not functioning adequately, whether through insufficient oxygen supply or an inability to use it, atp regenerates under anaerobic condition, and the protons are not consumed. hence, once the buffering capacity of the body is exceeded, acidosis occurs. in short, metabolic acidosis requires the ratio of glycolytic (i.e. anaerobic) atp hydrolysis to mitochondrial (i.e. aerobic) atp hydrolysis to reach a point at which the buffering systems can no longer cope. pathological changes in the buffering system can be a major determinant of when this occurs. high lactate levels have traditionally been seen not only as a marker for poor oxygen delivery in disease states, but also a consequence of it, and the cause of the acidosis. for some time hyperlactataemia has been regarded as a functionally relevant marker for a poor prognosis in both sepsis [ ] and malaria [ , , ] . although the sepsis world now discusses several origins for the lactate increase, including inflammation-induced mitochondrial dysfunction [ ] , in falciparum malaria it is still generally attributed to a reduced oxygen supply, mostly through microvascular occlusion by sequestered parasitised erythrocytes [ ] . other mechanisms are known to contribute to acidosis in malaria, independent of lactate production, e.g. acute renal failure [ ] . impaired hepatic clearance [ , ] , production by parasites, and, in some areas, thiamine deficiency [ ] are also argued to contribute to lactate accumulation independent of impaired cellular respiration. thus, as described below, although acidosis and hyperlactataemia can be associated, they are independent cellular mechanisms. lactate anion has complex roles in biology. hyperlactataemia may be associated with acidosis, a normal ph, or alkalosis [ ] . a recent editorial in critical care medicine [ ] has lucidly summarised the key points of the mechanism of metabolic acidosis in sepsis, a condition that shares systemic inflammation and a range of its consequences with severe malaria (tab. ). these authors argue against lactate as the cause of the acidosis associated with hypoxia. instead, they note the evidence that during hypoxia, be it from limited oxygen supply or utilisation, the unconsumed protons that cause acidosis arise from the hydrolysis of non-mitochondrial atp. since these reactions are independent of lactate levels, it is difficult to see how therapeutically reducing levels of this anion, as has been proposed [ ] , could increase survival rate in falciparum malaria any more than in sepsis [ ] . indeed, in theory it could harm comatose patients, since there is evidence that lactate helps brain tissue survive hypoxic and hypoglycaemic episodes [ ] [ ] [ ] , and the lactate shuttle is proving to be how astrocytes protect neurons from metabolic stress [ ] . even when considerable lactate is generated in acute inflammatory states, other, unidentified, anions contribute much more than it does to the strong ion difference that, through influencing the body's buffering capacity, influences acidosis in sepsis [ , ] and falciparum malaria [ , ] . thus, lactate accumulation can only partially account for the high anion gap observed during the metabolic acidosis associated with severe malaria. in summary, lactate is an imprecise but useful marker for metabolic acidosis in malaria. in turn, acidosis is an imprecise but useful marker of impaired cellular respiration. whether impaired cellular respiration arises from (a) poor supply of oxygen to mitochondria (through vaso-occlusion, low circulating volume, anaemia or cardiac insufficiency) or impaired mitochondrial function (in response to severe systemic inflammation) the outcome is essentially the same. the resulting high anion gap metabolic acidosis is strongly predictive of death in severe malaria. greater understanding of the multiple factors influencing the metabolic acidosis could provide further insight into the underlying pathophysiological process and may provide additional therapeutic options. when glycolysis is enhanced for any period glycogen stores are soon depleted, and gluconeogenesis supervenes. however, its substrate supplies are limiting [ ] , and the hypoglycaemia often reported in severe malaria [ ] and sepsis [ , ] occurs. hypoglycaemia is therefore a secondary cause of harm in these diseases, and is an inevitable consequence of exuberant, mostly anaerobic, glycolysis. cm is a clinical syndrome characterised by coma (inability to localise a painful stimulus) at least h after termination of a seizure or correction of hypoglycaemia, detection of asexual forms of p. falciparum malarial parasites on peripheral blood smears, and exclusion of other causes of encephalopathy [ ] . a relatively consistent feature of acute cm in children is raised intracranial pressure (icp). studies in african children have demonstrated a raised cerebrospinal fluid (csf) opening pressure during lumbar puncture in % of cm children [ ] , raised icp during intracranial pressure monitoring ( / icp > mmhg) [ ] and papilloedema (a late sign of raised icp) in % of cm patients who died [ ] . where computer tomography has been performed, there was evidence of diffuse brain swelling in % of patients [ ] . the cause of the raised icp is likely to be multi-factorial and has been postulated to involve both vasogenic and cytotoxic patterns of cerebral oedema. vasogenic oedema is characterised by accumulation of interstitial fluid within the brain secondary to increased permeability of the blood-brain barrier (bbb). it has been demonstrated in bacterial cerebral infections, but evidence of significant disruption of the bbb is not conclusive in cm [ ] . others have proposed that icam- binding by infected erythrocytes may generate a cascade of intracellular signalling events that disrupt the cytoskeletal-cell junction structure and cause focal disruption to the bbb [ ] . adult post-mortem analysis has shown cerebrovascular endothelial cell activation (increased icam- endothelial staining, reduction in cell junction staining, and disruption of junction proteins), particularly in vessels containing infected erythrocytes [ ] . however, disruption of intercellular junctions is not associated with significant leakage of plasma proteins (fibrinogen, igg, or c b- ) into perivascular areas or csf [ ] . in thai adults, transfer of radioactively labelled albumin into csf was not raised during unconsciousness compared with convalescence [ ] . similarly, the albumin index (ratio of concentrations of albumin in csf to those in blood) was not altered significantly in vietnamese adults [ ] or significantly different between malawian children with cm who died and those who survived [ ] . cytotoxic oedema is increasingly being recognised as an important mechanism of cerebral oedema in traumatic brain injury [ ] . as previously discussed, this type of cell swelling involves disturbance of the "pumpleak equilibrium" maintained, under physiological conditions via active elimination of osmotically active solutes through the energy-dependent na + /k + -atpase. thus, cytotoxic oedema can occur secondary to an imbalance in supply and demand of energy within the cells. several mechanisms, such as sustained increase in neuronal activation, impaired substrate delivery (structural and functional) and impaired mitochondrial utilisation of available substrates, including oxygen, may coexist to generate this imbalance. all these mechanisms could contribute to atp depletion and na + /k + atpase failure, leading to cytotoxic oedema in cm. cm is clearly associated with increased neuronal activity. a recent review identified that % of african children with cm have a history of seizures, with prolonged and recurrent seizures associated with a poor outcome [ ] . impaired vascular flow during acute cm may limit substrate delivery within the brain and contribute to energy imbalance. in the past, a common premise was that parasite sequestration precipitated cerebral vaso-occlusive/ischaemic (i.e. stroke-like) events that manifested clinically as cm. however, cm demonstrates several features that are atypical for stroke. in children, focal neurological signs do not tend to accompany coma, although a sub-set of patients do exhibit hemiparesis or focal brainstem deficits during the agonal period [ ] . the incidence of residual neurological deficits following recovery from coma is relatively low ( % [ ] ) when compared to childhood stroke ( % had residual neurological deficit [ ] ). where computer tomography has been performed in children, diffuse brain swelling was observed [ ] rather than focal lesions more typical of stroke. although retinal haemorrhages have been observed in % of malawian children with cm (and in % of patients who died), these lesions were also seen in % of children with sma in the same study [ ] . consequently, although associated with cm, retinal haemorrhages do not confirm that focal cerebral vaso-occlusive/ischaemic events underlie cm. similarly, histological examination of fatal cm cases of african children at autopsy demonstrated that one third had little or no evidence of local vascular change in the brain, as indicated by sequestered parasites, monocyte clusters, micro-haemorrhages, local vascular inos [ ] or haemoxygenase - (ho- ) [ ] staining. accepting that cm may occur without ischaemia does not exclude temporary or less severe reductions in vessel flow occurring during acute cm (associated or independent of parasite sequestration) that may contribute to impaired substrate delivery and lead to energy imbalance. as previously discussed, energy imbalance may also be impaired due to the uncoupling action of inflammatory cytokines on mitochondrial atp production. in gambian and ghanaian children, concentrations of tnf and its receptor were higher in those with cm than in those with mild or uncomplicated malaria [ , ] . polymorphisms in the tnf promoter region have also been associated with increased risk of cm and death [ ] or neurological sequelae [ ] . cytokines may also up-regulate inos in brain endothelial cells, increasing production of no, which could then diffuse into brain tissue and disrupt neuronal (and/or mitochondrial) function [ , ] . in the brain, mitochondrial function may also be influenced by neuronal excitotoxins. within the simplified model of dissociated neuronal culture, mitochondria appear to play a critical role in neuronal homeostasis during excitotoxin exposure. mitochondria are not only involved with maintaining atp production but also calcium homeostasis, and generation and detoxification of reactive oxygen species [ ] . excitotoxin production may also be influenced by cytokine release. tnf administration has been shown to alter brain metabolism of tryptophan to produce more kynurinine [ , ] . thus, as part of a general inflammatory reaction, increased excitotoxin generation during acute malaria may contribute to cellular energy imbalance. elevated levels of neuronal excitotoxins (quinolinic and picolinic acid) in the csf have been associated with a fatal outcome in malawian children with cm [ ] . similarly, a graded increment of quinolinic acid concentration in csf was observed across patient outcome groups of increasing severity in african children [ ] . although a subset of the malawaian autopsy patients [ ] demonstrated negligible histological change in their brains, they did demonstrate inflammation, as indicated by inos, mif [ ] and ho- [ ] , staining in other tissues. these systemic changes were shared with the comatose sepsis cases in the study, and therefore are consistent with the premise that coma may in part be secondary to a host inflammatory response to systemic infection. below are further examples of systemic responses to infection that present with diffuse cerebral syndromes, including coma. in the past, the term cm has been restricted to falciparum malaria, and patients with p. vivax infection exhibiting symptoms of severe malaria, including coma, have been dismissed as undiagnosed falciparum co-infections. however, the use of more sensitive diagnostic techniques makes such dismissal less tenable. two such studies report adults exhibiting severe malaria with p. vivax (but not p. falciparum) infection detectable on pcr and serological and testing [ , ] . the patients exhibited multiple organ failure including cerebral symptoms, renal failure, circulatory collapse, severe anaemia, haemoglobinuria, abnormal bleeding, acute respiratory distress syndrome, and jaundice. vivax malaria has been associated with a strong systemic inflammatory response [ ] , but this was not investigated in the above studies. sepsis-associated encephalopathy (sae) syndrome has multiple features that resemble cm. it is characterised by a diffuse disturbance of cerebral function (typically impairment of consciousness) that occurs in the context of systemic response to infection without direct neuroinvasion (i.e. meningitis, macroscopic cerebritis and brain abscesses are excluded). sae is associated with generalised slow waves on the electroencephalogram (eeg), with the depth of coma linked with mortality. mild sae cases often recover completely, while survivors of severe sae may have persistent neurological deficit [ ] ). in line with adult cm, the severity of encephalopathy parallels the severity of systemic organ failure [ ] . inflammatory cytokines have been demonstrated to be higher in the serum than in the csf, suggesting that sepsis encephalopathy is a consequence of the systemic inflammatory response to infection [ ] . an animal model in which prior administration of a neutralising antibody to tnf prevented the sepsis encephalopathy of pancreatitis [ ] is consistent with this. further postulated reversible mechanisms of pathogenesis include changes in regional cerebral blood flow, neurotransmitter imbalance, mitochondrial dysfunction, bbb impairment and oxidative stress [ ] . severe influenza infection can present with encephalopathy, yet as in malaria, the pathogen is not neuroinvasive [ ] . seizures and coma occur after high fever [ ] , commonly accompanied by thrombocytopaenia [ ] , with metabolic acidosis and hyperlactataemia in severe cases (t. ichiyama, personal communication). similar to adult malaria, neurological sequelae occur concurrently with multiple organ failure [ ] . tnf, il- , stnfri, and soluble e-selectin are increased in serum and csf [ , ] , and serum nitrite/nitrate levels are increased [ ] . detailed examination of brain has revealed apoptosis of neurons and glial cell, histological evidence of active caspase- and caspase-cleaved parp, cerebral oedema, and bbb impairment [ ] . these parallel changes are set out in table . it is clear, therefore, that the presence of sequestering parasitised red cells is not necessary to generate these changes, which are also demonstrable in the falciparum malaria encephalopathy. notably, high levels of inflammatory cytokine are present in each disease. seizures are a very common component of acute malaria illness in children. a recent review documented that % of african children had a history of seizures, with % exhibiting seizures during hospital admission [ ] . the molecular basis of the seizures is unclear. multiple mechanisms have been postulated, including fever, hypoxia and/or cytokine stimulation leading to an imbalance of neurotransmitters and excitotoxins or neuronal damage [ , ] . recently, lang and co-workers [ ] demonstrated that falciparum parasitaemia is associated with the generation of specific antibodies for voltage-gated calcium channels directed against neurones. higher antibody concentrations were detectable in sera from patients exhibiting cm or malaria with seizures than uncomplicated malaria, suggesting that these antibodies may influence seizure propensity. only the erythrocytic form of malaria is associated with disease, so valuable information about which african children are likely to have more, or less, severe malaria has inevitably been obtained from examining the inborn rbc abnormalities that endemic malaria has selected across the tropics. the coinciding geographic distributions of malaria transmission and the thalassaemias prompted haldane to put forward the 'malaria hypothesis', which proposed that common erythrocyte abnormalities are selected because of the fitness advantage they confer against malaria [ ] . sickle cell haemoglobin (hbs) has also been repeatedly shown to be associated with malaria resistance, with heterozygotes for the hbs trait demonstrating % of the population at risk for severe malaria in certain populations [ ] . other haemoglobinopathies (e.g. hbc [ , ] and hbe [ ] ) and deficiencies in rbc enzymes (e.g. glucose- -phosphate dehydrogenase deficiency [ ] ) have also been linked with protection against severe malaria. the mechanisms of protection afforded by haemoglobinopathies are likely to be multi-factorial. studies have demonstrated evidence to support several independent mechanisms including: reduced parasite invasion of rbcs and diminished intraerythrocytic growth of parasites in patients with the hbs trait [ ] , enhanced phagocytosis of parasite-infected erythrocytes (ies) [ ] and enhanced immune responses against ies [ ] . recent in vitro studies observed that hbc modifies the quantity and distribution of the variant antigen p. falciparum erythrocyte membrane protein (pfemp ) on the ie surface. pfemp has been implicated in numerous ie adhesive interactions. in the latter study the authors demonstrated that hbc reduces the level of ie adhesion to endothelial monolayers, in addition to ie rosetting (the adhesion of ies to uninfected erythrocytes) and ie agglutination by sera. these findings provide the prospect that hbc pro-tects against severe malaria by mitigating the obstruction and inflammation caused by the pfemp -mediated adherence of ies [ ] . however, sequestration is believed to enhance parasite survival by enabling ies to avoid splenic clearance, so any reduction of sequestration by hbc can be expected to limit parasite fitness. multiple epidemiology studies (e.g. [ , , ] ) have failed to identify any significant impact of hbc on the frequency or density of parasitaemia in naturally exposed populations. consequently, the influence of the changes in ie surface conformation needs to be confirmed and further examined in vivo [ ] . a recent study re-confirmed that african children with -thalassaemia trait are significantly less likely to be hospitalised with severe malaria, particularly with coma or severe anaemia (hb < g/ ml). it is intriguing that the -thalassaemia patients did not demonstrate a lower incidence of uncomplicated malaria nor any reduction in peripheral parasite density [ ] . thalassaemia has also been associated with increased incidence of clinical vivax and falciparum malaria during early life [ ] . the findings raise speculation that the trait may indirectly afford enhanced immunity through increased non-lethal exposure to malarial parasites. such a mechanism is appealing, since it would be equally plausible across a range of haemoglobinopathies, including hbc. variations in erythrocyte membrane proteins also have a profound influence on malaria susceptibility. most notably the absence of duffy antigen protein confers absolute protection to p. vivax infection. more recently, the duffy antigen has also been associated with a protection against falciparum malaria [ ] . enzymes involved with iron handling may also have a critical influence on malaria morbidity. a recent study from the gambia demonstrated that children in an endemic malaria area possessing the haptoglobin , , isotype had a significantly increased risk of anaemia [ ] . however, a lack of parallel alterations in other haematinic indices leaves the mechanism of this process unclear. malarial protection within individuals exhibiting multiple rbc abnormalities appears even more complex. a recent study observed that the concurrent presence of sickle cell and -thalassaemia trait among african children had a negative influence on the risk of malaria infection [ ] . the results warn geneticists that gene epistasis may have a profound influence on overall malarial susceptibility. in tropical countries many hospital deaths from falciparum malaria happen before anti-malarial drugs have had time to kill the parasites. two approaches could help rectify this -addressing public-health problems resulting in delayed presentation, and identifying the physiological processes and molecular pathways that lead to these early deaths, with a view to developing evidence-based adjunct therapies. therapies being explored in sepsis, and based on disease pathogenesis data common to sepsis and malaria, may prove to be transferable from either of these diseases to the other. as noted above, circulating levels of a late-appearing inflammatory cytokine, hmgb , are increased in falciparum malaria [ ] as well as in sepsis. results from animal models on the role of hmgb , although untested in humans, have inspired enthusiasm for inhibition of this molecule as a potential intervention for human sepsis. for instance, anti-hmgb antibodies provided dose-dependent protection [ ] and reduced mortality [ ] against experimental sepsis in mice. late administration of ethyl pyruvate, which inhibits hmgb release from macrophages, also conferred protection against endotoxaemia in mice [ ] . treatments directed towards critical downstream consequences of malaria infection and inflammation, such as those intended to limit acidosis, are also a focus of investigation. one current approach is to identify which acute malaria patients most benefit from early volume expansion [ ] . controlling lactic acidosis via sodium dichloroacetate (dca), an inhibitor of pyruvate dehydrogenate kinase (maintaining pyruvate dehydrogenase in its active form), is also being examined. dca reduced lactate levels in acute malaria patients [ ] , although the study was unable to determine whether treatment improved outcome. an earlier large sepsis study also demonstrated that dca reduced lactate, but again with no improvement in outcome [ ] . as outlined in the section 'is hyperlactataemia a cause or marker of the acidosis of malaria?', some researchers argue, in view of the strong ion difference contributing to acidosis and the postulated mitochondrial dysfunction during acute malaria infection, that lactate reduction per se may have limited impact on prognosis. other adjunct therapies are also being examined. improving rbc deformability provides one potential therapeutic approach. in vitro studies with n-acetylcysteine (nac), reported to scavenge free radicals, showed improvement in red cell deformability through in vitro studies [ ] . unfortunately, an initial in vivo trial of nac in malaria patients had no effect on mortality [ ] . blocking endothelial activation is also a focus of research, with initial in vitro studies providing some encouraging results [ ] . in conclusion, continuing to identify the host responses to malaria infection that lead to disease is providing insights into novel molecular mechanisms. this information is beginning to guide the design of much needed additional therapies against this disease. there is little doubt that poor oxygen supply through vascular occlusion or anaemia could contribute to the body relying on excessive glycolysis to generate energy, resulting in hyperlactataemia, hypoglycaemia, and metabolic acidosis, and altered consciousness. however, inflammatory cytokines control these changes, as well as inhibit the capacity of mitochondria to use oxygen. thus, as described throughout this review, inflammatory cytokines are likely to have various pivotal roles across the multiple pathological processes involved in malarial disease (fig. ). indicators of life-threatening malaria in african children the pathophysiologic and prognostic significance of acidosis in severe adult malaria malaria and the lung indicators of mortality in african adults with malaria severe falciparum malaria in children: 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prevents lethality in mice with established lethal sepsis and systemic inflammation randomized trial of volume expansion with albumin or saline in children with severe malaria: preliminary evidence of albumin benefit pharmacokinetics and pharmacodynamics of dichloroacetate in children with lactic acidosis due to severe malaria oxidative stress and rheology in severe malaria a pilot study of n-acetylcysteine as adjunctive therapy for severe malaria inhibition of endothelial activation: a new way to treat cerebral malaria key: cord- -icsey p authors: fernandez-botran, rafael; uriarte, silvia m.; arnold, forest w.; rodriguez-hernandez, lisandra; rane, madhavi j.; peyrani, paula; wiemken, timothy; kelley, robert; uppatla, srinivas; cavallazzi, rodrigo; blasi, francesco; morlacchi, letizia; aliberti, stefano; jonsson, colleen; ramirez, julio a.; bordon, jose title: contrasting inflammatory responses in severe and non-severe community-acquired pneumonia date: - - journal: inflammation doi: . /s - - - sha: doc_id: cord_uid: icsey p the objective of this study was to compare systemic and local cytokine profiles and neutrophil responses in patients with severe versus non-severe community-acquired pneumonia (cap). hospitalized patients with cap were grouped according to the pneumonia severity index (psi), as non-severe (psi < points) or severe (psi ≥ points). blood and sputum samples were collected upon admission. compared to non-severe cap patients, the severe cap group showed higher plasma levels of pro- and anti-inflammatory cytokines but in contrast, lower sputum concentrations of pro-inflammatory cytokines. blood neutrophil functional responses were elevated in cap patients compared to healthy controls. however, neutrophils from severe cap patients showed reduced respiratory burst activity compared to the non-severe group. results indicate that patients with severe cap fail to mount a robust local pro-inflammatory response but exhibit instead a more substantial systemic inflammatory response, suggesting that a key driver of cap severity may be the ability of the patient to generate an optimal local inflammatory response. electronic supplementary material: the online version of this article (doi: . /s - - - ) contains supplementary material, which is available to authorized users. community-acquired pneumonia (cap) is one of the major infectious disease-related causes of death in both developed and underdeveloped countries [ , ] . cap initiates with an inflammatory response elicited by resident alveolar macrophages in response to invading microorganisms [ ] . these cells produce a cascade of pro-inflammatory cytokines and chemokines that vigorously recruit and activate blood leukocytes, particularly neutrophils, into the lungs, where they play a major role in curbing the infection [ ] . in hospitalized cap patients, severity of disease is influenced by the ability of the host′s immune response to control the pathogenic microorganism, as well as through regulation of the local pro-inflammatory response and neutrophil accumulation. during the inflammatory response, neutrophils undergo sequential release of various granule subtypes and enhance respiratory burst activity with production of reactive oxygen species, with the potential to rafael fernandez-botran and silvia. m. uriarte contributed equally to this work. electronic supplementary material the online version of this article (doi: . /s - - - ) contains supplementary material, which is available to authorized users. cause tissue damage [ , ] . evidence suggests that failure to control excessive inflammation and/or neutrophil activation may result in exaggerated systemic inflammation and organ damage, leading to severe disease [ ] [ ] [ ] [ ] . data examining the association between local the inflammatory response and severity of disease are limited. since severe cap is associated with significantly higher mortality than non-severe cap, a better understanding of the local and systemic inflammatory responses in hospitalized patients with severe cap may be necessary to develop novel strategies for the management of these patients. the objectives of this study were to characterize and contrast the lung and systemic cytokine profiles as well as blood neutrophil responses in patients with severe versus non-severe cap at the time of hospital admission. this was a prospective observational study of hospitalized patients with cap at the university of louisville hospital and the louisville′s veteran administration hospital from / / to / / . the university of louisville human subjects program protection office and the robley rex veterans affairs medical center institutional review boards approved this study prior to any data collection (approval nos.: . and , respectively). inclusion criteria/criteria for cap. patients with cap, defined as evidence of a new pulmonary infiltrate at chest radiograph associated with at least one of the following: ( ) new or increased cough; ( ) fever or hypothermia; and ( ) leukocytosis, left shift, or leucopenia, were included in this study following previous written consent. the inclusion and exclusion criteria and full case report forms for this study can be found at the community acquired pneumonia organization study site at www.caposite.com. exclusion criteria. unstable psychiatric or psychological condition rendering the subject unlikely to be cooperative or to complete the study requirements. patients were excluded if they had a medical history that, in the investigator′s opinion, precluded subject compliance with the protocol. immunosuppression was not part of the exclusion criteria. none of the patients included in this study was neutropenic (< . × neutrophils/mm ). healthy control group. in order to compare results of the plasma cytokines and neutrophil functional assays from cap patients with those of healthy individuals, blood samples were also obtained from a control group (n= ) of healthy adult donors (approved by the university of louisville′s irb # . ). microbiologic analysis. testing of sputum samples, blood cultures, tracheal aspirates, pleural fluid; serology for respiratory viruses and atypical organisms; and urinary antigens for legionella spp. and streptococcus pneumoniae was performed according to standard clinical practice. the identification of microorganisms and susceptibility testing were performed according to standard methods [ ] . severity of disease. as one of the most commonly used predictive scores [ ] , the psi was used in this study to define cap severity. hospitalized patients were considered to have severe cap if their psi was risk class ivor v ( points or higher). patients with a psi risk class of i-iii (< points) were defined as having non-severe cap. plasma samples. blood samples were obtained on the day of admission at the hospital. venous blood was collected using sodium citrate vacutainer tubes. following centrifugation at ×g for min, the plasma was separated by aspiration, aliquoted, and stored frozen at − °c until assayed. sputum samples. sputum samples were collected from a total of cap patients ( with non-severe, with severe cap) on the day of admission to the hospital. patients were instructed to rinse their mouth with water and collect any spontaneously-produced sputum into a -ml sterile disposable polypropylene centrifuge tube. the sample was kept at °c and transported to the laboratory within h of collection. sputum samples were processed following the method described by pizzichini et al. [ ] . upon receipt, sputum samples were diluted with an equal amount of a . % dithiothreitol (dtt) solution in pbs and incubated in a rocking platform for min in order to digest the mucus. an equal volume of sterile saline was added followed by additional min of incubation. the samples were then filtered through nylon gauze and centrifuged at ×g for min. the cell-free supernatants were then aliquoted and stored frozen at − °c until used for the measurement of cytokine levels. the concentrations of interleukin (il)- β, il- receptor antagonist (il- ra), il- , cxcl (il- ), il- , il- p , il- , interferon (ifn)γ, tumor necrosis factor (tnf)α, and cxcl (ip- ) in plasma and sputum samples were determined using milliplex map multiplex kits (emd millipore, billerica, ma). following thawing, plasma and processed sputum samples were centrifuged at , ×g for min and the supernatants used in the assay according to the manufacturer′s instructions. in order to control for the potential interference of the dtt contained in the sputum samples on the measurement of the different cytokines in the multiplex assay, we evaluated the effect of dtt by running samples of known cytokine concentrations in the presence and absence of dtt (at . %, the same final concentration in the sputum samples). for eight out of the ten cytokines measured (il- β, il- ra, il- , il- p , il- , ifnγ, tnfα, and cxcl ), there was little if any interference, with measurements in the presence of dtt being, on average, within ± % of controls. in the case of il- and cxcl , measurements in the presence of dtt showed average reductions of - % compared to controls. comparisons of cytokine levels in the sputum samples between the different patient groups (severe vs. non-severe cap) were performed only among similarly processed samples (e.g., in the presence of dtt). neutrophil functional assays were performed using whole blood samples. basal or formyl-methionyl-leucylphenylalanine (fmlf)-stimulated exocytosis were determined by measuring the plasma membrane expression of secretory vesicles (cd , clone e ) and specific granules (cd b, clone g f , both from biolegend, san diego, ca) on a facscalibur instrument (becton dickinson, franklin lakes, nj). following antibody treatment, red cells were lysed with bd red cell lysis buffer (becton dickinson), followed by two washes with . % sodium azide, and fixation in % paraformaldehyde before analysis. phagocytosis and phagocytosis-stimulated respiratory burst activity were measured using a flow cytometric assay based on the production of h o stimulated by the phagocytosis of staphylococcus aureus, as previously described [ ] . briefly, neutrophils ( × cells/ml) were incubated with ′, ′-dichlorofluorescein diacetate ( . μm final concentration) for min at °c. then, μl aliquots of the cell suspension were sampled before and min after the addition of opsonized, propidium iodide-labelled s. aureus (at a final concentration of bacteria/ml). after fixation in % paraformaldehyde, the cells were then analyzed for phagocytosis and h o production by flow cytometry on a facscalibur instrument. statistical analysis was performed using graphpad prism . (graphpad software, la jolla, ca) and r version . (www.r-project.org). all cytokine and neutrophil data are presented as the median and intraquartile ranges. data distribution was analyzed using the d′agostino and pearson omnibus normality test. statistical comparisons between the groups were performed using the mann-whitney test. correlation analyses were performed by the spearman′s method. p values of ≤ . were considered statistically significant. a total of hospitalized patients with cap were included in the study. non-severe and severe cap groups were made up of and patients, respectively. the main demographic, medical history, and clinical/laboratory characteristics of the two groups are summarized in tables and . there was a statistically significant difference (p= . ) in the ages of the two groups, with patients in the severe cap group having a median age of . years vs. . years for the non-severe cap group. no differences in gender or residence at a nursing home were observed. cerebrovascular disease, congestive heart failure, and copd were more frequent in the severe cap group, but there were no significant differences with other underlying conditions. blood urea nitrogen levels (p= . ) and peripheral blood neutrophil counts (p= . ) were higher in the severe cap group. in-hospital or -day mortality and re-hospitalization were not significantly different between the groups. patients in the non-severe cap group were more likely to have a time to clinical stability (tcs) of less than days compared to the severe cap group ( vs. %, respectively; p= . ). the etiologic agent could not be identified by standard laboratory procedures in approximately half of the patients in each group ( patients each) and in approximately a quarter of the cases in each group ( patients), the etiologic agent was identified as s. pneumoniae (table ) . the plasma cytokine profiles for both groups are shown in fig. (data in table form in supplementary material- table ). generally, patients in the severe cap group showed a pattern with median plasma concentrations of both pro-and anti-inflammatory cytokines that were higher in comparison with the non-severe cap group and the healthy control group. the severe cap group of patients had median plasma concentrations of il- , tnfα, and cxcl that were approximately twice as high as those of the non-severe cap group. moreover, while most patients in either group had undetectable concentrations of il- β in their plasma (≤ . pg/ml), when measurable, il- β tended to be higher in the severe cap group. the differences in the median concentrations in the two groups were statistically significant (p< . ) for all of the above cytokines. no statistically significant differences were found for cxcl or il- p . in the case of the anti-inflammatory iqr interquartile range, pao partial oxygen pressure, bnp brain natriuretic peptide, bun blood urea nitrogen, crp c-reactive protein, wbc white cell blood count, rr respiratory rate, hr heart rate, bp blood pressure, tcs time to clinical stability, los length of stay, psi pneumonia severity index cytokines, il- ra and il- , a similar pattern was observed, with the median concentrations for these cytokines being three to four times higher in the severe cap group (p< . ). finally, the median concentrations of adaptive immunityrelated cytokines, ifnγ and il- , were also four to five times higher in the severe cap group (p< . ). the sputum cytokine profiles are shown in fig. (data in table form in supplementary material- table ). the concentrations of cytokines in the sputum were not only much higher, but their patterns did not mirror those observed in the plasma. when related to the severity groups, the patterns in sputum were opposite to those seen in plasma for several of the cytokines, with the median values being significantly higher in the non-severe cap group, particularly for the proinflammatory cytokines. for example, median concentrations of il- β, il- , and tnfα were, respectively, -, -, and times higher than those in the severe cap group (p< . for il- β; p< . for il- ; and tnfα). however, no significant differences were observed for the anti-inflammatory cytokines. in the case of adaptive-immunity-related cytokines, the median concentrations of il- were found to be also higher in the non-severe cap group (p< . ). no differences were observed in the concentrations of ifnγ. table shows that the basal and fmlf-stimulated expression of both granule markers cd and cd b, as well as respiratory burst activity, were higher in the cap groups (both non-severe and severe) compared to healthy donors, indicating that their neutrophils were primed/pre-activated in the cap patients. comparison between the non-severe and severe cap groups showed a moderate but statistically-significant higher median basal level of cd b, with no differences in fmlf-stimulated expression in severe cap patients. in contrast, patients in the severe cap group had significantly lower median levels of respiratory burst activity compared to the non-severe cap group. analysis of potential correlations between plasma cytokine levels and neutrophil functional studies indicated a moderate (spearman′s r= . ) but statistically significant (p= . ) positive correlation between table . the neutrophil respiratory burst and plasma levels of cxcl when all patients′ samples were analyzed (fig. ) . this study indicates that, at the time of hospital admission, patients with severe cap have a decreased local inflammatory response and an exaggerated systemic inflammatory response when compared to patients with non-severe cap. this observation suggests that patients with severe cap may have a lower and suboptimal lung inflammatory response resulting in an inability to control the microbial invasion at the local level. in our analysis of the plasma cytokine response, we found that the patients with severe cap presented with significantly higher levels of both pro-and anti-inflammatory cytokines, arguing against a potential skewing of the systemic response towards anti-inflammatory or immunosuppressive cytokines. although it was expected that the higher systemic levels of pro-and anti-inflammatory cytokines would correlate with more severe pneumonia, we found that hospitalized patients with non-severe cap had significantly higher levels of pro-inflammatory cytokines (il- β, il- , and tnfα) and il- in the sputum. interestingly, no significant differences between the two groups were observed in the sputum concentrations of chemokines (cxcl and cxcl ) or anti-inflammatory cytokines (il- ra and il- ). taken together, these observations suggest that the local (sputum) pro-inflammatory response in hospitalized patients with non-severe cap was more robust than in hospitalized patients with severe cap. despite the activated status of neutrophils in hospitalized patients with cap compared to healthy controls, neutrophils in patients with severe cap showed moderately but significantly reduced levels of respiratory burst activity when compared to patients with non-severe cap. this is consistent with reduced bactericidal capacity and an inability to control bacterial infection. indeed, reduced neutrophil respiratory burst activity has been reported in cases of sepsis [ ] . the causes for the reduced respiratory burst in hospitalized patients with severe cap are not clear. nevertheless, the positive and statistically significant correlation found between neutrophil respiratory burst activity and the systemic levels of cxcl is interesting. a role for ros in the activation of signaling mechanisms, including stat- phosphorylation, involved in the secretion of cxcl has been reported [ ] . moreover, a reduced production of cxcl may also correlate with decreased ability to attract t lymphocytes, particularly the th subset, to the lungs, therefore further compromising the ability to adequately control infection [ ] . our study is in line with previous reports of positive associations between higher serum pro-inflammatory and anti-inflammatory cytokine levels with measures of pneumonia severity, such as the pneumonia severity index, crb, or crb- scores [ , [ ] [ ] [ ] [ ] [ ] [ ] . however, our findings of higher pro-inflammatory cytokine levels in the sputum of non-severe vs. severe cap patients have not been, to our knowledge, previously reported. a study by paats et al. [ ] found significantly higher levels of il- , cxcl , and ifnγ in the bronchoalveolar lavage (bal) fluids of pneumonia patients compared to healthy controls but no correlations with disease severity scores (psi or crb- ). however, it is not clear how closely cytokine levels in sputum correlate with those in bal. because of the heavy predominance of neutrophils in sputum compared to bal, cytokine levels in the former may be more representative of a neutrophil response. a separate study by lee et al. [ ] did find significantly higher levels of tnfα in bal from patients with severe cap who died versus those that survived. comparisons with our studies are not possible since all of the patients analyzed in lee′s study had severe cap. our study had relatively few numbers of patients and sputum samples, which limits the ability of our study to generalize, and prevented adjustment for confounding effects. another potential limitation of our study was the inclusion of cap patients with different microbial etiologies. it has been reported that different microorganisms may elicit different cytokine activation patterns in cap with the lowest inflammatory patterns being associated with cap of unknown etiology and the highest with s. pneumoniae and enterobacteriaceae [ ] . the use of the psi score to define severe cap is not without its limitations. a recent meta-analysis indicates that some of the most commonly-used scores such as psi and curb- , previously derived and validated to predict day mortality, have only moderate performance to predict icu admission [ ] . moreover, it needs to be remarked that several factors influencing the psi score may have direct effects on the inflammatory/immune response and, thus, be partially responsible for the observed results. our two psibased severity groups differed significantly in terms of age and several co-morbidities. aging is known to affect immune function at multiple levels, with both innate and adaptive immune responses being compromised with increased age [ ] [ ] [ ] . multiple studies, both in mice and humans, have analyzed the effect of senescence in macrophages and neutrophils, which are the main cells involved in the innate responses during cap. generally, macrophages and neutrophils from older individuals have been found to have deficits in the expression of toll-like receptors (tlrs), opsonization and phagocytosis, bacterial killing, chemotactic responses, and production of cytokines and chemokines [ ] [ ] [ ] . however, age by itself does not seem to be the only factor accounting for the differences observed in our study since comparison of elderly (> years old) vs. non-elderly patients showed no significant differences in neutrophil functional parameters (results not shown). another limitation to our study is the inclusion of patients with conditions potentially affecting their immunocompetence status. the presence of co-morbidities such as congestive heart failure, cerebrovascular diseases, and copd, which are characterized by the presence of chronic inflammatory responses, neoplasia, and therapeutic corticosteroid use, may result not only in several defects in the function of immune cells, but also in the activation of several immunoregulatory mechanisms, including immunosuppressive cytokines and myeloid and lymphocytic regulatory cells [ ] . further studies are needed to better understand the contribution of the immune defects associated with these co-morbidities to the local lung response in cap. in this regard, targeting of immunosuppressive mechanisms, particularly in the lung, may thus have a potential benefit for high risk cap patients. a common pathophysiological explanation of the inflammatory response in patients with severe cap proposes that, in response to invading microorganisms, first there is an initial exaggerated local inflammatory response. when the excessive local inflammatory response fills the pulmonary compartment, there is a spill of cytokines and inflammation into the systemic circulation, generating the systemic inflammatory response that is associated with severe cap. the data from our study seem to indicate that, an exaggerated local inflammatory response is not associated with severe cap, and on the contrary, patients with severe cap have a significantly lower pulmonary inflammatory response in relation to patients with non-severe cap. in agreement with the literature [ , [ ] [ ] [ ] [ ] [ ] [ ] , we found that patients with severe cap have a significantly higher level of systemic inflammation, but the initial step to reach this systemic inflammation may not be an excessive local pulmonary inflammation. it can be hypothesized that, patients who are able to produce an adequate local pulmonary response are able to contain the microorganisms, do not develop an exaggerated systemic response, and present with a non-severe cap. if a first response at the local pulmonary level is not sufficient due to some local level of immune deficiency, then a second response at a systemic level is elicited as the last attempt to control the infection. if this paradigm were to be correct, and patients with severe cap have dual inflammatory responses with a low pulmonary response and a high systemic response, then the therapeutic management of inflammation in patients with severe cap should be different for the lung and for the systemic circulation. financial disclosures. dr. uriarte was supported by nih grant k /r hl . trends in infectious disease mortality in the united states during the th century epidemiology of community-acquired pneumonia in adults; a population-based study neutrophil recruitment to the lungs during bacterial pneumonia leukocyte response and anti-inflammatory cytokines in community acquired pneumonia neutrophil granules: a library of innate immunity proteins structural organization of the neutrophil nadph oxidase: phosphorylation and translocation during priming and activation systemic cytokine levels in community-acquired pneumonia and their association with disease severity understanding the roles of cytokines, neutrophil activity and neutrophil apoptosis in the protective versus deleterious inflammatory response in pneumonia subpopulation of hyperresponsive polymorphonuclear neutrophils in patients with adult respiratory distress syndrome. role of cytokine production contribution of neutrophils to acute lung injury manual of clinical microbiology prediction of severe community-acquired pneumonia: a systematic review and meta-analysis indices of airway inflammation in induced sputum: reproducibility and validity of cell and fluid-phase measurements priming of the neutrophil respiratory burst involves p mitogen-activated protein kinase-dependent exocytosis of flavocytochrome b -containing granules a controlled study of leukocyte activation in septic patients tlr -triggered reactive oxygen species contribute to inflammatory responses by activating signal inducer and activator of transcription- ip- mediates selective mononuclear cell accumulation and activation in response to intrapulmonary transgenic expression and during adenovirusinduced pulmonary inflammation circulating cytokines as markers of systemic inflammatory response in severe community-acquired pneumonia systemic and bronchoalveolar cytokines as predictors of in-hospital mortality in severe community-acquired pneumonia systemic cytokine response in patients with community-acquired pneumonia inflammatory biomarkers and prediction for intensive care unit admission in severe community-acquired pneumonia interleukin , lipopolysaccharide-binding protein and interleukin in the prediction of risk and etiologic patterns in patients with community-acquired pneumonia: results from the german competence network capnetz local and systemic cytokine profiles in nonsevere and severe community-acquired pneumonia cytokine activation patterns and biomarkers are influenced by microorganisms in community-acquired pneumonia effect of age on human neutrophil function innate immunity in aging: impact on macrophage function reduced neutrophil chemotaxis and infiltration contributes to delayed resolution of cutaneous would infection with advanced age new insights into chronic inflammation-induced immunosuppression key: cord- -zfudssm authors: fong, i. w. title: emergence of new tickborne infections date: - - journal: emerging zoonoses doi: . / - - - - _ sha: doc_id: cord_uid: zfudssm several tickborne infectious diseases such as lyme borreliosis, ehrlichiosis, anaplasmosis, babesiosis, and others have been expanding to new endemic regions in the world for over a decade. moreover, new pathogens transmitted by ticks have recently been recognized in animals and humans from diverse regions of the globe, widely separated in distance. these include new phleboviruses of the bunyaviridae family, exemplified by severe fever with thrombocytopenia syndrome virus [sftsv] recognized in china in , and the heartland virus, a closely related but distinct virus, presenting with similar clinical features and discovered in missouri in . other newly recognized tickborne infections include a novel spirochete of the relapsing fever group, borrelia miyamotoi, first reported to cause human infection in russia in and subsequently discovered to cause clinical disease in the netherlands, japan, and the united states, with transmission by the black-legged deer tick ixodes scapularis. in europe a new tickborne disease, neoehrlichiosis caused by candidatus neoehrlichia mikurensis belonging to the anaplasmataceae family, has been described recently. furthermore, new tickborne rickettsial infections continue to be recognized in europe such as tickborne lymphadenopathy identified in and caused by rickettsia slovaca. novel tickborne infectious diseases will continue to emerge worldwide for the foreseeable future and be a challenge to the health of human populations. innovative methods of prevention for a broad variety of tick-transmitted diseases are needed, and one approach is to develop a universal tick vaccine that can be given to animal hosts or humans. there are at least species or subspecies of ticks recognized [ ] . these comprise of two major families: the ixodidae or hard-ticks because of their sclerotized, hard dorsal plate, which represent the most important group medically and numerically, and the argasidae or soft-ticks because of their flexible cuticle. a third family consisting of a single species, the nuttalliellidae, is confined to southern africa [ ] . ticks have three stages in their life cycle: the larval, nymphal, and adult forms. hard-ticks and soft-ticks differ morphologically and in their life cycle and ecology. ixodid ticks have several advantageous attributes for transmitting infectious pathogens as vectors more than soft-ticks or argasids. ixodid ticks feed for long periods [several days], firmly attached to the host, and remain unnoticed as their bite is painless, and their feeding hosts include a large variety of vertebrates in diverse habitats [ ] . argasids, conversely, feed briefly and often on a single host species, and they live in dry areas, mostly in sheltered habitat near their hosts [ ] . it is believed that ticks evolve about million years ago in the paleozoic or early mesozoic eras, initially parasitizing reptiles [ ] . ixodids or hard-tick adult size averages - mm, females longer than males, and the larva is about mm with progressive increase in size to the nymph, which is under cm, and then to the adult form [ ] . each stage of the tick attaches and feeds on a single host over several days; then once satiated it detaches and finds a resting place to molt to the next stage. the larva and nymph are the main transmitters of diseases, as the adult feeds only briefly and the males may not feed at all. ixodid ticks actually spend > % of their life cycle unattached from the host and mainly live in open areas of meadows but are transported to woods by their hosts [ ] . these ticks exhibit active host-seeking behavior by climbing plants and attaches to passing animal hosts or emerge from their habitat and attack nearby animals [ ] . the life cycle of the ixodid ticks usually extends to years but may vary from months to years, depending on the environmental conditions such as temperature and humidity [ ] . some tick species are host specific, feeding only a limited variety of animals, but this may vary with the stage, and some ticks have different host for each feeding stage. in general the animals in their habitat influence host selection and diverse species of ticks have different affinities for attacking humans [ ] . the relationship with ecology or environment, natural animal inhabitants and host specificity or range, and geographic distribution are all interrelated with respect to the life cycle of these ticks. the brown dog tick, rhipicephalus sanguineus, for instance, is well adapted to the vegetation and climate in the mediterranean region and other areas with similar conditions such as mexico, arizona, california, texas, and parts of brazil, all regions where the brown dog tick can transmit rickettsial pathogens. each feeding stage of r. sanguineus has high specificity, and it readily feeds on dogs and is important vectors for mediterranean spotted fever in the mediterranean region, and occasionally rocky mountain spotted fever in southwestern us and mexico. in contrast, ixodes scapularis [black-legged deer tick] in the us and ixodes ricinus [european sheep tick] in europe are adapted to the woods and forests with relative high humidity and are sensitive to desiccation from dry places. ticks inhabiting wooded areas, i. scapularis, i. ricinus, amblyomma american [lone star tick], and others, feed on different host species, such as small and large mammals and birds [ , ] . the life cycle of ixodid wooded ticks, as exemplified by i. ricinus, starts with the gravid female laying - of eggs on the grass, and the larvae hatch in - weeks, which attach and feed on small mammals [rodents] and birds, become engorged, detach in grass, and molt to adults in - weeks and which then attach and feed on small or large mammals and birds, then the engorged female mates with males to continue the cycle [ ] . soft-bodied ticks [argasids] are inhabitants of sheltered environments, nests, caves, burrows, and primitive man-made shelters. unlike ixodids, they lack cement from the salivary glands for firm attachment but produce anticoagulants and cytolytic substances to facilitate brief multiple feeding times [ ] . thus, the time spent on the host is relatively short, up to a few hours, and after meals they can be found in cracks and crevices or below the soil surface in their home environment. ticks have been recognized to inflict bites on humans for thousands of years by ancient greek scholars [ ] . their ability to transmit infectious disease was first recognized in animals; cattle fever caused by tickborne protozoan [babesia bigemina] was described in texas at the end of the nineteenth century [ ] . subsequently, within the first decade of the twentieth century, ticks were implicated in the transmission of microbes to humans. borrelia duttoni, transmitted by a soft-bodied tick [ornithodoros moubata], was found to cause relapsing fever in [ ] , and in rocky mountain spotted fever agent, rickettsia rickettsii, was shown to be transmitted by the wood hard-tick dermacentor andersoni [ ] . although mediterranean spotted fever was first described in tunis in [ ] , it was not until the s that the brown dog tick [rhipicephalus sanguineus] was recognized as the vector of the causative rickettsia conorii [ ] . tularemia was recognized in squirrels in and the first human case may have been described in japan in [ ] , but the first documented human case was reported in [ ] . however, the epidemiology and the role of diverse ticks in the transmission of francisella tularensis were not reported until [ ] . in the era following world war ii, many tickborne diseases of animal and humans were discovered of bacterial, viral, and protozoan etiologies [ ] . q-fever, caused by coxiella burnetii, was first described in australia in [ ] , and although the organism can be found in > species, it rarely is a vector-transmitted disease [most commonly transmitted by inhalation or ingestion], and tick transmission was initially reported in [ ] . lyme borreliosis, named after the town old lyme in connecticut [us] in , was discovered to be caused by borrelia burgdorferi in and transmitted by ixodid ticks [ ] , the black-legged deer tick i. scapularis in the northeastern and . historical aspects upper midwestern us and the western black-legged tick [i. pacificus] along the pacific coast. a similar condition was described in europe more than years before, with a rash of "erythema chronicum migrans," which was subsequently found to be caused by borrelia garinii, b. burgdorferi, and borrelia afzelii and transmitted by ixodid ticks [i. ricinus] [ ] . lyme borreliosis is the most common vector-transmitted disease in the us and europe, but also occurs widely in other countries of the former soviet union, china, japan, australia, and probably in north africa [ ] . the main reservoir of lyme borreliosis are small mammals, especially rodents such as the white-footed mouse [peromyscus leucopus] in northeastern us and apodemus species in europe [ , ] . although deer are hosts for the black-legged ticks, they are not reservoirs for lyme borreliosis. in europe b. afzelii life cycle is maintained in rodents and b. garinii in avian reservoirs [ ] . rickettsiosis, caused by intracellular bacteria, has been recognized to be expanding globally for several decades and represent one of the major zoonoses [ ] . prior to only four tickborne rickettsiosis were known with one tickborne spotted fever group identified in separate regions: rocky mountain spotted fever due to r. rickettsii was first described as a clinical entity in in the americas; r. conorii present in europe, southeast asia, and north africa causing mediterranean spotted fever; r. siberia recognized in siberia and western russia; and r. australis found in australia [ ] . the members of the rickettsia genus are classified into four groups: the spotted fever group rickettsiae, typhus group rickettsiae, rickettsia bellii group, and rickettsia canadensis [ ] . the recognition and expansion of pathogenic rickettsia have been facilitated by molecular techniques in recent times. rickettsiae once considered nonpathogenic to humans and new species have been identified in the past years. there are now rickettsia species validated in addition to subspecies mentioned. some new species identified since include rickettsia asiatica, rickettsia heilongjiangesis, rickettsia hoogstraalii, rickettsia raoulti, and rickettsia tamourae [ ] . tickborne rickettsiosis is now present in all continents of the world and some caribbean and pacific islands. rickettsia africae which causes african tick-bite fever is believed to have been exported from africa during the slave trade of the s to several caribbean islands, guadeloupe, st. kitts, nevis, dominica, us virgin islands, montserrat, st. lucia, martinique, and antigua [ ] . tickborne infections of humans, farm, and domestic animals are primarily associated with wildlife animal reservoirs. humans and domestic animals are incidental hosts that result from infringement of the usual circulation between wildlife and tick vectors in their natural habitats. the risks of tickborne diseases vary geographically and are determined by the climate, environment, presence of rodents and other vertebrate reservoirs, and the species of ticks parasitizing wild and domestic animals [ ] . these zoonoses can emerge in previously non-endemic areas when climate conditions and circumstances favorable to the maintenance and transmission arise. this may include displacement of the hosts of vectors and reservoir animals by human expansion and development. birds are important reservoirs for several pathogens and act as vehicles for infected ticks and probably play an important role in the dispersal of tickborne zoonoses [ ] . although for each pathogen one or more tick vectors and several animal reservoirs may exist, various combinations of coinfection with different microbes may coexist, and studies have demonstrated mixed infection in ticks of - . % [ , ] . for example, the black-legged deer tick [i. scapularis], from the northeastern and upper midwestern us, can be coinfected and transmit multiple pathogens to humans such as lyme and myamotoi borreliosis, anaplasma, babesia, and powassan virus. concurrent infection by two or more tickborne microbes is fairly common and frequently detected in clinical and veterinary practice [ ] . coinfection with multiple pathogens may be transmitted by ticks simultaneously during the same blood meal or at separate times. the natural reservoir hosts of many microbes [b. burgdorferi, anaplasma phagocytophilum, and babesia microti] transmitted by ixodes ticks are rodents, which are often coinfected with multiple organisms and may transfer the microbes to the larvae or nymphs and then to animals and humans [ ] . simultaneous coinfection of several pathogens may result in diverse host responses, which can conflict and antagonize each other, allowing the pathogens to synergistically infect the host more successfully, with more prolonged and severe disease with multitude of clinical manifestations, than normally reported with a single pathogen infection [ ] . sequential infection with a new microbe may allow the establishment of a new infection which otherwise may not have occurred because of elimination by the host immune defense mechanisms [ ] . the majority of tickborne infections are transmitted during the course of the blood meal from contaminated tick saliva [i.e., b. burgdorferi, relapsing fever borreliae, and spotted fever rickettsiae], regurgitated midgut contents [b. burgdorferi], feces [coxiella burnetii], or coxal fluid in argasid ticks with relapsing fever borreliae [ ] . transmission from contamination of abraded skin or mucosa of the eye following crushing of the ticks with the fingers is possible. transmission by accidental ingestion of the arthropods is well known in animals and is possible to occur in humans. transmission of several tickborne zoonoses can occur by blood transfusion and sharing of needles and syringes from an unsuspected blood donor or drug abuser. different babesia species, for instance, have been reported to be transmitted from blood transfusions in both animals [dogs] and humans [ ] . the bunyaviridae family of viruses constitutes the largest group of rna viruses with more than identified [ ] . they are enveloped, spherical virions with a diameter of - μm, containing three single-stranded rna segments [negative sense], with no matrix proteins [ ] . [ , ] . presently there are at least viruses of the bunyaviridae family not assigned a genera or species, including bhanja virus, forecariah virus, and the kismayo virus [ ] . the bhanja serogroup viruses are most closely related to sftsv and the heartland virus, and members of this group have been isolated from ticks on livestock and wild animals in india, southern europe, and africa [ ] . the bunyaviridae are generally found in arthropods and wild animals, especially rodents, and some members can infect humans. the hantavirus is the only member of this family not transmitted by vectors, but by rodent excreta [ ] . the orthobunyavirus [i.e., la crosse virus] infects only mosquito vectors, the nairovirus [i.e., cchfv] is largely limited to ticks, and the phlebovirus [i.e., sandfly fever] is transmitted by sandflies and midges, but rvfv which is a member of this group can infect a wide range of arthropods and is primarily transmitted by mosquitoes [ ] . sfts was first recognized as a distinct clinical entity in henan province of china in . patients presented with high fever, gastrointestinal bleeding, abdominal pain, bloating, nausea, and vomiting, with low platelets and white blood count, elevated alanine and aspartate transaminases, and proteinuria [ ] . in , a novel bunyavirus designated huaiyangshan virus and subsequently changed to sftsv was isolated from patients with this syndrome [ ] . it is believed that the sftsv actually originated in china about - years ago [ ] . the sftsv and heartland virus are assigned in the phlebovirus genus, but they are different from other known phleboviruses. the phleboviruses consist of about antigenically distinct serotypes classed into two groups. the phlebovirus fever group is transmitted by the phlebotominae sandflies or mosquitoes and the uukuniemi group by ticks [ ] . although sftsv have limited sequence similarities to other viruses of the uukuniemi group, it was assigned to this subtype due to serological similarities and lack of small nonstructural protein on the m segment, and ticks are also the common vector [ ] . isolates of sftsv from different geographical regions share % genetic sequence similarity and are grouped into five sublineages a-e [ ] . lineage a constitutes isolates from animals, dogs, cats, goats, buffaloes, and cattle, with no geographical clustering pattern [ ] . segmented-genome viruses such as the bunyaviruses are capable of rapid recombination, which is associated with pathogenicity and transmissibility among vectors and hosts and increases the risk for new outbreaks [ ] . recently two strains of sftsv were found to have reassortment in the small rna segment, which may drive rapid changes in the in the virus [ ] . the basis of genetic diversity of sftsv, although not completely understood, may be explained by lack of proofreading function of its rna-dependent rna polymerase, with high mutation rate [about - substitution per site each year] during its replication [ ] . sfts as a distinct syndrome was first reported in rural regions in henan and hubei provinces of central china in [ ] . since then the disease has been found in province of china with over reported cases. there is evidence that the microbe is widely distributed in china and only a small proportion of infected subjects develop clinical disease. serosurveillance studies of populations in hilly regions of china showed that . - . % of people had sftsv antibodies [ ] [ ] [ ] . there is also evidence that the virus has been circulating naturally in some regions of china showing seasonal variance with most cases occurring from may to july, and clinical disease occurred mainly in older subjects [ %] [ ] . it has been estimated that the annual incidence of disease is about per , of the rural population [ ] . seroprevalence and asymptomatic viremia of sftsv in blood donors from endemic regions have also been reported. in a study of , blood donors, seropositivity ranged from . % to . %, but very low-grade viremia was detected in only two subjects [ ] . overall, sfts is mainly reported from rural areas of central and northeastern china from may to september, targeting farmers > years of age [ ] . subjects were predominantly affected from farming-related exposures and numerous domestic and wild animals were infected by sftsv [ ] . sfts was first reported outside china in north korea in [ ] and a subsequent fatal case was reported from south korea in [ ] . during sfts was diagnosed in patients in south korea, and phylogenic analysis of sftsv isolates from south korea and china was closely related [ ] . locally transmitted cases have also been identified in japan with cases recognized by , all living in western japan, and were > years of age, and there were six fatalities [ ] . phylogenetic analysis of the japanese isolates indicated that the genotype was independent from those in china. it has been estimated that sftsv was likely circulating unrecognized in animals and humans in korea and japan for sometimes before these reports. in , there were suspected cases of sfts in japan and were confirmed cases by pcr [ ] . the tick vector of sftsv is considered to be a widely distributed hard-tick of the ixodidae family, haemaphysalis longicornis, present in china and other countries of asia [ ] . in endemic areas of china, the prevalence of sftsv in these ticks collected from domestic animals ranged from . % to . %, and the rna sequences of viral isolates were very closely related to those in patients [ , ] . the virus is also detected at a lower rate [ . %] of another widely distributed tick, boophilus (rhipicephalus) microplus, in endemic and non-endemic areas [ ] . viral rna of sftsv was also detected in h. longicornis ticks at all stages of its life cycle, indicating both transstadial and transovarial routes of transmission and the ability of ticks to play a role as a vector and a reservoir of the virus [ ] . in a study from south korea, , ticks were examined for sftsv from nine provinces, and the minimum detection rate in h. longicornis was . %, with the highest prevalence in the southern region [ ] . in south korea sftsv was also detected in other hard-ticks at lower rates, ixodes nipponensis and amblyomma testudinarium [ ] , and in china the virus was also detected in mites from field mice and goats in endemic regions [ ] . common with many tickborne infections, not all patients with clinical disease have a history of tick bites, and in patients from china only % with sfts recall tick exposure [ ] . a variety of domestic and wild animals have been found to carry sftsv, at different rates in various endemic regions of china. seroprevalence in domestic animals ranged from in cattle - %, goats - %, chickens - %, dogs - %, and pigs - % [ , , , ] . low levels of viral rna were found in a small fraction [ . - . %] of the animals studied [ , ] . the viruses from animals and those from patients and ticks shared . % of the genomic sequences [ ] . many wild animals such as deer, hedgehog, weasel, possum, and some bird species are hosts for the vector ticks and could carry the virus. however, rodents are known reservoirs of many bunyaviruses, but the rates of infection of rats [ . - %] with sftsv are lower than in livestock [ , ] . hence, it appears that domestic animals may be the amplifying hosts of sftsv and are probably the main reservoir of the virus. transmission of sftsv is considered mainly from tick bites, but there is also evidence from multiple reports that the virus can be transmitted from human to human by direct contact with blood of infected patients [ ] [ ] [ ] [ ] [ ] . a cluster of cases in families/households have been reported to be transmitted by blood contact, and blood transfusion and laboratory accidents from handling infected blood are potential means of transmission of sftsv. although the pathogenesis of sfts is not fully understood, major strides have been made in the few years since its description in understanding the mechanisms of the disease. like other severe viral infections, cytokine and chemokine imbalance appears to be important in the pathogenesis, and severe, fatal cases of sfts usually demonstrate "cytokine storm." increased levels of tumor necrosis factor-alpha [tnf-α], interferon-gamma [ifn-γ], and ifn-induced protein- were associated with disease severity [ ] . sftsv gain entry to many human and animal cells, including macrophages and dendritic cells, by binding of the virus glycoprotein [gn/gc] to a receptor, the c-type lectin dc-sign [ ] . dynamic changes in viral load, t-cell subsets, and cytokines have been measured and analyzed in patients with sfts and correlated with outcome. high levels of peak viral rna load, serum liver enzymes, and serum interleukin [il]- and il- were associated with higher fatality rates [ ] . these markers declined within weeks of onset in survivors, and cd + t-cells were elevated early after infection, while hla-dr+ and ctla + t-cells elevated during the recovery phase of survivors [ ] . hence, high sftsv viral load, very low platelets, high transaminases, marked elevation of proinflammatory and anti-inflammatory cytokines, and activation of cd + t-cells were markers of severe disease and poor outcome. cytokine storm with suppression of cd + and cd + lymphocytes with progressive decline but higher b lymphocytes has also been reported in severe and fatal cases of sfts [ , ] . analysis of the immune response during the course of illness may also assist in understanding the pathogenesis of disease. in a study of confirmed cases of sfts and followed after convalescence, during the first week of illness, there was a loss of t, b, and nk lymphocytes which were subsequently restored, but severe disease was associated with slower recovery and lower humeral immunity [ ] . sftsv-specific igm antibody could be detected within days, peaked at weeks, and persisted for months. igg antibody could be detected in most patients within weeks, peaked at months, and persisted for at least years [ ] . there is also evidence that sftsv is capable of infecting monocytes and suppresses ifnbeta and nf-kappa b promoter activities, facilitating the virus replication in human monocytes by restricting the innate immune response [ ] . the virus can also disrupt type interferon signaling by the nonstructural protein-mediated sequestering of signal molecules [stats i and ii] into inclusion bodies [ ] . ifn-β production is an important host defense mechanism against viral pathogens, and inhibition of this response has been reported with other bunyaviruses [ , ] . limited pathological studies of fatal sfts cases have been reported and the characteristic feature is the presence of necrotizing lymphadenitis of lymphoid tissues [ ] . leucopenia and thrombocytopenia which are hallmarks of the disease were not due to bone marrow suppression or aplasia, but are likely related to peripheral destruction or sequestration [ ] . animal models are useful for elucidating the pathogenic mechanisms and development of new therapies for many human diseases. newborn mice and rats, especially kunming mice, are highly susceptible to sftsv, and infected mice demonstrate pathological changes with large areas of necrosis only in the liver, but the virus can be detected in numerous organs [ ] . interferon-α/interferon-β knockout mice are also highly susceptible to infection with the virus, with % mortality in - days after inoculation [ ] . the virus is found in numerous organs with heavy viral burden in mesenteric lymph nodes and spleen but no detectable histological changes. it appears that c /bl mouse may be a better model for sftsv infection, as the animals demonstrate leucopenia and thrombocytopenia similar to humans [ ] . moreover, histopathological changes were found in the spleen, liver, and kidney, but the spleen appears to be the primary target as viral replication could be demonstrated in this organ. the thrombocytopenia is probably caused by virus-bound platelets that underwent phagocytosis by splenic macrophages [ ] . nonhuman primates are considered the gold standard animal models for studying human disease pathogenesis, and a recent study has been reported by infecting rhesus macaques. sftsv infection of macaca mulatta did not result in severe disease symptoms or death but caused fever, thrombocytopenia, leucopenia, and elevated transaminases and cardiac enzymes [ ] . minor pathological lesions were found in the liver and kidney during the late stages of infection, and elevation of inflammatory cytokines was present in the blood. thus, infection of this primate model resembles mild sfts in humans. other animal studies assessed potential therapeutic and preventative interventions. the nonstructural protein of the s segment [sftsv/nss] fraction of sftsv appears to antagonize interferon and suppress the host's innate immunity to facilitate infection. however, immunization with recombinant sftsv/nss was ineffective in promoting virus clearance in infected c l/ j mice [ ] . in another study using a mouse model, antiserum from a recovered patient with sfts prevented lethal infection with the virus and improved clinical signs in nonlethal infection [ ] . other agents tested including steroids, ribavirin, and a site-i protease inhibitor were ineffective. the incubation period of sfts after a tick bite is estimated to be - days [ , ] and after exposure to infected blood as - days [ ] . the major stages of the disease recognized include an initial febrile flu-like illness, a second stage of multiorgan failure, and a subsequent convalescent phase [ , ] . the first stage of the disease is characterized by sudden onset of fever from to °c, headache, fatigue, myalgia, nausea, vomiting, and diarrhea, associated with high viral load, leucopenia, thrombocytopenia, lymphadenopathy, and elevation of transaminases and creatine phosphokinase. this stage may last for - days but may resolve after a week and enter a convalescent phase in patients with mild disease. the second stage occurs after days and in more severe disease progress to develop multiorgan failure, first involving the liver, then the heart, lungs, and kidneys, which can persist for - days or lead to death [ , ] . coma may occur in about % but as high as % and confusion is seen in - % [ , ] . a recent report of patients with sfts described development of encephalitis in % of cases, with a fatality rate of . % in this subgroup [ ] . in nonfatal cases the biomarkers start to decrease with decline of the viral load and improvement of the platelet count by day - , whereas in fatal cases the viral load, biomarkers, and thrombocytopenia continue to increase. the overall case fatality rate of reported cases in china averages at . % [ ] , but the initial report was - % [ ] . convalescence in survivors varies from days to days after onset of illness but in most severe cases occurred after days [ , ] . clinical symptoms improve and the laboratory abnormalities gradually return to normal after - weeks [ ] . prognostic factors for disease severity and outcome of sfts are related to host factors, clinical manifestations, and laboratory parameters. age is a key factor in the severity of disease and outcome, with older age carrying a worst prognosis [ , , ] . children rarely become infected [possibly from less tick exposures and more robust immunity] and those who become infected manifest mild symptoms with fever, malaise and gastrointestinal symptoms, and minor laboratory abnormalities [ ] . clinical features associated with adverse prognosis include neurological manifestations, acute respiratory distress syndrome, and disseminated intravascular coagulopathy [ ] . host immune responses and viral replication are important factors in determining clinical severity and outcome, and high viral load in blood at admission is associated with a worst prognosis and fatality [ ] . laboratory parameters associated with poor prognosis and fatality include hypoalbuminemia, hyponatremia, coagulation disturbance, elevated transaminases, elevated creatinine, decreased lymphocyte count, and elevated lactic dehydrogenase levels in the late stage [ , ] . diagnosis of sfts is based on epidemiological exposure risk and clinical features with presence of fever, thrombocytopenia, and leucopenia. laboratory diagnosis can be confirmed by viral nucleic acid test with polymerase chain reaction [pcr] or by serology. rapid diagnosis in acute illness is best accomplished with a reverse transcriptase [rt] pcr which is highly sensitive and specific [ ] . potential detection limit of viral rna copies/μl was achieved using quantitative real-time pcr with . % sensitivity and over % specificity [ ] . moreover, the quantitative pcr at acute diagnosis can assist in the prognosis based on the viral load [ ] . serological methods for diagnosis include conventional immunofluorescence and serum neutralization assays which are not helpful in acute cases or early diagnosis, and they are costly and require well-trained personnel [ ] . indirect enzyme immunoassay [eia] and double-antigen sandwich eia have been used to detect total antibodies or viral-specific igm and igg [ ] . a highly sensitive and specific eia utilizing a glycoprotein n from the nucleocapsid has been developed [ ] . in serological diagnosis acute and convalescent sera are best tested, thus providing a delayed diagnosis, although the presence of specific igm antibodies in acute disease can be diagnostic. management of patients with sfts is largely supportive with correction of fluid and metabolic disturbances and blood transfusion if necessary for significant blood losses. platelet transfusion may be required for severe thrombocytopenia [< × - /l] with significant bleeding, and appropriate antibiotics for suspected or proven secondary bacterial infections [ ] . mechanical ventilation may be needed in severe cases with ards or coma for airway protection and hemodialysis for severe renal failure. no specific therapy has been shown to be effective for sftsv infection. ribavirin was of interest as it had been considered effective or approved for treatment of other bunyavirus infections, such as rift valley fever virus and crimean-congo hemorrhagic fever virus [ , ] . while ribavirin has some in vitro activity against sftsv, it did not effectively reduce virus replication in pre-infected cells [ ] . moreover, ribavirin treatment of patients with sfts produced no significant clinical benefit and had no effect on platelet counts or viral loads [ ] . convalescent sera from recovered patients infected with sftsv have high neutralizing antibodies against the virus and may have a role in treatment of severe cases or postexposure prevention after contact with infected blood [ , ] . heartland virus infection was first described from a patient in missouri [us] in and the agent is a phlebovirus closely related to sftsv [ ] . phylogenetic and serological analysis revealed that heartland virus should belong to the bhanja group of viruses in the phlebovirus genus [ ] . bhanja viruses have been isolated from various species of hard-ticks and are divided into the african and eurasian lineages. amblyomma americanum [lone star tick] is the vector of heartland virus [hrtv] . investigations had revealed the presence of viable virus in a. americanum nymphs in a patient's farm and nearby conservation area, with > . % sequence identity to human strains [ ] . ticks probably become infected by feeding on viremic hosts during their larval stage and may transmit the virus to humans during the spring and early summer when nymphs are plentiful. serological investigations in wild and domestic animals in missouri have detected high antibody prevalence to hrtv in raccoon [ . %] , horse [ . %] , and white-tailed deer [ . %] , which suggest that these species are possible candidate reservoir hosts [ ] . antibodies were also found in dogs [ . %], but no hrtv was isolated from any animal sera or ticks. the original two index cases were male farmers, aged and years, who presented with fever, leucopenia, and thrombocytopenia [ ] . both patients survived without hemorrhagic complications or multiorgan failure. an additional five nonfatal cases were subsequently identified through active surveillance in missouri [ ] . more recently a fatal case of hrtv infection was described in -year-old male on a farm in tennessee [ ] . the patient had a history of multiple tick bites, with detectable tick on his body weeks prior to onset of illness, and had comorbid conditions of chronic obstructive lung disease and alcoholism. he presented with weakness, fever, and altered mental status, and tests showed persistent leucopenia, progressive thrombocytopenia, anemia, and elevated transaminases. his clinical course deteriorated on broad-spectrum antibiotics and subsequent hypotension, hypoxia, and renal dysfunction occurred before his death. autopsy findings were largely nonspecific and the bone marrow revealed myeloid hyperplasia and trilineage hematopoiesis. although this fatal case resembles severe cases of sfts, there was no evidence of necrotizing lymphadenitis as described in fatal cases of infection with sftsv, but the spleen demonstrated white-pulp depletion and scattered immunoblasts [ ] . the differential diagnosis of hrtv infection would include infections with borrelia, rickettsia, ehrlichia, anaplasma, viruses, thrombotic thrombocytopenia, and hematological malignancies. a subsequent fatal case has been described in oklahoma [ ] , bringing the total number of hrtv infection detected to date to nine cases with two fatalities from three states. however, more cases are likely to be recognized in the future from other regions of the us, as wildlife serological studies have determined that hrtv is widespread within the central and eastern us [ ] . of animals tested, were seropositive for the hrtv including deer, raccoon, coyote, and moose. thirteen states had seropositive animals from florida, georgia, illinois, indiana, kansas, kentucky, maine, missouri, new hampshire, north carolina, tennessee, texas, and vermont [ ] . similar to sfts, there is no known specific therapy for hrtv infection. management is primarily supportive, but doxycycline should be used initially in severe cases for treatable differential diagnoses [borreliosis, rickettsiosis, ehrlichiosis, etc.] until these conditions are excluded. there is no data yet on pathogenesis and animal models of hrtv infection. however, the mechanisms in disease pathogenesis maybe similar to that of sftsv since the two viruses are closely related. borrelia miyamotoi is a spirochete that is closely related to species that cause relapsing fever. it was first discovered in ixodes persulcatus ticks in japan in and subsequently documented in ticks and rodents in europe and north america, but was not recognized to cause human infection until reported from russia in [ ] . in , the first case of b. miyamotoi infection was described in the us with clinical manifestation of meningoencephalitis [ ] . subsequently two more cases were reported in the us presenting like human granulocytic anaplasmosis [ ] . cases of b. miyamotoi infection were also reported from new england [ ] , the netherlands [ ] , and japan [ ] . [ ] . whole blood pcr for presence of specific dna sequences of common tickborne pathogens and bmd were performed. among , patients tested, bmd cases were identified by pcr. most of the patients presented with fever, chills, marked headache, myalgia, or arthralgia, and % required hospitalization [ ] . elevated liver enzymes [ %], leucopenia, and thrombocytopenia were common, and symptoms resolved after doxycycline and patients treated with amoxicillin or ceftriaxone also improved. serology for the acute diagnosis of bmd was poor, as only of cases with circulating dna had b. miyamotoi igm and none with igg, but convalescent sera were reactive in %. in this study of acute febrile patients, bmd was confirmed less frequently than babesiosis and human granulocytic anaplasmosis. b. miyamotoi infection is transmitted by the black-legged deer ticks [i. scapularis] and has a similar range and distribution as lyme borreliosis. most cases of bmd occurred in july and august, suggesting transmission by larval ticks which have their peak activity in these months. lyme disease, babesiosis, and human anaplasmosis mainly occur in june and early july, when the nymphal deer ticks are most abundant [ ] . b. miyamotoi had been shown to undergo transovarial transmission and can be transmitted experimentally by larvae [ ] . in regions where tickborne infections are possible, with or without known tick bites/exposure such as in many parts of northeastern, north-central, and far western us, empiric treatment with doxycycline may be reasonable for acutely sick, febrile patients for possible borrelia, rickettsia, ehrlichia, and anaplasma infections until a diagnosis is confirmed. novel tickborne infections continue to emerge in distant unrelated regions of the world with worrisome frequency. new, previously unrecognized, pathogens from tick exposure will continue to be discovered in the future. this is further exemplified by the recent detection of a novel orthomyxovirus of the genus thogotovirus, labeled bourbon virus, associated with a fatal infection in a previously healthy male from kansas after tick bites in [ ] . the differential diagnosis of acute febrile illness after tick exposure is quite diverse and largely depends on the local region or geography at the time of exposure. for instance, in the us, there are tickborne diseases listed by cdc, and now there will be with the addition of bourbon virus [see table . ] tickborne diseases are theoretically preventable with appropriate clothing [long sleeve shirts and pants], avoidance of high-risk exposures, use of insecticides or acaricides to control tick population, or use of personal insect repellants. despite these known preventative measures, for decades tickborne diseases have continued to flourish with worldwide expansion and emergence. thus, new approaches for prevention of tickborne infections are needed. directed mass education of targeted atrisk populations [farmers, outdoor campers, hikers, etc.] in endemic areas may be of benefit to inform the public of the risk of these diseases and preventative measures, including large visible signs in these areas. development of specific vaccines for these various pathogens is not feasible nor would be cost-effective due to their relatively low frequency. however, it may be worthwhile to evaluate new strategies such as a vaccine to prevent tick bites that could be used to prevent many tickborne diseases. anti-tick vaccines could potentially reduce tickborne infections by reducing the tick burden or interference in host-tick-human transmission by vaccination of the vertebrate hosts [ ] . anti-tick vaccines would be environmentally safe, unlikely to select resistance as compared to insecticides, and can include multiple antigens to target a broad range of ticks [ ] . metagenetic analysis of fever, thrombocytopenia and leucopenia syndrome [ftls] in henan province, china: discovery of a new bunyavirus a new phlebovirus associated with 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presenting as human granulocytic anasplasmosis human borrelia miyamotoi infection in the united states a case of meningoencephalitis by the relapsing fever spirochete borrelia miyamotoi in europe human infections with borrelia miyamotoi borrelia miyamotoi disease in northeastern united states. a case series surveillance for lyme disease united states a relapsing fever group transmitted by ixodes scapularis ticks novel thogotovirus associated febrile illness and death hard tick factors implicated in pathogen transmission exposed and concealed antigens as vaccine targets for controlling ticks and tick-borne diseases key: cord- -lq tp z authors: khanafer, nagham; sicot, nicolas; vanhems, philippe; dumitrescu, oana; meyssonier, vanina; tristan, anne; bès, michèle; lina, gérard; vandenesch, françois; gillet, yves; etienne, jérôme title: severe leukopenia in staphylococcus aureus-necrotizing, community-acquired pneumonia: risk factors and impact on survival date: - - journal: bmc infect dis doi: . / - - - sha: doc_id: cord_uid: lq tp z background: necrotizing pneumonia attributed to panton-valentine leukocidin-positive staphylococcus aureus has mainly been reported in otherwise healthy children and young adults, with a high mortality rate. erythroderma, airway bleeding, and leukopenia have been shown to be predictive of mortality. the objectives of this study were to define the characteristics of patients with severe leukopenia at -h hospitalization and to update our data regarding mortality predicting factors in a larger population than we had previously described. methods: it was designed as a case-case study nested in a cohort study. a total of cases of community-acquired, necrotizing pneumonia were included. the following data were collected: basic demographic information, medical history, signs and symptoms, radiological findings and laboratory results during the first h of hospitalization. the study population was divided into groups: ( ) with severe leukopenia (leukocyte count ≤ , leukocytes/ml, n= ) and ( ) without severe leukopenia (> , leukocytes/ml, n= ). results: median age was years, and the male-to-female gender ratio was . . the overall in-hospital mortality rate was . %. death occurred in . % of severe leukopenia cases with median survival time of days, and in . % of cases with leukocyte count > , /ml (p< . ). multivariate analysis indicated that the factors associated with severe leukopenia were influenza-like illness (adjusted odds ratio (aor) . , % ci ( % confidence interval) . - . , p= . ), airway bleeding (aor . , % ci . - . , p= . ) and age over years (aor . , % ci . - . , p= . ). a personal history of furuncles appeared to be protective (or . , % ci . - . , p= . ). conclusion: s. aureus-necrotizing pneumonia is still an extremely severe disease in patients with severe leukopenia. some factors could distinguish these patients, allowing better initial identification to initiate adapted, rapid administration of appropriate therapy. since the description of necrotizing pneumonia due to panton-valentine leukocidin (pvl)-positive staphylococcus aureus by gillet et al. in , numerous cases have been reported worldwide [ ] [ ] [ ] [ ] [ ] [ ] [ ] . they mainly impacted otherwise healthy children and young adults, with a median age of years. they are attributed to either methicillin-sensitive or -resistant s. aureus strains of various genetic backgrounds, but have all pvl genes in common [ , ] . necrotizing pneumonia is characterized by rapid, extensive, bilateral pneumonia frequently evolving towards acute respiratory distress syndrome (ards), despite intensive medical interventions with mechanical ventilation and inotrope support. no specific treatment guidelines have been published so far, but the addition of toxin-suppressing antibiotics, such as clindamycin, linezolid and rifampicin, has been suggested [ ] [ ] [ ] . the overall mortality rate ranges from to % of cases [ , ] . the fatal outcome is rapid, with median survival time between to days from the onset of symptoms [ , , ] . it is associated with classic severity factors, such as the need for mechanical ventilation or inotrope support, and the onset of ards. erythroderma, airway bleeding, and leukopenia have been shown to be predictive of mortality. gillet et al. reported significant differences, by multivariate analysis, in median leukocyte count between patients who survived and those who did not. the survival rate was < % when the leukocyte count was < , leukocytes/ml. the leukopenia observed in patients could reflect pvl cytotoxicity, demonstrated in vitro on human neutrophils [ , ] . as patients with leukopenia face a high risk of mortality, better initial recognition of these severe cases would allow rapid administration of appropriate treatment. in this study, a series of cases of s. aureus-necrotizing pneumonia was analyzed. our aim was to define the characteristics of patients with severe leukopenia at -h hospitalization and to update our data regarding mortality predicting factors in a larger population than we had previously described [ ] . since , the french national reference centre of staphylococci (lyon, france) has collected case reports of documented pneumonia caused by s. aureus strains carrying pvl genes (luks-pv-lukf-pv). informed consent was waived because data were extracted from the surveillance database. according to french law, a study like this one does not require ethics committee approval because it is observational and based on a surveillance database approved under national regulations. the protocol design was approved by the hospital's institutional review board (comité national informatique et liberté). pneumonia was defined by signs and symptoms of lower respiratory tract infection (e.g., cough, expectoration, and chest pain) and pulmonary infiltrates on chest x ray reviewed by a radiologist, that were not attributable to other causes, but coinciding with s. aureus isolation by at least of the following procedures: ( ) pleural effusion or lung abscess; ( ) broncho-alveolar lavage fluid culture ( cfu/ml), wimberley brushing ( cfu/ ml), or protected tracheal aspiration ( cfu/ml); and ( ) blood culture revealing s. aureus as the sole potential pathogen. cases with respiratory symptoms starting at least h after hospitalization were classified as nosocomial and were excluded from the study. leukopenia was defined as severe if median leukocyte count was < , leukocytes/ml within the first h after hospital admission. s. aureus isolates were tested for antimicrobial susceptibility and toxin production. testing of isolates for antimicrobial susceptibility by broth microdilution was undertaken according to the interpretive criteria of the clinical and laboratory standards institute (formerly the national committee for clinical laboratory standards). the following antimicrobial agents were used: penicillin, oxacillin, kanamycin, tobramycin, gentamicin, erythromycin, clindamycin, tetracycline, ofloxacin, fusidic acid, rifampicin, vancomycin, teicoplanin, fosfomycin, trimethoprimsulfamethoxazole, and linezolid. gene sequences encoding superantigens (enterotoxins a-e, g-i, and toxic shock syndrome toxin), pvl and meca gene, which codes for methicillin resistance, were detected by pcr, as described elsewhere [ ] . only cases caused by pvl-positive s. aureus strains were included. designed as a case-case study nested in a cohort study, analyses were restricted to cases where all clinical and biological data were available. the following data were collected by a standardized form and comprised basic demographic information, medical history (including risk factors for infection and history of personal or familial abscesses or furuncles), signs and symptoms, radiological findings and laboratory results during the first h of hospitalization. severity was rated by pediatric risk of mortality (prism) scores for patients < years and the simplified acute physiology score (saps) ii for patients ≥ years, when available. some biological and radiological data were missing because of death shortly after admission to hospital. categorical variables were compared by the chi-square or fisher's exact test, and continuous variables, by student's t-test or mann-whitney tests. survival probability according to median leukocyte count, was estimated by the kaplan-meier method. initially the cases were divided into groups: - , ; , - , and > , leukocytes/ml but we merged the data of the last two groups since no significant difference was found for variables included in the final multivariate regression model (data not shown in this paper). the roc analysis showed a % sensitivity for a cut-off of leukocytes/ml. baseline was the day of admission to hospital because of pneumonia, and patients who survived were censored at hospital discharge. when patients died within h after admission, the observation period was rounded to day. survival distributions were compared by the logrank test. variables independently associated with survival were identified with a cox regression model based on hazard ratios with % confidence interval ( % ci). variables associated with severe leukopenia were tested by the multivariate logistic regression model. when p values of variables described in the first table, were < . in univariate analysis, they were submitted to the multivariate model. variables in multivariate analysis were subjected to the forced entry procedure, with stepwise and backward elimination, using p values of . as criteria for inclusion and elimination of risk variables based on best subset logistic regression with chi-square score fit criteria. the hosmer-lemeshow test assessed the model's goodness-of-fit. adjusted odds ratios (aor) and corresponding % ci were calculated. for all tests performed, -tailed p values < . were regarded as denoting statistical significance. analyses were performed with spss . software (spss inc., chicago, il, usa). from through , cases of communityacquired, necrotizing pneumonia were collected and documented; case reports were excluded because of missing data concerning leukocytes count. in total, cases of community-acquired necrotizing pneumonia were included. median age was years (interquartile range [iqr] . - . ) and the male-to-female gender ratio was . ( males and females). smoking was reported in . % patients. common risk factors for staphylococcal infection, such as diabetes, steroid therapy, and immunosuppressive treatment, were noted for . %, . % and . % of patients, respectively. among the patients for whom data were available, . % had a personal history of furuncles or skin abscess. the median duration of symptoms prior to hospitalization was . days (iqr . - . days), with preceding influenzalike syndrome in . % ( of ), and pre-existing skin and soft tissue infection (ssti) in . % ( of ). the clinical course during the first h after hospital admission was usually severe, with . % of patients requiring mechanical ventilation. the most remarkable clinical feature was airway bleeding, which occurred in . % of patients. s. aureus was recovered from blood culture in . %, in pleural fluid from . %, and in tracheal aspirates from . % of patients. median minimal leukocyte count during the first h of hospitalization was , leukocytes/ml (iqr , - , leukocytes/ml), and . % had leukocyte count ≤ , leukocytes/ml. the minimal platelet count was also low, with a median of , platelets/ml (iqr , - , platelets/ml). the overall in-hospital mortality rate was . %. . % of deaths were attributed to s. aureus and were secondary to refractory shock and/or respiratory failure. the study population was divided into groups: ( ) with severe leukopenia (leukocyte count ≤ , leukocytes/ ml, n= ) and ( ) without severe leukopenia (> , leukocytes/ml, n= ). the group with severe leukopenia was characterized by a significantly higher rate of female patients (p= . ) ( table ) . prior influenza-like illness (ili: . % vs. . %, p< . ) and airway bleeding ( . % vs. . % (p< . ) were also associated with severe leukopenia. conversely, a personal history of furuncles or skin abscess (p= . ) and ssti at admission (p= . ) were more frequent in the group without severe leukopenia. the group with severe leukopenia presented ards onset more frequently and needed artificial ventilation or inotrope support (p< . ). multivariate analysis indicated that ili, airway bleeding and age over years were independent factors associated with severe leukopenia (table ) . a personal history of furuncles appeared to be protective. leukocyte count was negatively correlated with mortality. death occurred in . % of cases ( of ) with severe leukopenia (≤ , leukocytes/ml) with median survival time of days (figure ). only . % of cases ( of ) with leukocyte count > , /ml (p< . ) died. mortality was % in cases with airway bleeding versus . % in those without (p<. ). cox multivariate analysis indicated that the only factors associated with fatal outcome were leukopenia, airway hemorrhage and age (table ). in this article, we compared the characteristics of a group of cases of severe leukopenia (≤ , leukocytes/ml) with a group of patients without severe leukopenia (> , leukocytes/ml) at h of hospitalization. the group with severe leukopenia was exemplified by a significantly higher rate of female patients (p= . ), prior ili (p< . ) and airway bleeding (p< . ). conversely, a personal history of furuncles or skin abscess (p= . ) and ssti at admission (p= . ) were more frequent in the group without severe leukopenia. the mortality rate was significantly different between groups ( . % vs . %, p< . ). necrotizing pneumonia largely occurred in young people with a median age of years, only slightly higher than reported earlier in smaller series [ , , , , ] . the increasing of age of included patients might be attibuable to a delayed exposure to our study confirms the association of leukocyte count with disease severity. by multivariate logistic regression, age over years was associated with severe leukopenia (aor . , p= . ). the reason of this association was not clear. the increasing of age of patients might be attibuable to a delayed exposure to the causative organism. this epidemiological trend should be challenged by similar surveys in other countries. an association between time and lymphocyte count was actually reported in different populations [ , ] . female gender was associated with severe leukopenia (or . , p= . ) by univariate analysis but not by multivariate analysis. females in general mount a more profound immune response after antigenic challenge, and these differences have mainly been attributed to the immunomodulatory effects of sex hormones, despite the lack of human in vivo data. even though we still found some papers reporting an association of neutropenia with female gender. vaneijk et al. demonstrated, in an in vivo study, that females exert a more proinflammatory pattern of cytokine release compared to males during systemic inflammation after the administration of escherichia coli endotoxin [ ] . this difference is associated with more leukocyte sequestration in females [ ] . sterling et al. reported gender-based differences in host immune responses [ ] . in a study analyzing the risk factors for developing neutropenia after mitomycin c administration, multivariate logistic regression showed that female gender was an independent risk factor for neutropenia and the reasons for this association are unknown [ ] . the significant association with a previous ili (aor . , p= . ) might be related to a particular linkage with influenza virus. possible epithelial damage, due to viral infection, could promote the pathogenicity of pvlproducing strains with increasing affinity for collagen (i and iv) and laminin [ ] . a personal history of furuncles appears to be protective (or . , % ci . - . , p= . ). it is conceivable that patients with such a history had previously been exposed to pvl and had developed a degree of protective immunity [ ] . another hypothesis could explain this association: after bacteria reach the lungs through the bloodstream, the necrotizing effect is less serious. the present investigation has some limitations. first, case reporting to the reference center was unsolicited and may not reflect accurate epidemiology and disease severity. severe and dramatic cases among previouslyhealthy young people are more likely to be reported. a bias of notification would not be excluded. this would be caused by some missing data possibly not declared after due to the dispersion of laboratories dealing with pvl gene detection. moreover the median age was slightly higher than had been previously reported which could be related to the introduction of several novelties in the treatment of severely ill patients since the identification of this pathology. another limitation concerns clinical data collection: they were missing or incomplete in % of cases. analysis included only cases for which microbiological and demographic information were available; cases were excluded from further analysis because they lacked data on leukocyte count. comparisons between the excluded cases and the cases analyzed revealed no significant differences. leukopenia may be simply considered as a reflection of disease severity. this is in agreement with in vitro data showing that pvl induces both apoptosis and necrosis in human leukocytes [ ] . the time period between the onset of symptoms and hospitalization could be important to reach a certain level of severity and impaired leukocyte count, but it was not different between both groups. on the other hand, multivariate logistic regression was adjusted to the time factor. furthermore we noticed that factors associated with severe leukopenia were quite different from those associated with mortality. based on our results, we can suggest two syndromes of necrotizing pneumonia: ) those that are mainly related to ili and direct inoculation of staphylococci to damaged respiratory epithelium, airway hemorrhage, severe leukopenia and death; and ) those that are mainly related to hematogenous spread from ssti, less airway hemorrhage, without leukopenia and improved survival. in conclusion, our study emphasizes that s. aureus-necrotizing pneumonia is still an extremely severe disease. we found that some factors could distinguish patients with severe leukopenia from those without leukopenia. leukocytes count is promptly available at admission to the hospital and can be easily used to asses the severity of disease as suggested by this paper. the impact of this marker, on patient management, need to be clarified. empiric therapy should include coverage for s. aureus as soon as possible, without waiting for the bacteriological results. clinical data indicate that neutralizing toxin production improves the outcome [ , ] . the toxin can be blocked by combining a toxin-suppressing agent (e.g., clindamycin, linezolid or rifampin) with bactericidal antibiotics acting on the cell wall [ ] . s. aureus-necrotizing pneumonia is still an extremely severe disease in patients with severe leukopenia. some factors like leukocytes count could distinguish these patients, allowing better initial identification to initiate adapted, rapid administration of appropriate therapy. this paper could be regarded as a preliminary work. variables were adjusted to gender, mechanical ventilation, personal history of furuncles, platelet count, pleural effusion, and previous influenza-like illness. experts are invited to work on a widely accepted score, validate a score including leukocytes count, to predict the severity of this disease. association between staphylococcus aureus strains carrying gene for panton-valentine leukocidin and highly lethal necrotising pneumonia in young immunocompetent patients life-threatening hemoptysis in adults with community-acquired pneumonia due to panton-valentine leukocidinsecreting staphylococcus aureus severe community-onset pneumonia in healthy adults caused by methicillin-resistant staphylococcus aureus carrying the panton-valentine leukocidin genes severe community-acquired pneumonia due to staphylococcus aureus, - influenza season severe community-acquired pneumonia 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i and iv collagens and laminin a history of panton-valentine leukocidin (pvl)-associated infection protects against death in pvl-associated pneumonia staphylococcus aureus panton-valentine leukocidin directly targets mitochondria and induces bax-independent apoptosis of human neutrophils expanded clinical presentation of community-acquired methicillin-resistant staphylococcus aureus pneumonia prompt and successful toxin-targeting treatment of three patients with necrotizing pneumonia due to staphylococcus aureus strains carrying the panton-valentine leukocidin genes submit your next manuscript to biomed central and take full advantage of: • convenient online submission • thorough peer review • no space constraints or color figure charges • immediate publication on acceptance • inclusion in pubmed, cas, scopus and google scholar • research which is freely available for redistribution we thank the clinicians and microbiologists who sent us clinical data and isolates. we acknowledge the contribution of prof. rené ecochard and dr. muriel rabilloud from the department of biostatistics in lyon university hospital. special thanks to mr ovid da silva for editing this manuscript. the authors declare that they have no competing interests.authors' contributions je, pv, fv, yg, od, vm, at, mb and gl conceived of and designed the study; nk, ns, and pv performed the data analysis; and nk, ns, and je wrote the paper. all authors read and approved the final manuscript. key: cord- - ri x y authors: zhou, bo-ping; lu, pu-xuan; chen, qiu; xiao-ping, tang; yu-juan, guan; jin-xin, liu; xing, lu; zhen-wei, lang; xin-chun, chen title: sars date: - - journal: diagnostic imaging of emerging infectious diseases doi: . / - - - - _ sha: doc_id: cord_uid: ri x y severe acute respiratory syndrome (sars) is an acute respiratory tract infectious disease induced by sars-cov and mainly transmitted through the short-distance air droplets and close contact. its main clinical characteristics is abrupt onset of the disease and the initial symptom is fever accompanied with systematic symptoms of headache, soreness and fatigue, and respiratory tract symptoms such as cough, chest dullness, and dyspnea. a few cases may progress to acute respiratory distress syndrome (ards). due to its self-limiting feature, the prognosis is predominantly good but may be poor in severe cases, with mortality about . %. some patients may develop such complications such as lung fibrosis and necrosis of the head of femur. on april , , sars was defined as a legal infectious disease by the ministry of heath of china. . the fi rst sars case was reported around the world on january , ; a hospital in heyuan city of guangdong province hospitalized two patients' with severe pulmonary infection of unknown cause, which was the fi rst traceable report of "sars" around the global. . on february , , sars was fi rst diagnosed by the who expert carlor urbani in hanoi of vietnam, who also named this new unexplained disease " s evere a cute r espiratory s yndrome" (sars). following the emergence of sars, who and governments around the world attached great importance and took consecutively effective control and prevention measures, which reined in the infection rapidly controlled in a short time. on april , , who eliminated vietnam from the name list of the sars-infected area, which made vietnam the fi rst to get rid of sars outbreaks. since then, numerous countries were also ticked off the list. on july , , who announced to remove taiwan of china from sars-affected area list, which is also the last. at present, there is no sars-affected area around the world, signaling the victory of global battle against sars. by the end of september , , there were in total , people infected and deceased, with the mortality at . %. on april , , a new coronavirus never seen was announced as the cause of sars by who. it is the joint efforts of global scientists and biological scientifi c and technological advances that enabled establishment of international sars research network, separation and identifi cation of sars coronavirus, and confi rmation of sars pathogenesis. some canadian laboratories excluded infl uenza virus a and b; para-infl uenza virus type , , and ; adenovirus; and respiratory syncytial virus by scanning electron microscopy and direct fl uorescent antibody test. they also conducted immunohistochemistry on the corpse tissues of dead patients about viruses including infl uenza virus a and b, respiratory syncytial virus, adenovirus, circovirus, hantaan virus, measles virus, intestinal tract virus, and pneumonia mycoplasma and chlamydia, which showed negative. scholars from german, france, america, hong kong, and taiwan applied specifi c pcr tests targeted at corresponding pathogens to examine pneumonia mycoplasma, pneumonia chlamydia, human cytomegalovirus, circovirus, herpes virus, human coronavirus oc and e and arenaviruses, bunyavirus, hantaan virus, crimean-congo hemorrhagic fever virus, which also indicated negative. on march , , hong kong university fi rstly separated and cultured coronavirus from the nasopharyngeal specimen of sars patients by vero cells, and then several laboratories of canada and america disease center and sars international coordination group also cultured coronavirus. on april , , based on the aforementioned research fi ndings, who declared a new coronavirus as the pathogen of sars and named it as sars coronavirus (sars-cov). sars-cov is a single-stranded and positive rna virus belonging to the genus coronavirus of coronaviridae family of nidovirales order. it bears great morphological resemblance with known human coronaviruses. under the electric microscopes, sars coronavirus presents with pleomorphic spherical particles with envelopes in vero e cell in vitro and with a diameter of about - mm. it has distinct coronavirus morphological characteristics that rodlike surface projections about - nm long protrude outside viral envelope to form a corona. for separation and culture of viruses in vero e cells, viral particles are mostly found in vacuoles inside cells under electric microscope, mainly hollow particle, and viruses outside cells observed with spikes. the coronavirus is mainly composed of nucleic acid, protein, carbonate, and lipid. sars-cov are relatively stable in human specimens and environment but relatively sensitive to chemical and physical factors. the viruses could be killed by exposure to ultraviolet rays for min. as the envelope of sars-cov contains lipoids, they are sensitive to lipid solvents and could be inactivated by ether, chloroform, tween, % ethyl alcohol, methanol, pancreatic enzymes, and ultraviolet rays. ether could sterilize the virus completely under °cafter h, % ethyl alcohol could deactivate virus after min, and disinfectants containing chlorine may kill the virus aft. sars is the fi rst newly emerging infectious disease in the twenty-fi rst century. since the fi rst case was detected in guangdong on january , , sars spread to countries at fi ve states at an unprecedented speed with several months. according to the report of who, sars epidemics was found in total the main source of infection is sars patients, and other infectious sources such as animals need to be further consolidated. it is regarded that only symptomatic patients can effectively spread sars-cov. at present there still lacks clinical proof that asymptomatic sars patients could spread sars coronavirus. few cases are highly contagious and able to directly infect over patients, who are called super-spreader. the fi ve super-spreaders in singapore have been found infecting people. such powerful infectiousness is attributable to the high viral load inside patients who could discharge abundant viruses via respiratory tract in a short time. specialists from the department of microbiology of hong kong university and guangdong provincial disease control and prevention center tested abundant sars coronaviruses from the samples collected from civet cats sold in guangzhou and shenzhen, suggesting that the human sars coronavirus might originate from civet cats. sars virus is spread mainly through the respiratory tract and also spread by aerosol and droplet. high level of sars coronavirus could be detected in the throat swabs and sputum specimens of the patients in acute phase. viruses discharged by patients via cough would exist in the air within certain radius, thus forming short-distance respiratory tract droplet spread. it is a main route of sars dissemination. indirect or direct contact spread could also spread sars coronavirus. it could be spread through contact with the respiratory tract secretion or through the mouth and nose via contaminated hands and toys. as viruses proliferate abundantly in the respiratory tract, exfoliated cells containing infectious viruses could be expelled outside the body through patients' respiration, cough, and sneeze and suspend in the air, thus forming the aerosol of sars virus. cough and sneeze could greatly contribute to production of sars virus aerosol. sars is a new type of infectious diseases, and people effectively exposed to sars pathogens are vulnerable. present data reveal that sars is an acute self-limiting infectious disease, and neutralizing antibodies to sars could be detected weeks after onset. however, the change patterns of neutralizing antibody titer have not been fully elucidated. the epidemics of sars mainly take place in the winter and spring. sars outbreaks are mainly concentrated at coastal tourism cities with prosperous economy, dense population, and convenient transport, particularly air transport. from the perspective of international geography, sars primarily occurs in southeastern countries or regions, including china mainland, hong kong, taiwan, singapore, and vietnam. analysis of the statistics of various countries shows that sars onset is mainly recorded in young populations aged - years old, occupying around % of the total. sars coronavirus (sars-cov), a mutant of coronaviridae, is pathogenic to hosts in the following two aspects: fi rstly, viral infections could directly damage the structure and functions of infected cells and trigger cell apoptosis; secondly, immune reactions and release of various cytokines induced by viral infections could not only eliminate infectious pathogens but also may deal severe damages on histiocytes due to excessive immune reactions. the member of coronaviridae is a single-chain positive rna virus, which taxonomically belongs to nidovirales with arteriviridae. the homological analysis of sars-cov structural protein amino-acid sequences shows that the main structural protein s (spike protein), m (membrane protein), n (nucleocapsid protein), and e (envelope protein) bear very low homology with other known coronaviruses, between and %. besides, bioinformation analysis infers that sars-cov may also encode - nonstructural proteins of unknown functions. all these sars-cov specifi c proteins may play key roles in determining the viral virulence. s-protein, existent on the viral surface in form of trimer, is the main component in constructing the coronal structure and major structural protein for the integration of virus and host cell receptors and fusion of viral envelope and cell membrane to enable invasion of viruses. after infection with sensitive cells, sars-cov could replicate enormously and release virions, which may disturb cell metabolism and directly result in injury of body tissues and cells. in the early phase of the disease and in the continual progression phase, the active immune cells such as cd + and cd + t lymphocytes, nk cells, and dc cells are decreased prominently, to an extent positively correlated with severity of conditions and speed of progression. besides, all dead patients are detected with irreversible signifi cant decline. however, along with remission of conditions and entry into recovery phase, the number of aforementioned immune cells could be recovered to some extent, whose change presents distinct par-allel correlation with disease evolution. dynamic observations suggest that serum pre-infl ammatory cytokine il- , il- , il- , and inf-α level of sars patients could reach the peak at about second week of disease course, which relatively corresponds to the acute aggravation phase. all these consolidate the immune damage of sars-cov infections. autopsies of the over patients who died after infected with sars revealed that sars is a disease detrimental to organs of the whole body, particularly lung tissue and immunity system. primary pathologic changes can be classifi ed into severe lung lesions, immune organ impairment, toxic changes of other organs, and secondary infections. the causes of death were as follows: ( ) diffusive lung alveolar damages lead to progressive respiratory failure; ( ) impairment of liver, kidney, heart, and several other organs may induce multiple organ failures and exacerbate conditions; ( ) due to injury of immune organs particularly lymph nodes and spleen, the lymphocytes are decreased and immunity compromised, therefore resulting in secondary mycotic infections. lung tissues become swollen and heavier. according to domestic report, the left lung may reach - g (average . g) and right lung - g (average . g), with the total lung up to - g (average . g). the lung tissues appear in dark red. texture is hard, indicative of consolidation. the surface is relatively smooth and free of pleural adhesions. lung tissues are observed with blood vessel dilation and congestion, spotted and patchy necrosis, and hemorrhagic infarct foci and focal compensatory emphysema. sections of various lung lobes are visualized with outfl ows of light red or/and a few foamy blood-stained fl uids. the trachea and bronchi are found with exudations of slight mucoid or blood-stained secretions. pulmonary hilar lymph nodes are revealed with slight enlargement and pleural cavity without or with a few effusions. microscopy reveals bilateral diffusive severe injuries of alveoli, characterized by acute exudative, hemorrhagic, and fi brinous infl ammation. however, the injury may be inhomogeneous and diverse at different lung lobes and different parts of the same lobe. the characteristic change in early phase is pulmonary edema, with homogeneous pink serous or fi brinous effusions fi lling alveolar cavities as well as erythrocytes leakages in some cavities. the effusions from some alveolar cavities may condense to form thin-layer membranelike substance which adheres to alveolar walls, namely, formation of hyaline membrane ( fig. . ). alveolar epithelia present with such diffusive injuries as degeneration and necrosis, with exfoliated or/and apoptotic type ii alveolar epithelial cells observable in alveolar cavities, which are similar to changes of desquamative pneumonia ( fig. . ). among them, some appear like apoptotic bodies. partial regional type ii alveolar epithelial cells and macrophages proliferate actively, with obvious increase of cell volume and distinct nucleolus. in some case, cells are detected fusing with each other to form syncytial monocytes and multinucleated giant cells (fig. . ) . the alveolar epithelia of some cases are examined with structures similar to viral inclusion bodies, mostly spherical and with eosinophilic staining, around which a transparent halo is visible (fig. . ) . pulmonary interstitium and alveolar spaces are detected with high dilation and congestion of capillaries, swelling and exfoliation of epithelial cells, as well as widening of the latter, which present slight lymphocyte and monocyte infi ltration, small vessel proliferation, and enlargement characteristic of vasculitis changes. some regions are observed with pulmonary vascular embolism or fi brinous microthrombi in alveolar capillaries, while some alveolar capillary cavities manifest dilation but free of blood components. in addition, some patients are detected with lung tissue focal hemorrhage, compensatory emphysema, and small airway necrotizing infl ammation and alveolar collapse or shrinkage. patients with a disease course of over three weeks manifest fi broblast proliferation in alveolar septa, and individual cases demonstrate organization of fi brinous exudates from alveoli and fi broblast proliferation, which gives rise to glomeruloid structures, named "glomeruloid organizing pneumonia" (fig. . ). the bronchial epithelial cells are revealed with exfoliation, scarce and exfoliated cilia, as well as structures similar to viral inclusions and squamous cell metaplasia at the inside. the bronchioli are detected with submucosal edema, infl ammatory cell infi ltration, and serous grandular hyperplasia with hyperactive secretion. pulmonary hilar lymph nodes demonstrate light swelling and present different degrees of congestion, hemorrhagic necrosis, and decrease of innate lymphocytes under the microscope. blood sinus in the lymph nodes presents with grave congestion and dilatation, lymphoid nodule atrophy or disappearance, prominently the cortex, and lymphatic tissue focal necrosis, accompanied by apoptosis of some lymphocytes ( fig. . ). the spleen mostly shrinks with soft texture, and has patchy and focal hemorrhage at the surface and sections. splenic white pulp is detected obviously atrophic or absent, with central arterial wall thickened, endothelial cells swollen and partially shed, peripheral lymphatic sheath lymphocyte depleted greatly, and germinal center disappeared. splenic corpuscle is observed with plasma protein sedimentation in central arterial walls, white pulp and marginal sinus lymph tissues with large patchy necrosis, and partial remaining lymphocytes with apoptosis. the splenic sinus inside red pulp is revealed with obvious congestion and focal hemorrhagic necrosis as well as proliferation of histiocytes ( fig. . ). the number of hematopoietic cells in bone marrow declines, and granulocyte system and megakaryocytic system are relatively inhibited. polychromatic erythroblasts present with small focal proliferation. tissues of the brain, liver, kidney, and heart demonstrate varying degrees of congestion, edema, degeneration, and necrosis as well as infi ltration of a few lymphocytes. secondary bacterial pneumonia and fungal infection may occur ( fig. . ). the initial symptom of sars is frequently fever, accompanied by headache and general muscular and joint souring soreness, and often without nasal discharge, sore throat, and other upper respiratory tract catarrhal symptoms. pulmonary signs are not obvious and signs and distinct systematic symptoms are inconsistent, which is typical to sars. the clinical manifestations of sars may vary, which can be divided into light type, common type, severe type, and atypical type, with the common type most commonly seen. based on the clinical development, it could be divided into early phase, progression phase, and recovery phase. generally, the incubation phase lasts - days, with an average of - days and a median of days, predominantly - days. the initial symptom primarily is fever, mostly high fever, accounting for . - %. patient body temperature may reach - °c and °c at the highest. the fever type varies, including remittent fever, irregular fever and continuous fever. the duration of fever is mostly - days. it presents as headache, joint or/and systematic soreness, and chest pain. it is reported that the incidence of headache in sars patients is - %. most sars patients have cough, which is dry with scanty sputum. generally sars does not incur upper respiratory tract catarrhal symptoms. severe cases may suffer from accelerated respiration, shortness of breath, or even acute respiratory distress syndrome, characterized by progressive dyspnea or even distress with respiratory frequency up to > times. patients complain of heart beat or palpitation accompanied often by precordial discomfort. the incidence is - %. the incidence of diarrhea is reported mostly at - %, with single case up to %. one characteristic of sars is severe symptoms and light signs. the signs are mostly not obvious or even absent. dyspnea presents irregular respiratory frequency, depth, and rhythm. the incidence of dyspnea of sars patients is - %. severe patients may manifest nose fanning, orthopnea, and cyanosis, involvement of accessory respiratory muscles in respiration, as well as "three depression signs" upon inhalation. according to report, % of typical patients and % of severe patients have sinus tachycardia, and . % of sars patients suffer from paroxysmal supraventricular tachycardia. the pulmonary signs of sars patients are frequently unobvious or absent in the early phase. light moist rales may be audible ( - %) . patients may also demonstrate pulmonary consolidation sign and attenuated breath sounds. there may also be a few pleural effusions. in light of conditions, it could be classifi ed in clinics into light type, common (typical) type, severe type, and special (atypical) type. the follow-up visit of sars patients via x-ray and hrct by choi et al. found that about - % patients developed fi brotic changes of lung interstitium and shrinkage of lung volume. application of glucocorticoids could to some extent alleviate lung injury of patients with severe infectious atypi-cal pneumonia. however, long-term and abundant use of glucocorticoids may generate numerous side effects, and it is an undisputable fact that hormones may induce femoral head ischemic necrosis. literature reports suggest that the incidence of femoral head ischemic necrosis is - %. the white blood cell count remains within normal range in majority patients and may decrease in some patients ( . %). % of severe patients without secondary infections are revealed with decreased total white blood cell, and normal neutrocyte and monocyte differential white blood cell count. however, % initially diagnosed patients manifest a decrease of absolute lymphocyte count, suggestive of a tendency of gradual decline, and morphocytology change, with abnormal lymphocytes observable. . - % patients are examined with thrombocytopenia. many evidences prove that sars-cov could directly invade the immune system and act mainly on lymphocytic system, especially the t lymphocytes, which could lower peripheral blood lymphocytes. the peripheral blood white cell count is normal or decreased in most sars patients, but cd +, cd +, and cd + t lymphocyte counts are distinctly lower compared with those of healthy population. the more severe the condition, the more drastically declined the t lymphocyte count. blood gas analysis reveals hypoxemia of different degrees and commonly no carbon dioxide retention. generally under - l/min oxygen uptake, patients with severe hypoxemia are measured of arterial partial pressure from oxygen (pao ) < mmhg or blood oxygen saturation (spo ) < %, or develop acute lung injury (ali) or acute respiratory distress syndrome (ards). one of the main immunopathogenesis changes in sars patients is insuffi ciency of t-lymphocyte-mediated specifi c cellular immunity, characterized by distinct damages of t lymphocytes and its subgroups, particularly cd + , cd + , and cd + . there exists distinct correlation between the extent of t lymphocyte damages and severity of the disease. the method recommended by who for test of serum sars-cov antibodies is enzyme-linked immunosorbent assay (elisa). upon test of the sars-cov-specifi c antibodies in the infected patients, including igg, igm, or total antibody, seroconversion or increase by times and above of any serum antibody in progress phase and recovery phase could serve as evidence for defi nite diagnosis. polymerase chain reaction (pcr), a type of molecular biological test, could identify sars virus genes from various specimens (blood, feces, respiratory tract secretions, tissue sections, etc.), that is, sars virus rna. presence of any of the following three circumstances could be sars virus positive : . clinical specimens from at least two different sites are tested positive (nasopharyngeal secretions and feces). . the same clinical specimens collected by at least days apart are tested positive (two or more specimens of nasopharyngeal excretions). . the same clinical specimen is tested positive by two different methods or in different laboratories. . repeated pcr examination indicated positive. clinical diagnosis . the patient has a history of intimate contact with patients or is one of the infected populations or consolidated having infected others by defi nite evidences. . the patient had been to or lived in the areas with sars patients and secondary infection epidemics. the onset is acute and initial symptom is fever, with body temperature > °c, and occasionally accompanied by concurrent chillness. patients may also suffer from headache, joint and muscular soreness, fatigue, and diarrhea, and generally there is no upper respiratory tract catarrhal symptom. patients may have cough, mostly dry with scanty sputum, and chest dullness. severe patients demonstrate accelerated respiration, shortness of breath, or obvious respiratory distress. pulmonary signs are not obvious, and light moist rales may be audible or pulmonary consolidation observable. there is generally no increase of peripheral white blood cell count or decrease. besides, the lymphocyte count is found generally declined. lungs present different degrees of patchy and plaque-shaped infi ltration shadows or reticulate changes, which may progress rapidly into large patchy shadows. these commonly occur at multiple lobes or both sides, and absorption and dissipation of shadows are relatively slow. pulmonary shadows and symptoms and signs may be inconsistent. if the test indicates negative, reexaminations shall be conducted after - days. there are no remarkable responses to antibiotics. patients in compliance with any of the following three circumstances are diagnosed as suspected cases. . the patient has a history of intimate contact with patients, or is one of the infected population, or is confi rmed having infecting other people by concrete evidences, who also manifest aforementioned clinical symptoms but not high white blood cell count by test of peripheral blood. . the patient has been to or lived in areas reported with infectious sars patients and patients suffering from secondary infections weeks before the disease onset, who also have abovementioned clinical symptoms and consistent pulmonary shadows on chest x-ray fi lms with the abovementioned characteristics. . despite the lack of epidemic data, the patient has abovementioned clinical manifestations and not high peripheral white blood cell count, as well as pulmonary shadows on chest x-ray that accord with abovementioned characteristics. in other words, the patient conform to the aforementioned ( ) + + or ( ) + + or + + . in case of any of the three following circumstances, the patient could be clinically diagnosed with sars. . the patient has a history of intimate contact with patients, or is one of the infected population, or is confi rmed having infecting other people by concrete evidences, who also manifest aforementioned clinical symptoms and pulmonary shadows on chest x-ray image. . the patient has been to or lived in areas reported with infectious sars patients and patients suffering from secondary infections weeks before the disease onset, who also have abovementioned clinical symptoms, not high peripheral blood white cell count and pulmonary shadows on chest x-ray fi lms. . the patient has been to or lived in areas reported with infectious sars patients and patients suffering from secondary infections weeks before the disease onset, who also have abovementioned clinical symptoms, pulmonary shadows on chest x-ray fi lms, and no obvious response to anti-infectious treatment. in other words, the patient conform to aforementioned ( ) + + or ( ) + + + or ( ) + + + . the conditions are mild without pulmonary fi brosis and other sequela. the prognosis is good. patients have typical clinical manifestations of the early phase, progression phase, and recovery phase. the onset is acute, and the fi rst symptom is fever, with body temperature generally > °c. besides, the patients have no obvious respiratory distress or hypoxemia. some patients are auscultated with slight moist rales or observed with pulmonary consolidation. the prognosis is good without sequelae. severe type accounts for about %. it is same with the common type in clinical process but presents severe conditions and rapid progression. some cases may progress to acute pulmonary injury or ards, rapidly incurring respiratory failure or even death. the diagnosis could be reached in case of any of the following three circumstances: . dyspnea adult respiratory frequency is ≥ times /min, accompanied by one of the following conditions: the chest imaging shows that the total surface of multilobar lesions or foci occupies more than / of the two lungs on orthotopic imaging. conditions progress, with the focal area increased by over % within h or accounting for over / of the two lungs on orthotopic imaging. . obvious hypoxemia is detected, with the oxygenation index lower than mmhg. . shock or multi-organic dysfunction syndrome occurs. there are still some atypical cases of this disease. this generally refers to the fi rst - days. the onset is acute and initial symptom is fever. half of the patients are accompanied by the nonspecifi c systemic symptoms such as chillness, headache, and joint and muscular soreness. this phase spans the day - of the disease course. infectious toxic symptoms such as fever, fatigue, muscular soreness continue to exist or even aggravate. patient's body temperature rises, possibly to °c or higher in a short time, and the high fever is persistent. lung lesions demonstrate progressive exacerbation. a few patients ( - %) may develop ards, which is life threatening. it mostly takes place at day - of disease course. most patients have good prognosis after recovery for about weeks. some severe patients could gradually recover within - months after discharge. these mainly encompass standing chest x-ray and bedside chest x-ray, which are economic and effective examination methods and fi rst optional examination for preliminary diagnosis and reexamination. mobile x-ray bedside imaging should be applied for patients under observation and inpatients. as lung disease of sars patients progress rapidly, chest x-ray reexamination within a short period could help observe changes of conditions, commonly once every - days at the early phase and progression phase. regular chest x-ray also need to be conducted during recovery phase so as to ascertain absorption and dissipation of lesions, residual foci, and pulmonary interstitial fi brosis. for sars patients, ct scan generally is not a preferred method and, if necessary, could be conducted under strict disinfected quarantine. if the foci could not be absorbed for a long term in sars recovery phase or patients still have symptoms but normal chest presentations, ct scan need to be carried out for further observation as it can better visualize the subtle pulmonary interstitial changes, such as lung interlobular septum thickening, intralobular septum thickening, subpleural linear shadow, and small ground-glassdensity lesion and regional and segmental bronchiectasis, and therefore is helpful for clinical diagnosis of pulmonary interstitial fi brosis. thin-layer ct scan or hrct scan can better visualize intrapulmonary low-density small lesions, thus contributory to early diagnosis of pulmonary interstitial fi brosis. ct examination could also aid identifi cation of other concurrent lung lesions. medical staff in the room of imaging examination and department of radiology shall prevent infections in strict compliance with sars disinfection and prevention requirements and meanwhile conscientiously implement x-ray protection. . ward shall be equipped with bedside x-ray machine; imaging for all sars inpatients shall be conducted in wards and patients shall wear masks. . the personal protection for imaging technician should be subject to the level-two protection requirements and, when necessary (imaging for critically ill patients), to level-three protection requirements. . x-ray fi lm magazine should be put into the isolation bag before imaging (one isolation bag for each person). . the imaging technicians should sterilize their hands before return to the radiological department and then remove protective clothing, gloves, and protective glasses at the designated place. they should wash hands thoroughly, gargle, take a shower, and change clean working clothes before work. . the department must strengthen interior ventilation and air disinfection . ultraviolet irradiation may be assumed at night, - times per day, minutes per time, and peracetic acid spray conducted times per day. . rigorous isolation measures: all subjects must wear masks. the operation staff must wear protective masks, gloves, and gowns, with masks changed every h and gowns replaced in time, and should use disposal bed sheet and shoe covers, and dispose of and replace them timely after the examination. . sars examination zone should be separated from conventional patient examination zone and examination staff shall be fi xed. if feasible, special chest x-ray examination room and x-ray machines may be set up in the isolation zone to avoid cross-infections. . the protective measures for subjects and operation staff are similar to those required about isolation protection in radiological department. . all the subjects must wear masks and, for examination in ct room, shall minimize the stay time as much as possible. . after the patient leaves, the environment and articles which may be polluted by patients shall be given terminal disinfection. . the department must enhance interior ventilation and air disinfection and, in addition to use of chemical disinfectant spray, may also initiate long-term ultraviolet irradiation at night for disinfection. . a pathway specifi c for sars patients should be established. chest x-ray diagnosis the imaging manifestations of sars are closely related to its pathological changes. besides edema, infl ammatory cell infi ltration and other nonspecifi c infl ammatory changes, the more prominent pulmonary pathological feature of sars is abundant exfoliation of epithelial cells, inducing obvious thickening and damage of alveolar septum, as well as significant organization of effusions inside alveolar cavities. all the three changes of infl ammation in early phase (degeneration, exudation, hyperplasia) are observable. the mechanism of acute lung injury induced by sars may be ascribed to sars coronavirus direct attack of alveolar epithelia and alveolar capillary epithelia or (and) indirect damage via lymphocyte, macrophage and effector cells, and lymphokine, cytokine, and infl ammatory mediator released by them. this suggests that besides viral direct infection, immune reactions are also involved in lung injury. in severe lung injury, pulmonary interstitial and pulmonary parenchymal air cavity changes detected by autopsy pathology, especially pulmonary air cavity not completely fi lled by lesion or fi lled but coexistent with unfi lled alveoli, are visualized as ground-glass-density shadows on x-ray and ct. varying degrees of residual lung cavity present with different densities of ground-glass shadows on thin-layer ct or high-resolution ct (hrct). complete fi lling of the alveolar space leads to pulmonary consolidation shadows. once pulmonary edema and alveolar hyaline membrane are formed, patients clinically manifest decreased lung compliance, progressive dyspnea, and intractable hypoxemia. it is such rapid pathological changes of sars that give rise to characteristic presentations by chest x-ray examination, including rapid progression of lesions, diverse morphological states, and wide range, often spanning several segments and lobes. therefore, dynamic observation of chest image holds great signifi cance for diagnosis of sars. ground-glass-density shadows are pathologically the result of lesions mainly at pulmonary interstitium and alveoli, with pulmonary interstitial lesions frequently complicated by alveolitis. ground-glass-density shadows incurred by alveolar consolidation are attributable to partial fi lling of alveoli or coexistence of fi lling and unfi lled alveoli. such shadows on x-ray and ct can be determined if the density of lesions is lower than that of blood vessels, and blood vessel presentations are observable inside. the density of lesions lower than that of pulmonary hila on x-ray could also help identify ground-glass-density shadows. pulmonary consolidation shadows are pathologically due to fi lling of alveoli by pathological tissues, which is frequently complicated with lung interstitial lesions. pulmonary consolidation could be confi rmed if the lesions are found with higher density than vascular shadows and without vascular shadows inside but with air bronchogram on x-ray and ct. it could also be determined if the lesions present higher density than pulmonary hila or similar density to mediastinum. the lesions are solitary or multiple in small patches, and some lesions are distributed along lung lobes and segments. % of sars patients may develop subsequently solitary and multiple small patchy shadows within seven days of onset, with relatively low density, blurry borders, and irregular shapes, predominantly solitary. occasionally, the pulmonary textures around lesions are revealed increased and thickened and mainly distributed at peripheral area, more commonly at bilateral inferior lungs. since chest x-ray can poorly visualize relatively small lesions and at sometimes posteroanterior x-ray can hardly display lesions overlapping with heart shadows, orthotopic chest imaging should be conducted at the same time ( fig. . ). the conditions of most patients become worse within days after onset. the small patchy shadows in the early phase may progress into large patchy, multiple, or diffusive lesions. the lesions may spread from unilateral lung to both lungs and from one lung fi eld to several fi elds. severe patients may demonstrate obvious changes - days after onset. lesions present mainly as ground-glass-density shadows or coexist with lung consolidation shadows. the center of some shadows with high density indicates consolidation, and relatively low-density peripheral area suggests ground-glass fig. . the -year-old male patient diagnosed with sars. the specifi c segment rt-pcr of srs virus is positive. on day of onset, the right inferior lung fi eld was observed with patchy ill-demarcated shadows of increased density density which may be complicated with thickening and increase of pulmonary textures. severe patients may develop "white lung." some cases may have pneumothorax, pneumomediastinum, and subcutaneous emphysema after the use of ventilator. a small part of lesions adjacent to the pleura may be complicated with local pleural thickening or revealed with mild tentiform adhesion. pleural changes may reside after absorption of intrapulmonary lesions. complication of slight pleural effusions may also be observable (fig. . ). the lesions of sars patients begin to be absorbed in - days, which can be completely absorbed for majority patients. a few patients may show pulmonary fi brosis or pulmonary interstitial hyperplasia, with apparent pulmonary interstitial hyperplasia occurring - days after the onset. the intrapulmonary patchy shadows present with shrinkage and increase of density to gradually form high-density cordlike and honeycomb-shaped shadows in lungs. severe pulmonary interstitial proliferation could dwindle the lung volume and make the mediastinum shift to the affected side. the imaging may reveal local irregular high-density plaque and cord-shaped shadows. occurrence of intrapulmonary honeycomb-shaped shadows and tractional bronchiectasis is the characteristic of pulmonary interstitial fi brosis (fig. . ). as rapid change is an important feature of sars chest x-ray manifestations, dynamic x-ray studies and observations could provide evidences for sars defi nite diagnosis. therefore, observation of the disease dynamic change is critical to sars chest x-ray, which is a major difference from x-ray examinations of other pneumonia. in the early phase and progression phase of sars, lung foci may undergo remarkable changes within a short time ( h at the shortest), such as expansion and dissemination, which are characterized by changes in shape, extent, and site of the foci. this suggests that clinical physicians shall conduct chest orthotopic imaging once every - h after the sars patients are hospitalized so as to ascertain the disease changes. imaging dynamic changes are associated with multiple factors, such as age, original underlying disease, treatment method, and effi cacy. the lesions are generally absorbed days after onset but possibly on day in some mild patients. the focal shadows may become smaller with the density gradually decreased. one characteristic dynamic change of sars is migrating change of lesions in some patients, suggesting that the foci fi rstly occur at the unilateral inferior lung fi eld and then spread to contralateral and/or superior and middle lung fi eld. patients with worse lesions may manifest wider imaging extent and new foci, and severe patients may develop "white lung." ards is a key cause leading to death of patients (fig. . ) . lung imaging examination is an important basis for sars diagnosis, and continual imaging examination could visualize the dynamic change characteristics of the disease. after sars infection, chest abnormal changes frequently occur on day - . in initial phase, focal shadows are frequently seen, which might be unilateral but dominantly bilateral. ct reveals small patchy ground-glass-density shadows, some of which are quasi-circular. lesions present as solitary small patchy lung consolidation and multiple small patchy or relatively large patchy shadows. large foci may appear as ground-glass-density shadows and concurrent lung consolidation shadows, with relatively higher-density vascular shadows observable in the former. besides, some foci are found with increased vascular shadows at the periphery. in addition, lesions are unveiled frequently at the bilateral lung inferior fi elds and lung margins (fig. . ). lesions in majority patients may progress and worsen within days after onset. the small patchy shadows in early phase could expand into large patchy, multiple, or diffusive lesions within - days. besides, lesions extend from unilateral lung to bilateral lungs, from one lung fi eld to multiple lung fi elds. most patients could develop the severest infi ltration of lungs on day - , namely, peak phase or "critical" phase. ct presentations are still predominated by ground-glass-density shadows, which may be complicated by pulmonary consolidation. lesions are commonly multiple and in diffusive distribution at both lungs, and lesions of varying shapes may coexist, with quasi-circular foci relatively common. some cases appear with ground-glass-density shadows all the time from onset to absorption of lesions. when ground-glass-density shadows are complicated by pulmonary consolidation, the large patchy, small patchy, or quasi-circular ground-glass-density shadows could be observed with pulmonary consolidation shadows of relatively high density. ground-glass density and pulmonary consolidation may also occur at different parts of the lung and could be visualized on the same layer or different layers of ct. for lesions typical of pulmonary consolidation, pulmonary consolidation appears as plaque-shaped high-density shadows or consolidation signs at lung lobes and segments. in addition, lesions are frequently recorded at lung lobe segment of the inferior lobe and outer band of lung fi eld (fig. . ) . lesions usually begin to be absorbed - weeks after onset, with the shadow shrunk and density gradually reduced and absorbed. although the clinical symptoms of some patients improve and disappear, light ground-glass-density shadows could still be seen in lungs on ct. it may prolong a relatively long time. during the absorption of intrapulmonary lesions, there is also concurrent pulmonary interstitial proliferation, which begins to be absorbed under dynamic observation. some lesions may develop into pulmonary interstitial fi brosis, characterized by lobular septum, intralobular septum, interstitial thickening, and subpleural curvature imaging signs as well as regional irregular high-density plaque and cord-like presentations. occurrence of intrapulmonary honeycomb manifestations and tractional bronchiectasis signifi es pulmonary interstitial fi brosis ( fig. . ). sars complication is an important factor affecting prognosis, which should be given enormous attentions. during sars treatment, antiviral drugs are administered as well as glucocorticoids and broad-spectrum antibiotics. some patients may still need airway positive pressure ventilation, tracheotomy, and tracheal intubation. reasonable use of such measures is of great signifi cance for improving the treatment rate and lowering mortality of sars patients. however, to be noted, such treatment drugs and methods may incur adverse reactions and cause complications. sars complications already detected encompass pulmonary secondary infection, mediastinal and subcutaneous emphysema, pneumothorax, bone aseptic necrosis, empyema, brain secondary infection, and pulmonary interstitial interstitial fibrosis in the early phase, sars manifests bronchiolar and peripheral interstitial pneumonia, which could further progress into patchy ground-glass blurry shadows at bilateral superior, middle and inferior lung fi elds, and consolidation and air bronchogram at different layers alveolar consolidation, with interstitial lesions present persistently and mostly reversible. in the recovery phase, only some cases develop into pulmonary interstitial proliferation and then cause fi brotic changes. pulmonary interstitial proliferation and fi brosis degree and outcome are associated with the extent of pulmonary involvement in peak phase, occurrence of complications, and treatment. generally, if lesions on chest image are light in peak phase, there will be fewer pulmonary changes left in recovery phase. contrarily, patients with diffusive lung interstitial thickening in recovery phase are often those suffering from relatively severe pulmonary injury in peak phase and involvement of the great mass of lungs, particularly patients with recurrent conditions, protracted course, and concurrent infections. researches demonstrate that - % of sars patients have pulmonary fi brosis of varying degrees and male patients display relatively more obvious pulmonary impairment than female patients. primary manifestations are lung dysfunction, such as decrease of pulmonary diffusion capacity, lung capacity impairment, and change of total lung capacity particularly residual volume. generally obvious pulmonary interstitial proliferation occurs at about - days after onset, which presents fi rstly lobular septum and intralobular thickening and subpleural curvature shadows. then there is the shrinkage of intrapulmonary patchy shadows with increase of density and gradual formation of high-density cord and honeycomb shadows inside lungs. severe pulmonary interstitial proliferation could diminish lung volume and compel the mediastinum to affected side. besides, pulmonary interstitial proliferation could occur extensively inside lungs, predominated by lobular septum, intralobular interstitial thickening, and subpleural curvature shadows, as well as irregular high-density plaque and cord-like shadows at some regions. honeycombshaped shadows and tractional bronchiectasis are signs of pulmonary interstitial fi brosis. ct reveals cord, reticulate, and honeycomb-shaped shadows, and hrct could better visualize the subtle abnormal changes of pulmonary interstitium, such as lobular septum thickening, intralobular interstitial thickening, subpleural curvature shadows, and honeycomb-shaped shadows. whether the cord-like and reticulate changes visualized by ct are completely absorbed or induce persistent fi brosis still entails long-term follow-up observation ( fig. . ). numerous sars patients develop osteonecrosis after recovery, which mostly appears as femoral head necrosis and also possibly necrosis of the ankle joint, knee joint, and shoulder joint. scans of bilateral hips and knees of sars patients in rehabilitation phase disclosed that patients ( %) suffered from osseous abnormality, of which bone ischemia accounts for % and implication of three sites for %. its pathogenesis still remains not completely ascertained, and it may be ascribed to improper use of hormones and injury of sclerotin from sars viruses, bacterial embolism, lipid embolism, or multiple mixed factors. the primary pathological change of aseptic necrosis of femoral head is degenerative alteration due to femoral head blood circulation disturbance, mainly characterized by limping and pain. the lower limb of the affected side is relatively shorter and presents with light fl exion and adduction, as well as with slightly limited abduction and intorsion. initial x-ray examinations suggest that the involved femoral head has normal appearance but inhomogeneous density, with spotted osteoporosis. gradually ensue the increase of density as a result of osteonecrosis, which meanwhile is interspersed with some transparent shadows. femoral head may become fl at, fragmented, and irregular in contour from pressures. typical femoral head necrosis appears in the shape of wedge, with the base toward articular surface and the top toward metaphysis. since lesions are dominantly confi ned at the anterosuperior area of femoral head, lateral imaging of hip joints could clearly reproduce the extent of lesions. the necrotic area is encircled by a ring of transparent band. hip articular cavity may become wider and irregular due to fl attening of the femur. sometimes, there may be complication of dislocation. the fragments of femoral head may be shed off into the joint, giving rise to joint mouse. normal femoral head is located at the acetabular center in the transverse ct image and may appear in the shape of sphere or hook on different layers, with delicate high-density bone cortex at the periphery. the high-density trabeculae inside femoral head are arranged in stelliform formation from the center continually to bone cortex, named as "stelliform sign." ct manifestations of aseptic necrosis of femoral head can be divided into early and advanced phase. in the early phase, the stelliform sign formed by the trabeculae of femoral head is deformed or absent, presenting spotted and patchy shadows of increased density inside femoral head and fasciculate changes or mutual fusion at its periphery. in the advanced phase, femoral head become ruptured and out of shape, and absorption of sclerotin between bone fragments appears as irregular low-density area, with stelliform sign disappeared. mri manifestations of aseptic necrosis of femoral head include: necrotic areas of femoral head of early cases have no repair reactions or mechanic collapse and still maintain normal lipid signals of medullary bones. however, lowsignal margins incurred by sclerosis reactions could be seen at the periphery of foci. the t high signals of normal femoral head on t -weighted image are detected with black linear low signals (representing demarcation between normal and ischemic bone tissues). on t -weighted image, a high-signal line is visible at the medial low-signal sclerotic reaction line, thus forming the typical double-line sign. the double-line sign could refl ect congestion and infl ammation of granula-tion tissues and proves specifi c to aseptic necrosis of femoral head. severe infl ammation, congestion, fi brosis, and sclerosis could greatly deplete the amount of lipids of femoral head, which presents liquid-like signals on mri, low signals on t -weighted image, and high signals on t -weighted images. advanced cases are typical of fi brosis and sclerosis, and the affected area present with fi brotic mri features, that is, low signals on t -and t -weighted and proton densityweighted imaging. complication with effusions in hip joints appears as t low signals and t high signals. based on mri presentations, aseptic necrosis of femoral head could be classifi ed into four phases: phase i is characterized by homogenous or inhomogeneous low-signal areas adjacent to the joint above the femoral head on t -weighted imaging; phase ii is visualized with wedge-shaped lowsignal bands; phase iii presents with sequestrum crescent sign and cortex collapse; phase iv manifests joint degeneration and joint space narrowing besides presentations of phase iii. in light of the aforementioned mr staging criteria, sars cases complicated by aseptic necrosis of femoral head already reported are mostly in phase ii. additionally, aseptic necrosis after use of hormones, besides most frequently involvement of the femoral head, could also implicate other joints, such as the knee joint, elbow joint, ankle joint, and acetabulum. therefore, if necessary, mr examinations of other joints shall be conducted to ascertain the extent and degree of lesions. if sars patients complain of pain at both hip joints and limited movement, ct scan or mri examination of both hip joints is recommended to conclude defi nite diagnosis. x-ray plain scan is not sensitive to early bone ischemic necrosis, which therefore is 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radiographic and ct fi ndings imaging of severe acute respiratory syndrome in hong kong chest ct manifestations of severe acute respiratory syndrome chest roentgenographic features of severe acute respiratory syndrome imaging follow-up of sars patients complicated with pulmonary fi brosis x-ray contrast study of severe and ordinary sars patients complicated with pulmonary fi brosis and femoral head necrosis overview and outlook of treatment of sars patients clinical presentations and outcome of severe acute respiratory syndrome in children transmission dynamics and control of severe acute respiratory syndrome key: cord- -upwe cpj authors: sullivan, kathleen e.; bassiri, hamid; bousfiha, ahmed a.; costa-carvalho, beatriz t.; freeman, alexandra f.; hagin, david; lau, yu l.; lionakis, michail s.; moreira, ileana; pinto, jorge a.; de moraes-pinto, m. isabel; rawat, amit; reda, shereen m.; reyes, saul oswaldo lugo; seppänen, mikko; tang, mimi l. k. title: emerging infections and pertinent infections related to travel for patients with primary immunodeficiencies date: - - journal: j clin immunol doi: . /s - - - sha: doc_id: cord_uid: upwe cpj in today’s global economy and affordable vacation travel, it is increasingly important that visitors to another country and their physician be familiar with emerging infections, infections unique to a specific geographic region, and risks related to the process of travel. this is never more important than for patients with primary immunodeficiency disorders (pidd). a recent review addressing common causes of fever in travelers provides important information for the general population thwaites and day (n engl j med : - , ). this review covers critical infectious and management concerns specifically related to travel for patients with pidd. this review will discuss the context of the changing landscape of infections, highlight specific infections of concern, and profile distinct infection phenotypes in patients who are immune compromised. the organization of this review will address the environment driving emerging infections and several concerns unique to patients with pidd. the first section addresses general considerations, the second section profiles specific infections organized according to mechanism of transmission, and the third section focuses on unique phenotypes and unique susceptibilities in patients with pidds. this review does not address most parasitic diseases. reference tables provide easily accessible information on a broader range of infections than is described in the text. . this review covers critical infectious and management concerns specifically related to travel for patients with pidd. this review will discuss the context of the changing landscape of infections, highlight specific infections of concern, and profile distinct infection phenotypes in patients who are immune compromised. the organization of this review will address the environment driving emerging infections and several concerns unique to patients with pidd. the first section addresses general considerations, the second section profiles specific infections organized according to mechanism of transmission, and the third section focuses on unique phenotypes and unique susceptibilities in patients with pidds. this review does not address most parasitic diseases. reference tables provide easily accessible information on a broader range of infections than is described in the text. physician be familiar with emerging infections, infections unique to a specific geographic region, and risks related to the process of travel. this is never more important than for patients with primary immunodeficiency disorders (pidd). a recent review addressing common causes of fever in travelers provides important information for the general population [ ] . this review covers critical infectious and management concerns specifically related to travel for patients with pidd. this review will discuss the context of the changing landscape of infections, highlight specific infections of concern, and profile distinct infection phenotypes in patients who are immune compromised. the organization of this review will address the environment driving emerging infections and several concerns unique to patients with pidd. the first section addresses general considerations, the second section profiles specific infections organized according to mechanism of transmission, and the third section focuses on unique phenotypes and unique susceptibilities in patients with pidds. this review does not address most parasitic diseases. reference tables provide easily accessible information on a broader range of infections than is described in the text. emerging infectious diseases are a result of a convergence of numerous factors and comprise complex interactions among multiple variables. some of those factors are human movement, land use change, encroachment and wildlife translocation, rapid transport, and climate change. several studies demonstrate that for a pathogen to persist in a population, a minimal host population size that is specific for the type of pathogen and host population. of particular relevance to emerging infections is the pattern of rapid population growth. in the tropics, before wwii, most regional ecosystems consisted of few large cities and scattered human settlements separated by large areas of cropland, pastureland, or undisturbed forest. today, the pattern is the opposite: many large cities have developed with only patches of undisturbed forest or grassland. domestic vectors have therefore expanded their population with increasing urbanization and this markedly impacts the interactions between vectors and pathogens [ ] . human activities such as deforestation, use of pesticides, pollution, etc. lead to the loss of predators that naturally regulate rodent and insect populations. this contributes to emerging zoonotic diseases and explains why they occur more frequently in areas recently settled. in today's global economy and affordable vacation travel, it is increasingly important that visitors to another country and their the southern, with a reduction in the number of cold days per year, changes in rainfall (more winter precipitation and summer droughts) [ ] , and together these changes increase the risk of several vector-borne diseases in new areas. climate change involves not only global warming but also changes in precipitation, wind, humidity, and the location and frequency of extreme weather events like floods, droughts, or heat waves. changes in climate produce changes in pathogens, vectors, hosts, and their living environment. increases in precipitation can increase mosquitoes for example, but heavy rainfalls may cause flooding that temporarily eliminates larval habitats and decreases mosquitoes. flooding may force rodents to look for new habitats in houses and increase the opportunities of vector-human interactions, as occurs for example in the case of epidemic leptospirosis. humidity is another very important factor of climate change in the development of vector-borne diseases. mosquitoes and ticks do not survive well in dry conditions. therefore, weather impacts infectious pathogen distribution in complex ways that are not predictable by the forecast. extreme weather events can precipitate outbreaks of infection. an increase in the frequency and intensity of natural disasters like hurricanes and tsunamis, in relation to the el niño/southern oscillation phenomenon, may result in more flooded grasslands, which favor the breeding of aedes and culex mosquitoes [ ] , and impact water sanitation fostering outbreaks such as cholera. flooded areas can displace rodents leading to plague. tornados and other severe weather can stir up soil leading to infections with soil fungi leading to episodic outbreaks of invasive fungal disease such as mucormycosis such as apophysomyces trapeziformis [ ] . malaria is a common disease that can vary dramatically depending on weather, and extreme weather can alter the very landscape, providing new bodies of water to support larval development. if the melting of glaciers and the polar ice caps bring coastal cities underwater, or if overpopulation and waste cause drinkable water shortages in certain regions of the world, we can expect mass migrations. these could change the patterns of infection and drive outbreaks. migrants traversing tropical forests, or feeding with meat from game or carcasses, are but two scenarios that could be envisioned for the emergence of zoonotic infectious diseases [ ] . several predictive models have been developed to evaluate the impact of climate change on the emerging infectious diseases: climex, dymex, miasma models. nevertheless, it remains difficult to predict when and where pathogen behavior will deviate from its typical pattern. climate change primarily affects vector-borne diseases by increasing rates of reproduction and biting and by shortening the incubation period of the pathogen they carry. ticks have gained spread from the mediterranean basin to northern and eastern europe, as well as appearing at higher altitudes. increased survival, density, and activity have also been reported following shorter, milder winters. climate change has also resulted in more days of activity per year for mosquitoes. as temperatures rise, more parasites are viable within regions ranging from the mediterranean and tropical zones, up to the balkans, russia, scandinavia, and the uk. for some ticks and fleas, temperatures over °c with relative humidity of over % are optimal for their proliferation and activity throughout the year [ , , ] . for example, dengue fever is usually limited to a tropical latitude and a low altitude, since mosquito eggs and larvae lose viability with sustained low temperatures. during unusually warm summers, however, dengue has been reported as high up as m above sea level. warmer temperatures also result in smaller adult mosquitoes, which need to bite more frequently to feed themselves and be able to lay eggs, thus increasing the rate of transmission [ ] . in contrast, the incidence of malaria has followed mixed patterns of increase and decrease along recent decades, and computer models have failed to predict the spread. the explanation for this is, in part, that climate change also results in diminished survival of the vectors (warming over °c affects the survival of both parasites and vectors), and in part, that the effect of climate change is non-linear and complex [ ] . the frequency of emerging vector-borne infections varies per changes in land use, human activity, intervention maneuvers to eradicate the vector or prevent transmission to humans, drug treatment, and vaccines. both ecologic and economic changes may bring together rodents and humans. hunting activities may change vector distribution and large-scale animal movements can impact disease distribution. impoverishment of cities and overcrowding in slums, but also reforestation, golf club development, and the urbanization of rural suburbs facilitate exposure to ticks and rodents [ ] . many patients with pidd require immunoglobulin replacement. immunoglobulin products have been demonstrated to improve outcomes in hepatitis a and measles [ , ] . at least some neutralizing antibody is present directed to rsv and group b streptococci [ , ] . this raises three distinct issues for patients: ( ) difficulty in the diagnosis of infections in travelers because locally produced immunoglobulin may have antibody titers to local infections that are not typical for other countries, ( ) safety concerns about locally produced immunoglobulin, if the patient resides in the location long enough to require immunoglobulin from a local provider, ( ) the decision to use locally produced immunoglobulin products to provide superior prevention of infection compared to the patient's usual product. there are limited data on each of these subjects. serologic diagnostic testing in patients on immunoglobulin therapy will be addressed below in terms of issues related to lack of specific antibody produced by the patient (potentially) after infection. the converse can also be an issue. patients arriving from countries with significant occurrences of infections unusual in their current country may have igg antibodies to those infections simply through their immunoglobulin product and not reflecting a true infectious event in the patient. this can lead to diagnostic confusion when serologic testing demonstrates the presence of antibodies due to the infusion product. patients will often ask if immunoglobulin products from other countries are held to the same rigorous standards as their home country. today, commercially produced immunoglobulin is safe and tightly regulated. all commercially produced immunoglobulin around the world has one of the time-tested viral inactivation procedures such as nanofiltration, caprylate absorption, pasteurization, solvent/detergent, vapor heating, and low ph treatment. these procedures uniformly inactivate enveloped viruses. many emerging viruses are specifically tested for their ability to withstand the purification process. much has been learned since the transmission of hepatitis c viruses through immunoglobulin products in the early s [ ] . nevertheless, vigilance is important. in , counterfeit immunoglobulin was identified. therefore, patients should ensure that they receive only brand name products while traveling. the subject of whether a patient's interests would be best served by using a local immunoglobulin product, with antibodies to pathogens that are prevalent in the community, or have their home physician ship their usual product, for which the patient has a known tolerance is hotly debated. patients with a history of intolerance to immunoglobulin products should not switch unless necessary. however, there are compelling reasons to consider a locally produced product when patients are in a foreign country for an extended period. it is known that antibodies to west nile virus have tracked with the distribution of the virus as it has emerged in several areas [ , ] . titers in products using donors from the usa have higher neutralizing titers to west nile virus than those using donors from europe, although there is a -fold difference in titers between lots from the usa [ ] . similarly, protective antibodies to concerning pathogens may be optimal in locally produced immunoglobulin. it is critical to inquire where the plasma source is derived. in most countries, the utilized immunoglobulin is produced in europe or the usa. having a different name does not ensure that the plasma pool comes from a different country. most lots of immunoglobulin are produced from to , plasma donors. the infrastructure to support such an endeavor is not easy to establish in each geographic area. serologic testing is commonly used for the diagnosis of infection. this approach relies upon detection or quantitation of antibodies made by the host against specific pathogens. the presence of igm antibodies against a specific pathogen indicates recent infection, while igg antibodies against a specific pathogen indicate past infection. importantly, serologic testing can only be applied for the diagnosis of infection if the host can mount a specific antibody response to the pathogen. conversely, serology cannot be relied upon to diagnose infection in the setting of immune deficiency where there is impairment of the specific antibody response, such as in the case of primary antibody deficiencies, cellular immune deficiencies, combined immune deficiencies, and other secondary immune deficiencies affecting t and/or b cell function. in these situations, the causative pathogen must be identified by alternate means such as culture of the organism, antigen detection, or molecular approaches (nucleic acid hybridization, nucleic acid sequencing, or oligonucleotide probe arrays). molecular approaches are particularly relevant for the diagnosis of infection in patients with pidd. signal and target amplification techniques can be coupled with nucleic acid hybridization or probe assays to allow detection of pathogen dna or rna that is present in very small amounts in clinical samples. in patients with pidd, vaccines could play an important role in preventing infections with vaccine-preventable diseases. even pidd patients may generate some protective responses [ , ] . the decision to immunize such patients or not depends on the type and severity of pidd as well as the type of vaccine to be administered (live or inactivated) ( table ). in general, inactivated vaccines are safe for pidd patients while immunization with live attenuated vaccines is a known hazard to patients with serious immunodeficiencies of t cell, b cell, and phagocytic cell origin (table ). in less severe pidd, the vaccine can induce adequate protection as in healthy individuals or the efficacy may be reduced [ ] [ ] [ ] . of note, immunoglobulin replacement therapy induces passive immune protection to some vaccine-preventable infections, such as measles, mumps, rubella (mmr), and varicella. in addition, live viral vaccines have greatly reduced efficacy while on immunoglobulin replacement. therefore, vaccine administration in patients receiving regular immunoglobulin replacement treatment should be withheld until at least to months (depending on dose) after cessation of such treatment, if cessation and vaccination are safe. in addition, pidd patients who have received hematopoietic stem cell transplantation but have incomplete immune reconstitution or are under immunosuppression should not receive live attenuated vaccines. in general, the decision of administering live viral vaccines should be made by clinical immunology experts [ ] . in developing countries where polio is still endemic and oral polio vaccine is essential for eradicating the disease, it is of utmost importance that all pidd patients and family members should not receive live oral polio (opv) because of the reported prolonged excretion of the virus for months and even years [ ] . in addition, vaccine-associated paralytic polio is a real risk for some with pidd. these patients and family members should receive inactivated polio vaccine (ipv) instead of opv. similarly, the hazards of administering bacillus calmette-guerin (bcg) vaccine to pidd patients have been documented. in a series of bcg, vaccinated severe combined immunodeficiency (scid) patients from centers in countries, % of scid patients developed disseminated bcg infection and had the worst outcome [ ] . patients with chronic granulomatous disease vaccinated with bcg also developed local and disseminated bcg infection. recently, vaccine strains of rubella virus were found to be associated with skin granulomas in pidd patients [ ] [ ] [ ] . siblings and household contacts of patients with suspected or diagnosed pidds should receive all the national immunization scheduled vaccines. ipv should be substituted for op in families where an antibody-deficient patient exists. yearly influenza vaccination of family members is recommended in order to reduce the risk of household-social transmission [ , , ] . diseases where routine vaccination has reduced incidence can occasionally experience a resurgence in times of economic hardship with reduced attention to vaccination. diphtheria is currently seen in venezuela for this reason. war and disruption of health infrastructure are other common reasons for resurgence in vaccine-preventable diseases. in other settings, antivaccination sentiment has led to outbreaks of diphtheria, measles, and mumps. an additional consideration is that not all countries provide the same level of vaccination, and therefore, vaccine-preventable illness can still be seen regionally. these outbreaks represent a significant risk to patients with pidd. a universal consideration for patients with pidd is the concern about antibiotic resistance, which varies dramatically around the world. for certain high impact infections, the emerging antibiotic resistance patterns will be discussed below. antimicrobial resistance occurs naturally, but misuse and overuse of antimicrobials are accelerating this process. in nearly every country, antibiotics are overused and misused in people and animals leading to antibiotic resistance in every country. key resistance patterns to common bacteria include emergence of carbapenem-resistant klebsiella pneumoniae around the world with high frequency of resistance (due to different mechanisms) in the mediterranean, with greece, italy, and turkey having endemic spread of this pathogen [ ] . carbapenem-resistant strains among other genera of enterobacteriaceae have also been recognized. they are particularly common in greece, but have been found widely distributed [ ] . the new delhi metallo-beta-lactamasemediated resistance, which is endemic in the indian subcontinent but becoming increasingly spread worldwide, is a growing concern [ , ] . as a common cause of urinary tract infections, colistin is the only recourse when carbapenemresistant enterobacteriaceae, and colistin resistance has recently emerged in small outbreaks throughout the world [ ] . in these cases, the infection is essentially untreatable. fluoroquinolone-resistant escherichia coli, a common cause of urinary tract infections, now accounts for over half of the isolates in some asian countries [ , ] . t he emergence of resistance to antibiotics in grampositive pathogens has become a major international problem as there are fewer, or even sometimes no effective, antimicrobial agents available for infections caused by these bacteria. methicillin-resistant staphylococcus aureus is common in many countries and in fact has spawned a nomenclature recognized by the general public: mrsa. several studies have reported resistance to the newer antimicrobial agents like linezolid, vancomycin, teicoplanin, and daptomycin [ ] . thus far, these isolates appear to be uncommon and have been found in < % of isolates in brazil, china, ireland, and italy, with nearly undetectable rates elsewhere. vancomycinresistant enterococcus (vre) is growing in frequency and can now be a cause of primary bacteremia in immunocompromised individuals [ ] . a key message is that antibiotic resistance is increasing (generally) and travelers should be alerted to resistance to commonly encountered organisms, and if antibiotic prophylaxis is required, their prophylaxis is adjusted. neisseria gonorrhoeae is a specific organism for which resistance has become especially problematic. it has progressively developed resistance to virtually all antimicrobial agents since introduction of sulfonamides in mid- s. the current treatment guidelines recommend dual antimicrobial therapy (ceftriaxone - mg × plus azithromycin - g × ) as first-line regimen [ , ] . although dual therapy is very effective, development of concomitant ceftriaxone and azithromycin resistance is likely to emerge [ ] . the risk of untreatable n. gonorrhoeae demands better global antimicrobial surveillance system, clinical trials on combined therapy of existing drugs as well as novel agents in monotherapy, and development of a gonococcal vaccine. for pidd patients, guidance on barrier methods for the prevention of sexually transmitted diseases should be a part of any pre-travel counseling. mycobacteria tuberculosis (mtb) is an age-old pathogen with emerging resistance. drug-resistant tb, fueled by the hiv epidemic, is a global threat. in , who estimated , new cases of multidrug-resistant tb (mdr-tb) and an additional , new cases of rifampin-resistant tb (rr-tb) who would also require mdr-tb treatment. treatment of mdr-tb and mycobacterium bovis disease is beyond the scope of this text and reader is referred to recent who mdr treatment guidelines [ ] . regions of the world with > % mdr tb include regions of azerbaijan, kazakhstan, russia, uzbekistan, china, georgia, and eastern europe. extensively drug-resistant tb (xdr tb) refers to mtb resistant to isoniazid, rifampin, any fluoroquinolone and at least one second-line drug. xdr tb has been reported in countries. on average, % of patients with mdr tb have xdr tb. as is true for all types of mtb, xdr tb is contagious and small outbreaks related to person-person transmission have been reported. non-tuberculous mycobacteria (ntm) cause significant systemic infections in patients with defects of the il- / ifnγ/stat axis as well as in gata deficiency and can cause significant pulmonary infections in pidd patients with bronchiectasis. compared to tb, ntm is acquired from the environment and not from person-to-person transmission; therefore, acquisition of antibiotic-resistant strains is less common. however, in these individuals with pidd, ntm disease is often chronic and can be difficult to eradicate, and resistance can then easily develop during therapy. using combination of antibiotics is essential, and consultation with those familiar with treatment of treatment refractory ntm disease is recommended. aspergillus species are ubiquitous inhaled molds with worldwide distribution that cause opportunistic infections in immunocompromised patients [ ] . aspergillosis also occurs in pidds associated with quantitative and/or qualitative phagocyte defects; it develops most frequently in chronic granulomatous disease (cgd) patients (prevalence,~ %), while it is seen less often in patients with gata deficiency, card deficiency, and congenital neutropenia syndromes [ , ] . upon inhalation, aspergillus species cause invasive pulmonary disease in susceptible hosts with the exception of card deficiency, where aspergillosis has a predilection for extrapulmonary tissues with sparing of the lungs [ ] . diagnosis is established by fungal culture and/or histopathology showing acute-angle septate hyphae and/or detection of galactomannan, an aspergillus cell wall component released during invasive infection, in serum or bronchoalveolar lavage fluid [ ] . while azole-susceptible aspergillus fumigatus is still the most common infecting species in pidd patients, the emergence of azole-resistant a. fumigatus and nonfumigatus aspergillus species underscores the importance of a high index of suspicion in patients who do not respond to front-line voriconazole treatment [ ] . the advent of fungal molecular diagnostics has demonstrated that patients with pidds are more prone to infections by uncommon low-virulence aspergillus species with intrinsic resistance to azole antifungal agents that do not infect patients with iatrogenic immunosuppression. these primarily include aspergillus viridinutans, aspergillus tanneri, and neosartorya udagawae, which pose major diagnostic and therapeutic challenges due to their impaired sporulation and propensity for contiguous and distant tissue spread, respectively. in addition, acquired azole resistance in a. fumigatus can be seen in patients on long-term exposure to azole drugs used as treatment and/or prophylaxis [ ] . azole resistance in these strains is predominantly caused by point mutations in the lanosterol α-demethylase gene that encodes the cyp a protein, the primary target of azole drugs. importantly, infection by azole-resistant a. fumigatus strains without prior exposure of patients to azole antifungals has recently emerged as an important global health concern due to the widespread use of sterol demethylase inhibitor fungicides in agriculture that results in cross-resistance with the triazole antifungals used in clinical practice [ ] [ ] [ ] . fungicide-driven azole-resistant environmental aspergillus strains were first observed in the netherlands in and have since then been documented in other parts of europe, south and north america, the middle east, australia, africa, and asia. the prevalence of these azole-resistant aspergillus strains among clinical aspergillus strains recovered from patients in european countries was reported to be . %, while alarmingly in some areas > % of recovered strains exhibited azole resistance [ ] . the emergence of such aspergillus environmental strains poses serious threats to the treatment of immunosuppressed patients. mortality rates as high as % have been seen due to delays in diagnosis and suboptimal treatment with azole antifungals [ ] . although no prospective data exist for the treatment of patients with such resistant fungi, the use of amphotericin b-and echinocandin-based regimens are preferred over azoles [ ] . in the absence of azoles, the lack of alternative oral antifungal agents is particularly challenging for pidd patients such as those with cgd who require long-term suppressive antifungal treatment. candida species are commensal yeast fungi that colonize the mucosal surfaces of~ % of healthy individuals [ ] . when perturbations in immunity and/or microbiota occur, candida causes opportunistic mucosal or systemic infections that depend on clearly segregated immune responses for host defense. specifically, t cells of the th program are critical for mucosal and phagocytes for systemic immunity [ ] . indeed, a proportion of patients with cgd and complete myeloperoxidase deficiency develop systemic, but not mucosal candidiasis [ ] , whereas patients with monogenic syndromes of chronic mucocutaneous candidiasis due to mutations in the il- signaling pathway (il ra, il rc, il f, act ) or in other genes that adversely affect th differentiation (rorc, stat , stat , aire, dock , stk , irf ) do not develop systemic candidiasis. the only known pidd to date that results in combined mucosal and systemic candida infection susceptibility is card deficiency. systemic candidiasis in card -deficient patients has a predilection for the central nervous system, associated with brain-specific impaired recruitment and effector function of neutrophils [ ] [ ] [ ] . diagnosis of candida infections is established by culture. azole-susceptible candida albicans is still the most common infecting species in pidd patients; however, emergence of azole-resistant c. albicans is not uncommon during chronic azole use, making long-term therapy challenging due to lack of alternative oral antifungal treatment options [ ] . beyond c. albicans, non-albicans candida species can rarely infect pidd patients, some of which have intrinsic resistance to azole antifungals, including candida glabrata and candida krusei [ ] . most recently, candida auris has emerged as a global public health concern due to its resistance to antifungal drugs, high virulence potential, propensity for health careassociated horizontal transmission and outbreaks in health care settings, persistence in the human skin and hospital environment, inherent resilience to antiseptics, and misidentification by routine biochemical methods as other yeasts (most often candida haemulonii, but also candida famata, rhodotorula glutinis, or saccharomyces cerevisiae). c. auris was first recovered from the ear canal of a patient in japan in and has since then been reported to cause life-threatening infections and hospital outbreaks in europe, asia, africa, the middle east, and south and north america [ ] [ ] [ ] [ ] . most of the reported strains of c. auris have intrinsic resistance to fluconazole and other triazole antifungal agents, while a significant proportion of strains has elevated minimum inhibitory concentrations to amphotericin b and echinocandins, with some strains reportedly resistant to all three classes of azoles, polyenes, and echinocandins [ ] . avoidance of azole antifungals is important in c. auris-infected patients, and echinocandinor amphotericin b-based regimens are preferred, guided by strain-specific in vitro susceptibility patterns. this section on vector-borne infections is a major focus of this review because the infections are often more severe in immunocompromised individuals and because there are mitigation strategies that should be considered even in the absence of defined medical treatments for infection. prevention of mosquito bites depends somewhat on the endemic species but there are generalizations. the use of a mosquito repellant such as deet, oil of lemon eucalyptus, ir , or picaridin is as important as using long sleeves and long pants while in an affected area. deet and picaridin are safe in pregnancy, and some data support their greater efficacy [ ] . air conditioning and fans tend to keep mosquitoes away but netting at night is essential in mosquito-prone areas. light-colored clothing is less attractive to mosquitoes than dark clothing, and scented detergents and use of dryer sheets tend to attract mosquitoes, hence should be avoided. aedes species prefer to bite indoors and thrive in urban areas with small puddles of water. they bite most frequently around dawn and dusk. anopheles species have very similar behaviors. culex mosquitoes, in contrast, bite primarily at night. tick and fly bite prevention is focused on physical and chemical prevention. for ticks, physical inspection for biting ticks should also be incorporated. arthropod-borne viruses (arboviruses) are transmitted to humans through the bites of infected insects: mosquitoes, ticks, sand flies, or midges. some arboviruses can be transmitted through blood transfusion, organ transplantation, perinatal transmission, consumption of unpasteurized dairy products, or breastfeeding. there are > arboviruses causing human disease. most arboviral infections are asymptomatic. infectious manifestations range from mild febrile illness to severe encephalitis. arboviral infections are often categorized into two primary groups: neuroinvasive disease and non-neuroinvasive disease. tables and list the encephalitigenic viruses and the febrile/hemorrhagic disease causing viruses. in this section, we will highlight west nile virus, the most common of the encephalitogenic arboviruses. west nile virus is a single-stranded mosquito-borne rna virus of the family flaviviridae. the natural transmission cycle of the virus occurs in culex mosquitoes and birds. humans and horses are dead-end hosts for the virus. the most common mode of transmission to humans is through the bite of infected mosquitoes. other less common modes of transmission include blood transfusions, organ transplantations, occupational exposure in laboratories and mother-to-child transmission during pregnancy, childbirth, and breastfeeding. west nile virus has been diagnosed in > people in the usa with slightly more than half having neuroinvasive disease. since , > , people in the usa have been infected. it is also common in africa, europe, and asia [ ] . infection with west nile virus is asymptomatic in most individuals [ , ] . the incubation period lasts for to days. however, it can be significantly longer in immunocompromised hosts. clinical manifestations following infection develop in - % of those infected and include fever, headache, myalgia, arthralgia, vomiting, and rash. severe neuroinvasive disease leading to meningitis, encephalitis, and flaccid paralysis develops in less than % of infected individuals. the overall case fatality is approximately % which is a disproportionately high mortality in patients with encephalitis and myelitis. diagnosis of west nile virus rests on demonstration of specific antibody responses especially specific igm antibodies in the serum or csf of infected individuals by enzyme immunoassays. plaque reduction neutralization tests can differentiate cross-reactive antibodies. detection of virus in csf, blood, or tissue specimens by culture or pcr is particularly useful in immunosuppressed individuals who may have impaired serological responses. west nile virus has been reported in the context of both primary and secondary immunodeficiency. severe neurological manifestations have been reported in hiv-positive individuals, individuals receiving immunosuppressive therapy including rituximab, and individuals with pidd. infection with wnv has been reported in individuals with common variable immunodeficiency, idiopathic cd lymphopenia, gata deficiency, and a case of probable good's syndrome [ ] [ ] [ ] . individuals with antibody defects, neutropenias, and impaired t cell responses are potentially at an increased risk of severe manifestations of wnv disease including severe neurological involvement. this section highlights the four important non-neuroinvasive arboviruses based on current geographical distribution: dengue, yellow fever, zika, and chikungunya (table ) . approximately countries/territories have reported local transmission for both chikungunya and dengue viruses [ ] . dengue is due to infection with one of four dengue virus serotypes, transmitted by a mosquito (typically aedes aegypti). this febrile illness affects all ages with symptoms appearing - days after the infective bite. symptoms range from mild to high fever, with severe headache, musculoskeletal pain, and rash. severe dengue (also known as dengue hemorrhagic fever) occurs in . - % of cases and is characterized by fever, abdominal pain, persistent vomiting, bleeding, and breathing difficulty and is a potentially lethal complication [ ] . paradoxically, the main risk factor for dengue hemorrhagic fever is pre-existing antibodies. early clinical diagnosis and comprehensive management by experienced clinicians increase survival. dengue is ubiquitous throughout the tropics with the highest infection rates in the americas and asia. dengue is now endemic in countries, causing up to million infections a year and , deaths, mainly among children. over half of the world's population inhabits areas at risk for dengue infection [ ] . the presence of a. aegypti in over countries potentially puts almost the whole world at risk of becoming infected with this virus. pcr is widely used as serologic methods to diagnose dengue. immunity to one serotype does not confer protection against the other three serotypes, and heterologous antibody may be a risk factor for hemorrhagic dengue [ ] . the natural history of dengue has been studied in hiv patients where hiv did not appear to increase severity. there have been no reports of patients with pidd having dengue; however, dengue infection after solid transplantation has been reported [ ] [ ] [ ] [ ] with some patients having severe complications suggesting that t cell compromise in pidd could be a risk for severe disease. there are no antiviral medications utilized for dengue virus. care of patients with hemorrhagic disease requires meticulous approach to fluids and coagulation status. one dengue vaccine has been registered in several countries (cyd-tdv) for individuals from to (or ) years old. it is a live attenuated recombinant tetravalent vaccine with backbone of the attenuated yellow fever d virus genome with the prm and e genes that encode the proteins from the four wild-type dengue viruses. the who has suggested its use in regions where seroprevalence of dengue virus of any serotype is % or greater, but has not recommended it to hiv-infected, immunocompromised individuals, nor pregnant or lactating women [ ] . most people infected with the yellow fever virus have no illness. symptoms of yellow fever include sudden onset of fever, chills, headache, musculoskeletal pain, nausea, vomiting, fatigue, and weakness. the incubation period is typically - days, and symptoms may appear after return from travel. most people improve after the initial presentation, but % of cases progress to develop a more severe form of the disease, usually after a day of presumed recovery. the severe form is characterized by high fever, jaundice, bleeding, and eventually shock and failure of multiple organs [ ] . yellow fever virus is an rna virus that belongs to the genus flavivirus. it is transmitted from mosquitoes after biting an infected primate. it is widely distributed in the equatorial tropics [ ] . aedes species of mosquitoes are primarily responsible for transmission. large epidemics of yellow fever occur when the infection enters heavily populated areas with a high mosquito density and where most people have little or no immunity. west africa has undergone a large-scale vaccination campaign with impressive results and yellow fever is now uncommon in west africa [ ] . serologic testing for yellow fever is the diagnostic standard. pcr can be performed on tissue samples. there are no published studies of yellow fever in immunocompromised people, but the elderly, very young, people with autoimmune disease, or who are post-thymectomy are at risk from the attenuated vaccine strain. thus, it seems likely that any immunodeficiency would be associated with more severe wildtype disease. currently, no specific antiviral drug for yellow fever exists. treatment of dehydration, liver and kidney failure, and fever improves outcomes. the yellow fever vaccine is highly effective; however, immunodeficient patients should not receive it. infection with zika virus is often asymptomatic. it represents a mild infection for those who have any symptoms [ ] . the zika virus has been detected in urine, semen, and saliva of infected individuals, and transmission from transfusion and sexual relations has been reported. it is also detectable in breast milk, but breastfeeding-associated transmission has not been reported so far [ ] [ ] [ ] . contact with highly infectious body fluids from patients with severe zika infection has also been suggested as a possible mode of transmission [ ] . of tremendous importance is the presence of prolonged shedding of zika virus in a congenitally infected newborn [ ] . the main public health risk of zika virus is microcephaly in newborns from infected mothers [ ] . zika virus is capable of infecting human neural progenitor cells in vitro. infection results in disruption of cell cycle, increased cell death, and attenuated neuron growth [ ] . zika is not thought to be a major risk for people with pidd (based on the experience with hiv patients, but our recognition of zika is very recent. there is no known specific treatment for zika; however, there is an important effort to develop a vaccine. chikungunya fever is an acute febrile illness caused by the alphavirus, chikungunya virus. the incubation period is usually - days after the bite of an infected aedes mosquito. there is abrupt onset of high fever, and the fevers can be biphasic [ , ] . severe polyarthraligias develop after the onset of fever. the joint pains can affect any joint, but the pattern is usually symmetric and a true acute arthritis is not uncommon. the proportion of infected people with rash has varied across studies. when seen, the rash appears after the fever as a truncal maculopapular type of rash [ ] . cervical adenopathy is another common feature of infection. death is uncommon in chikungunya, but serious complications such as myocarditis have been seen. over half of the patients report continued joint symptoms year after acute illness and % have long-term joint symptoms [ ] . chikungunya originated in central/east africa. in forests, the virus circulates in aedes mosquitoes and non-human primates. in urban centers, the virus circulates between humans and mosquitoes similar to the pattern of dengue. there have been periodic urban outbreaks in asia and africa since with an acceleration in spread since [ ] . an important consideration is the periodic outbreaks with high attack rates in naïve populations. areas at risk currently are east africa, central africa, la reunion, india, and southeast asia. diagnostic testing utilizes pcr or serology. the threat to immunodeficient patients is not entirely clear. there are a few provocative cases where the immunocompromised appears to have been associated with fewer joint symptoms, but there were two patients, medically immune compromised, who had very severe disease [ , ] . this suggests that the presentation may be atypical and the course may be severe in immunodeficient people. treatment is supportive, although chloroquine, acyclovir, ribavirin, interferon-ɑ, and steroids have limited preclinical data to support clinical trials. babesia microti (the main species in the usa) infection can be asymptomatic, but many people develop fever, chills, headache, myalgias, anorexia, nausea, or fatigue [ ] . babesiosis often causes hemolytic anemia. b. microti is spread by ixodes scapularis ticks in the usa and babesia divergens (the main species in europe) is spread by ixodes ricinus. symptoms begin - weeks after a bite from an infected tick with b. divergens having a higher mortality rate and greater symptomatology compared to b. microti. the main geographic areas involved are the coastal eastern usa and cattle breeding areas throughout europe. the diagnosis is usually by inspection of a blood smear or through serology. a pcr test has just been developed. immunodeficiency, asplenia, and older age are recognized risk factors for severe disease and even death [ ] [ ] [ ] . thus, congenital asplenia would be considered a major risk for severe disease. a combination of atovaquone and azithromycin is generally used for therapy, although clindamycin and quinine have been used with success. patients with severe illness have been treated with exchange transfusions. five different types of plasmodium (plasmodium falciparum, plasmodium vivax, plasmodium ovale, plasmodium malariae, and plasmodium knowelsi) infect humans. malaria is transmitted primarily by female anopheles mosquitoes. symptoms vary depending on the type of plasmodium involved but usually include high fever, chills, and headache. in some cases, the illness can progress to severe anemia, kidney and respiratory failure, and death. the incubation period typically ranges from to days for p. falciparum, to days for p. vivax and p. ovale, and to days for p. malariae. in p. vivax and p. ovale infections, relapses can occur months or even years without symptoms. p. vivax and p. ovale have dormant liver stage parasites that must be specifically eradicated through medical therapy. malaria has been a global health concern throughout history and is a leading cause of death and disease across many tropical and subtropical countries. over the last years, new control measures have reduced malaria by over half [ ] . the democratic republic of the congo and nigeria account for over % of the estimated total of malaria deaths globally. high rates of malaria are seen in india as well. nevertheless, malaria exists in most tropical regions of the americas, africa, and asia [ ] . the diagnosis of malaria depends on the demonstration of parasites in the blood, usually by microscopy. the threat to immunodeficient patients is not entirely clear, but patients with hiv seem to have no additional burden of disease other than an increase in placental malaria, suggesting that t cells are not central to the defense of malaria [ , ] . asplenia is a known risk factor for severe malaria [ ] . antibodies appear to be both protective and pathologic [ , ] . treatment and prophylaxis depend on the region of the world because the parasites and resistance are highly variable and highly dynamic. therefore, it is best to consult an infectious disease specialist familiar with the prophylaxis before travel and for treatment of acute cases. leishmaniasis is due to infection with an obligate macrophage intracellular protozoa of the genus leishmania. it causes a spectrum of disease ranging from a cutaneous ulcer to mucosal disease and the most severe form, visceral leishmaniasis (vl). the liver, spleen, and bone marrow are major sites of parasite growth and disease pathology in vl [ ] . purely cutaneous leishmaniasis is most often caused by leishmania major, leishmania. tropica, leishmania aethiopica, leishmania infantum, and parasites belonging to the leishmania mexicana complex, the leishmania braziliensis complex, and the leishmania guyanensis complex. mucocutaneous disease is most often due to l. braziliensis complex, leishmania panamensis, leishmania amazonensis, and rarely by leishmania guyanensis. vl is most often caused by leishmania donovani and leishmania infantum (previously l. chagasi) [ ] . cutaneous leishmaniasis can have many variations but is most often an ulcer that develops after an indolent papule. the incubation period ranges from weeks to months. the ulcer usually heals within months to years, and there can be mild adenopathy. mucocutaneous leishmaniasis follows a cutaneous ulcer and is only caused by l. braziliensis parasites. oral and respiratory mucosa are most often involved with granulomatous lesions that may be extremely destructive. vl is associated with fever, lymphadenopathy, hepatosplenomegaly, wasting, hypoalbuminemia, and pancytopenia. this picture evolves over months to years. there can be secondary immune deficiency due to the pancytopenia. the epidemiology has changed dramatically and has been impacted by climate change [ ] . the sand flies that spread the parasite are affected by temperature and rainfall. in most endemic regions, leishmania has a patchy distribution due to micro-ecologic factors. poverty has been demonstrated to be a major risk factor for leishmaniasis [ ] . it has been estimated that up to half a million new cases of vl occur every year, but the majority are in resource-poor countries such as bangladesh, nepal, india, sudan, ethiopia, and brazil. emergence of resistance to antimony-based drugs has also led to a major resurgence of disease. the primary reservoir for leishmania is forest rodents, but dogs are increasingly important. the growing spread of leishmania is due to a combination of factors, and now countries have reported cases. immunodeficient patients are more susceptible to infection, and relapse occurs more frequently [ ] . the risk of developing vl is estimated to be between and times higher in hiv-infected than in non-hiv-infected individuals [ ] , and these patients have higher rates of treatment failure with the illness often taking a prolonged chronic course and higher mortality rates [ ] . a similar picture has been seen in patients with vl-infected post-kidney transplantation [ ] . dendritic cells, t cells, and the generation of reactive oxygen species have been shown to be essential for parasite control [ ] [ ] [ ] . pidd with impaired il- production have been associated with severe disease [ ] . a patient with cd l deficiency, associated with poor il- production, had chronic leishmania and died in spite of aggressive treatment. vl has been reported in cgd patients [ ] . six cgd patients were infected by leishmania, and they developed hemophagocytic syndrome with a poor outcome for one of them [ ] . the diagnosis of leishmaniasis is usually by visual inspection for parasites. immunofluorescence microscopy, direct agglutination, skin test, and pcr have been used. treatment is long-term and difficult. emerging resistance to first-line treatment is increasingly problematic. pentavalent antimonials are the mainstay of treatment in most countries, but liposomal amphotericin is widely used where resistance occurs. newer drugs with more favorable side effect profiles have been used in certain geographic settings: miltefosine, paromycin, and sitamaquine. rickettsiae are small gram-negative bacteria. they are obligate intracellular parasites, and the primary target in humans appears to be endothelial cells with subsequent thrombosis and clinical presentation of vasculitis [ ] . the rickettsiaceae family, originally defined by non-specific phenotypic characteristics, was reclassified into different strains and subspecies based on gene sequencing and genetic phylogeny ( table ). the clinical presentation of rickettsial disease can vary, but the classic triad of fever, rash, and headache still provides major clues for the diagnosis [ ] [ ] [ ] . however, rash is not an obligatory sign, and the incidence of rash can range between % for rickettsia conorii infection,~ % for rickettsia rickettsii, % for rickettsia africae, and less than % in the case of anaplasma phagocytophilum infection. therefore, fever in patients with exposure to a potential vector should raise a concern for a rickettsial disease, especially if there is also evidence of rash, inoculation eschar, or localized lymphadenopathy. additional supporting laboratory findings can include neutropenia, thrombocytopenia, and increase in liver transaminases. the geographic distributions of rickettsioses and ehrlichioses are mostly dependent on their vector distribution [ ] . as such, louse-borne and flea-borne are worldwide, reflecting the worldwide distribution of lice and fleas, with a tendency to parasite poor people in cold places and, characteristically, during wars. ticks, on the other hand, depend on their environment and most do not have a worldwide distribution. with the exception of the dog tick, vector for r. conorii in asia and north africa, r. rickettsii in the usa, rickettsia massiiae and erhlichia canis worldwide, most other tick-borne diseases are restricted to areas of the world correlating with the distribution of their vector [ ] . for that reason, it should be anticipated that climate and environmental changes will affect vector distribution and its reservoir host and, hence, the geography and epidemiology of tick-borne diseases [ , , ] . diagnosis presents a challenge, as it is extremely difficult to grow these organisms in culture. immunohistochemistry and pcr can be helpful. the severity of rickettsial disease varies with the causative agent and the host. r. rickettsii, rickettsia prowazekii, and orienta tsutsugamushi are considered most pathogenic. as for host factors, although severe and fatal cases have been described in healthy immunocompetent hosts [ , ] , there is evidence to suggest that children under the age of [ ] and immunocompromised hosts either secondary to hematologic malignancies, immunosuppressant treatment for organ transplantation, or hiv infection are at a greater risk to develop more severe disease with higher case fatality rates [ , ] . all rickettsiaceae are intracellular pathogens, and one could expect an increased risk for severe disease in pidds with abnormal t cell function. five to days of doxycycline is the preferred treatment for non-pregnant adults and children. treatment should not be delayed while awaiting diagnostic testing [ ] and can be given to children despite a minimal risk for dental staining. alternative treatments include azithromycin for mild disease [ ] and chloramphenicol for pregnant women. anaplasma is an intracellular bacterium that infects wild and domestic mammals, including man. a. phagocytophilum was formerly known as human granulocytotropic ehrlighiosis but is now known as human granulocytotropic anaplasmosis [ ] . e. chaffeensis infects monocytes and causes human monocytic ehrlichiosis [ ] . anaplasma and ehrlichia have historically cycled within non-human enzootic hosts, and man has become infected through increasing interactions with the environment. ehrlichia and anaplasma are transmitted by ixodes species of ticks, and their ranges include the eastern usa, south central usa, and scattered regions of europe, as far north as sweden. these infections have not been seen in humans in the southern hemisphere, but there are reports of organisms being identified [ ] . a less common mode of transmission is through transfusions. the symptoms of ehrlichia and anaplasma infections are similar [ ] . abrupt onset of an influenza-like illness occurs about days after a tick bite. ehrlichia can cause a mild rash ( % of adults and % of children), but rash is uncommon in anaplasma infections. highly suggestive laboratory features are leukopenia and thrombocytopenia. mortality in the general populations appears to be < %, but icu admission is not uncommon. hemophagocytosis has been described with anaplasma [ ] and ehrlichia [ ] . both infections are more severe in any setting of immune compromised, including asplenia [ , ] . the diagnosis is typically made by pcr, and doxycycline is the recommended treatment. intracellular inclusions can be seen on cbc smears (more often in anaplasma than ehrlichia). an uncommon but well described facet of these infections is that the tick vector can also transmit borrelia burgdorferi and babesia microti, and simultaneous infection with multiple organisms can occur. c. burnetii is a highly pleomorphic gram-negative coccobacillus and the causative agent of q fever. q fever is a zoonosis, and the most common reservoirs are cattle, sheep, and goats but many other animals can be infected by c. burnetii [ , ] . when infected, these domestic animals can shed the organism in urine, feces, milk, and especially birth products. the pathogen survives within the phagolysosome of host cells, and a spore stage has been described. this spore stage explains the ability of c. burnetii to survive in unfavorable environmental conditions, and it can be an environmental risk for months to years after shedding from an infected animal. q fever is considered an occupational disease affecting people with direct contact with infected animals; however, indirect contact through exposure to contaminated animal products has also been described to cause disease outbreaks. humans are infected by inhalation of contaminated aerosols. following an average incubation period of days, infected patients can present with severe headache, fever, chills, fatigue, and myalgia. other signs and symptoms depend on the organs involved. in contrast to rickettsial diseases described above, rash rarely occurs in the early stages of the disease. c. burnetii can cause a range of clinical symptoms. a self-limited febrile illness is probably the most common form of q fever. pneumonia, either atypical or severe, is also common and can be a part of acute q fever syndrome. in contrast, a variety of manifestations can be recognized in chronic q fever, including endocarditis, endovascular infection, osteomyelitis, hepatitis, interstitial pulmonary fibrosis, prolonged fever, and purpuric vasculitic rash. q fever diagnosis is based on serologic testing with indirect immunofluorescence being the best for differentiating between acute and chronic q fever (high antiphase i antigen titer). the treatment of choice for acute q fever is doxycycline, with co-trimoxazole, chloramphenicol, or rifampin being an accepted alternative. there is no agreement on the treatment for q fever endocarditis, and a combination of doxycycline with either fluoroquinolone or hydroxychloroquine is recommended. there is also controversy regarding the duration of treatment, ranging from years to indefinite treatment. old evidence suggests that q fever is more common in immunocompromised patients. a french study showed higher incidence of antibodies to c. burnetii in hiv positive compared to hiv-negative patients ( . vs . %). in addition, out of hospitalized patients were hiv positive ( . %), suggesting a more frequent symptomatic disease [ ] . a smaller similar study performed in central africa failed to show increased incidence of seropositivity in hivpositive patients [ ] . two case reports describe severe disease in immunocompromised patients. the first was a case of fatal q fever disease in an -year-old male with cgd [ ] . the patient was treated with broad spectrum antibiotics, but without coverage for q fever. the second case was a -yearold asplenic male who presented with fever, jaundice, and encephalopathy and was diagnosed with acute q fever [ ] . the patient was successfully treated, but the two case reports could suggest susceptibility in cases of phagocytic disorders. the bartonellaceae are fastidious, facultative intracellular gram-negative bacilli (table ). most species infect primarily non-human animals, and in most cases, human are considered incidental hosts. the documented common human pathogens include bartonella bacilliformis, bartonella henselae, and bartonella quintana, and it is believed that humans are the primary mammalian reservoir of b. quintana. infection occurs through inoculation of bartonella-infected arthropod feces into breaks in the skin. the epidemiology of bartonella infection in humans follows the distribution of the vector. as such, infection with b. bacilliformis follows the distribution of the sand fly vector (lutzomya) and is confined to the andes mountain in peru, ecuador, and colombia at heights of between and m. the human body louse pediculus humanus is the vector of b. quintana, which explains the global distribution of this pathogen and worldwide outbreaks of trench fever, mostly in conditions of poor sanitation and upon exposure to body lice. trench fever was responsible for over a million infections during world war i. fever, either abrupt or indolent in onset, with a maculopapular rash, conjunctivitis, headache, myalgias (most often affecting legs), and splenomegaly was described. urban b. quintana infections occur most often among homeless and have a distinct clinical picture with fever as the most common manifestation. endocarditis occurs in many [ ] . bartonella henselae is globally endemic, and domestic cats are a major reservoir. the major arthropod vector of b. henselae is the cat flea, which is responsible for cat-to-cat transmission. human infection, called cat scratch disease, is assumed to involve inoculation of bartonella-infected flea feces into the skin during a cat scratch. b. henselae causes primarily adenopathy and neurologic symptoms [ ] . b. bacilliformis causes a condition with two phases: the acute phase with fever, anemia, and transient immunosuppression followed by nodular dermal eruption [ ] . recently appreciated are the ongoing systemic features during the eruptive phase such as arthralgias, adenopathy, and anorexia [ ] . diagnosis of bartonella-associated diseases can be achieved by direct examination of clinical material, bacteriologic culture methods, serologic and immunocytochemical studies, pcr-based assay, or combination of these methods. bartonella infection can present differently in immunocompromised hosts [ ] . in addition to higher prevalence of bartonella infection in hiv patients [ ] , both b. quintana and b. henselae can induce neovascular proliferation which might involve the skin, lymph nodes, and a variety of internal organs including the liver, spleen, bone, brain, lung, bowels, etc. these neovascular lesions, known as bacillary angiomatosis/peliosis (ba/bp), were initially described in hiv-infected patients with advanced disease and was later described in other immunocompromised hosts secondary to immunosuppressant treatment for solid organ transplantation or hematologic malignancy [ ] [ ] [ ] [ ] . cutaneous ba lesions can vary in presentation and can be subcutaneous, dermal nodules, single or multiple papule that can be flesh colored, red, or purple. lesions may ulcerate and bleed. they can change in number and size (millimeters to centimeters; few to hundreds) and can involve mucosal surfaces or deeper soft tissues. similar variation can be seen with visceral involvement. histologically, lesions consist of lobular proliferation of small blood vessels and neutrophilic predominant cell infiltration. the term bacilliary peliosis is used to describe bloodfilled cystic spaces mostly involving the liver, spleen, and lymph node. pathogenic bacteria can be isolated from vascular lesions. while both pathogens were associated with cutaneous lesions, only b. henselae was associated with visceral bp [ ] . based on hiv literature, it is reasonable to expect an unusual presentation of bartonella infection especially in pidds involving t cell dysfunction. bartonella infection was described as a cause for hepatitis in a single cd l deficiency patient [ ] . in addition, since cases of granulomatous disease due to bartonella infection [ ] [ ] [ ] have been described, it should be considered in the differential diagnosis of pidd with granulomatous inflammation. borrelia spp. the genus borrelia belongs to the spirochaetaceae family. it includes b. burgdorferi which causes lyme disease and species that cause relapsing fever. the latter are further divided into tick-borne species and louse-borne species. louse-borne relapsing fever (lbrf) is caused only by borrelia recurrentis and is spread by human body louse. the disease was epidemic in the early twentieth century, and it is estimated that more than , died of lbrf during world war ii. with sanitation improvement lbrf is now found only in the horn of africa and among homeless people in europe. more recently, cases of lbrf in refugees and migrants were described [ , ] . tick-borne relapsing fever (tbrf) is caused by a group of pathogens which are maintained by and survive in softticks (orinthodoros genus). each tbrf borrelia species depends on one specific species of soft-body tick. except for australia and antarctica, tbrf species can be found in all continents. the animal reservoir includes small animals and rodents. since the spirochetes persist in the tick salivary gland, disease transmission occurs when humans intrude the tick's environment. tick bites are painless, and history of a tick bite is often missing. therefore, a history of potential exposure can be valuable. the major clinical symptom is relapsing fever. after a median incubation period of days, patients present with febrile episode that can last - days, followed by afebrile period of - days. patients with tbrf can have up to febrile relapses, while lbrf is usually associated with only one relapse. other symptoms include myalgia, arthralgia headaches, and vomiting, and physical findings include lymphadenopathy and splenomegaly with rash occurring only in third of the patients. a range of neurologic complications as well as systemic inflammatory response syndrome also have been described [ ] . diagnosis is based on identifying the spirochetes on blood smear. sensitivity of blood smear is higher during febrile period (about %), and a negative blood smear does not exclude rf. in lbrf, the spirochete load can be low and specific stains can be helpful. other diagnostic methods include serologic testing, pcr, and mouse inoculation. doxycycline, tetracycline, and penicillin are the preferred treatment, with most patients treated with - days of doxycycline. jarisch-herxheimer reactions with high fever and leukopenia occur in half of the patients following the first antibiotic dose and can develop into a severe reaction with hypotension, respiratory distress, and death [ ] . without treatment, tbrf carries a mortality rate of up to % with even higher % mortality rate for untreated lbrf. two cases of meningoencephalitis with borrelia miyamotoi in heavily treated immunocompromised patients have been described [ , ] . lyme borreliosis is the most common vector-borne disease in the northern hemisphere caused by a group of at least genospecies. lyme disease is a multisystem illness affecting the skin, joints, nervous system, and heart. human infection is caused mainly by three species: b. burgdorferi is the most common cause in north america but also found in europe, and borrelia afzelii and borrelia garinii which cause the disease in europe and asia. emerging infections in the mid-western usa with borrelia mayonii cause a condition similar to lyme disease. most tick species do not carry borrelia species. the vectors of all borrelia species are the ixodid tick species; this includes the deer tick, i. scapularis, in the northeast and midwest of the usa, ixodes pacificus in the west, the sheep tick, ixodes ricinus, in europe and the taiga tick, ixodes persulcatus, in asia. the ixodid tick demonstrates a complex vector ecology with preferences for different hosts in different geographical regions and at different stages of its development. more than different species, including deer, rodents, birds, and reptiles, have been described. infection rates also show seasonal variation with highest rates during lyme disease follows several stages starting with localized disease at the site of inoculation, followed by dissemination stage and, later, persistent infection [ ] . however, an individual patient can show highly variable disease progression with different patterns of organ involvement and disease severity. erythema migrans (em) is often seen at the site of the tick bite after - days of incubation. regional lymphadenopathy can be seen. secondary skin lesions represent hematogenous dissemination. at this stage, constitutional symptoms of general fatigue, fever and headaches, migratory musculoskeletal pain, conjunctivitis, and cardiac involvement occur. in total, % of untreated patients can develop frank neurologic manifestations of meningitis, encephalitis, and variable forms of neuritis with fluctuating symptoms. persistence can occur in untreated (on inadequately) patients. antibiotic refractory arthritis is well described. however, even without treatment, intermittent or persistent attacks usually resolve completely within several years. co-infection with a. phagocytophilum and b. microti can cause diagnostic confusion [ , ] . the diagnosis of lyme disease is established based on clinical symptoms, history of potential exposure, and serologic studies. although positive culture can confirm the diagnosis, it can usually be obtained only from early em lesions. pcr testing is superior to cultures and can be performed on joint fluid samples [ ] . cdc recommendations for the diagnosis of lyme disease are based on serology which might be impossible in pidd patients with abnormal humoral response. cdc guidelines require both an elisa (or comparable test) to be positive and a western blot ( out of bands ( , , or kd) on the igm or out of bands on the igg ( , , , , , , , , , kd) . most lyme manifestations can be treated with oral antibiotics, while patients with neurologic abnormalities and some patient with lyme arthritis require intravenous therapy [ ] . doxycycline is the treatment of choice for early and disseminated disease, with amoxicillin as the second-line choice. jarisch-herxheimer-like reactions can appear in the first h in about % of the patients. for patients with clear neurologic symptoms, - weeks of iv ceftriaxone is the most commonly used therapy. few cases of neuroborreliosis and hiv have been described with a good response to treatment. descriptions of lyme disease in pidd patients are lacking. zoonoses are infectious diseases that pass between animals and humans and span the spectrum of pathogens including viruses, bacteria, fungi, and parasites. zoonoses are very common and range from mild such as certain forms of tinea to lifethreatening infections such as rabies. some of the zoonoses that are vector-borne will be covered in other sections. risk mitigation strategies for zoonoses include patient education, proactive advice about risk scenarios, and avoidance of infected animals. several zoonoses are associated with contact with mammals such as rodents or domestic farm animals through direct contact or through contact with their feces. for instance, hantavirus infections are often associated with exposures to mouse droppings when staying in cabins in the western usa. occupational exposures can occur with buffalopox or parapoxvirus (causing orf infection) through direct contact with buffalo and goats/sheep, respectively [ ] [ ] [ ] [ ] . in general, there are very few cases of pidd with zoonotic infections acquired from mammals. however, there are a few special considerations. for instance, lymphocytic choriomeningitis virus is acquired through exposure to house mice primarily, with hamsters being a less common source of infection. both domestic and wild mice can carry the infection without exhibiting symptoms. although infection is rare, there have been severe cases in patients with t/ nk cell dysfunction, such as a case in xlp and cases in solid organ transplant recipients [ ] . therefore, in patients with severe t/nk defects, consideration should be given to whether small rodents are appropriate household pets. tularemia is a disease of animals and humans caused by the bacterium francisella tularensis. rabbits, hares, and rodents are the main reservoirs. humans become infected through direct contact, ingestion of contaminated water, or inhalation of organisms. ticks and deer flies can also transmit the disease through bites. fever is universal, but other features depend on the mode of transmission. a patient with cgd had a complex course suggesting myeloid defects are a risk for more severe disease [ ] . rabies is an almost universally fatal infection caused by contact with oral secretions from infected mammals, typically raccoons, bats, or foxes, and there is no suggestion that pidd or immune compromised modifies the prognosis. for individuals with high-risk exposures, such as those working with wildlife or traveling in endemic areas, pre-exposure prophylaxis is given with vaccination, and if an exposure occurs, rabiesspecific immunoglobulin is provided as well as vaccination. however, for those with humoral immunodeficiencies who cannot respond to the typical pre-exposure vaccination, there needs to be counsel on the additional risk without vaccination. in table , several of the bacterial and viral zoonoses are summarized with their typical endemic areas, which is somewhat limited by diagnostic abilities and reporting, as well as the typical clinical scenarios, known cases in immunodeficiency and an approach to diagnosis and therapy. nipah virus causes a range of infectious phenotypes ranging from asymptomatic infection to acute respiratory distress and encephalitis. nipah virus was identified in on pig farms in malaysia, leading to identification of human cases, including human deaths and the culling of one million pigs [ ] . the natural host is the fruit bat: pteropodidae pteropus. symptoms of infection from the malaysian outbreak were primarily encephalitic in humans, but later, outbreaks have caused respiratory illness, increasing the likelihood of human-to-human transmission. fever, headache, cough, abdominal pain, nausea, vomiting, weakness, problems with swallowing, and blurred vision were common. seizures were seen in % and coma in %. relapses of encephalitis have been described [ ] . increasing infections due to nipah virus is thought to be due to an increasing overlap between bat habitats and pig sties in malaysia. all outbreaks thus far have been in india, bangladesh, or malaysia. the diagnosis of nipah virus relies on pcr of fluid samples, serology in convalescent samples, and immunofluorescence of tissue. there have been no infections of immune compromised patients reported. therapy is largely supportive, although preliminary reports of ribavirin use have been encouraging. a vaccine is under development. severe acute respiratory syndrome coronavirus (sars-cov) and the middle east respiratory syndrome coronavirus (mers-cov) are two zoonotic coronaviruses. the sars pandemic in - resulted in reported cases in countries. no further sars cases were reported after the pandemic except isolated cases linked to laboratory accidents. patients usually presented with fever and respiratory symptoms, but occasionally had diarrhea and vomiting. about - % of sars patients required mechanical ventilation, with a case fatality rate of about % [ ] [ ] [ ] . mers was first noted in saudi arabia in , and countries around the arabian peninsula are now endemic for mers-cov. patients usually present with fever, cough, chills, sore throat, myalgia, and arthralgia rapidly progressing to pneumonia with over % of patients requiring intensive care. about one-third of patients present with diarrhea and vomiting, and acute renal impairment is a striking feature of mers. risk factors for poor outcome include diabetes, hypertension, and renal and lung disease. cases have been exported to at least countries with travel occasionally causing cluster of secondary outbreaks. one such example is the mers-cov outbreak involving patients in south korea, and the median incubation period was estimated to be days with a range of to days [ ] . at the end of , there were confirmed mers with a % mortality rate [ ] [ ] [ ] . bats are the natural reservoirs of both sars-cov and mers-cov. sars-cov crossed the species barrier into palm civets and other animals in live animal markets in china, which then infected human, while a mers-cov ancestral virus crossed species barrier into dromedary camels. abundant circulation of mers-cov in camels results in continuous zoonotic transmission of this virus to human, while sars-cov was not found to circulate in the intermediate reservoirs, explaining sars being a one-off outbreak and mers a continuing zoonotic disease [ ] . aerosolgenerating procedures such as intubation were associated with increased viral transmission of both covs resulting in nosocomial outbreaks [ ] . super-spreaders are responsible for large and prolonged outbreaks [ ] . the diagnosis for sars and mers include both serological tests and pcr assays that can quantify viral loads [ ] . functional genetic polymorphisms leading to low serum mannose binding lectin (mbl) are associated with susceptibility to but not severity of sars in both southern and northern chinese [ ] [ ] [ ] . mbl was shown to bind to sars-cov and inhibit the infectivity [ ] , suggesting its role as first-line defense against sars-cov. although no patients with primary immunodeficiency infected with sars-cov or mers-cov were identified, likely due to the limited number of such infections, patients with t cell defect and type interferon pathway defects could suffer a more severe disease course [ , ] . virus-based and host-based treatment strategies are largely experimental with uncertain benefits. ribavirin, type interferons, small molecules, and monoclonal antibodies that block covs entry have been explored [ ] . passive immunotherapy and multiple candidate vaccines have been tested in various animal models. convalescent plasma immunotherapy has been considered, but clinical trials are lacking in mers [ ] , while for sars a systematic review concluded convalescent serum may reduce mortality and appear safe [ ] . the filoviridae family contains three known genera, the ebolaviruses, marburgviruses, and cuevavirus. ebolavirus and marburgvirus cause hemorrhagic fever syndromes in primates and humans, with high fatality rates. cuevavirus infects only bats. the ebolavirus genus contains five species, with two of the species (zaire ebolavirus and sudan ebolavirus) being responsible for the majority of cases of human disease, while marburgviruses contain two species (marburg virus and ravn virus). filoviruses are capable of replicating in a number of cell types (with the exception of neurons and lymphocytes). upon entry into the body of the host (via breaks in the skin, parenterally, or through mucosal surfaces), filoviruses employ a variety of mechanisms to evade the activity of the immune system [ ] . the incubation period (interval from infection to onset of symptoms) varies from to days. symptoms begin abruptly, with high fever, severe headache, malaise, myalgia, diarrhea, nausea, and vomiting. a rash can occur between and days after onset of symptoms. hemorrhagic manifestations occur between and days, and fatal cases usually have some form of active bleeding. in an outbreak setting, the symptoms are unmistakable but confusion with malaria can occur early in the disease or in sporadic cases. since their original descriptions in and , respectively, for marburg and ebola, there have been a number of sporadic cases and several major outbreaks. the largest marburg virus outbreak occurred in angola in (with a fatality rate of > %), while the largest ebola epidemic happened between and in west africa (sierra leone, guinea, and liberia) and claiming over , lives (fatality rate > %). although not definitively proven in the case of ebola, bats are believed to be the natural animal reservoir for these viruses [ , ] . these viruses are transmitted via contact with blood or body fluids from an infected host; notably, certain body fluids can harbor virus for weeks to months after resolution of disease. given the recent outbreak in west africa, there has been renewed interest in understanding the pathogenesis of filovirus infections and possible therapies. literature regarding how the pathogenesis of disease may be altered in patients with pidd is lacking. however, the assumption is that in the absence of an intact cellular and/or humoral immune response, the patient with a pidd may be at increased risk of mortality in the setting where mortality is already high. these viruses induce apoptosis of lymphocytes and macrophages, and there is therefore a profound secondary immune compromised [ , ] . filoviruses can be detected in multiple body fluids via pcr. although practiced for decades, a study in guinea in failed to show a decrease in mortality among patients receiving convalescent plasma from previously infected donors [ ] . a number of additional compounds (e.g., tkm-ebola, bcx , and gs- ) and biologics (zmapp) have been shown to offer protection in animal models of ebola, but to date, no controlled and appropriately powered clinical trials have addressed their efficacy in humans. finally, a number of vaccines for ebola are undergoing clinical studies (including four in phase iii trials). importantly, in late , the rvsv-zebov vaccine was shown to have displayed high efficacy in protecting immunized adults during the guinea ebola outbreak, and the data also suggested that the vaccine may even offer bherd immunity^to unimmunized persons in proximity to recipients of the vaccine [ , ] . hepatitis e virus is a single-strand rna virus of the hepeviridae family. it is an important zoonotic disease in asia and africa, and fecal-oral spread is the usual route of transmission [ ] . handling of pig or boar meat is a risk factor, and - % of pig livers sold in grocery stores in japan and the usa are infected [ , ] . swine represent the major reservoir, although antibodies to the virus have been found in many species [ ] . the incubation period is weeks to months, and viremia disappears with the onset of symptoms. the mortality rate is - % and can reach % in pregnancy [ ] . acute hepatitis usually resolves but can lead to liver failure in severe cases. patients with hepatitis e posttransplant have had severe courses in some cases [ ] . in immune compromised patients, the course can become chronic [ ] [ ] [ ] . in these cases, cirrhosis develops. the diagnosis is by serology or pcr, and the treatment is supportive. prevention modalities for infections transmitted by humans are conceptually different than infection prevention for vector-borne infections. hand hygiene is extremely important, and avoidance of clearly infected people can be helpful. recognition of infections with fecal-oral transmission and the importance of water purity are critical for patients with pidd. in contrast, infections transmitted by aerosols require prevention strategies related to droplet precautions. in outbreak scenarios, if the risk to the patient is high, specific chemoprophylaxis may be considered. influenza viruses type a and b cause annual epidemic influenza, while type c causes sporadic mild influenza-like illness. patients present with sudden onset of fever, chills, and myalgia, followed by sore throat and cough. other less common features include diarrhea, acute myositis, and encephalopathy [ , ] . co-infection with bacteria such as pneumococci results in more severe disease [ , ] . influenza pandemics occur yearly around the world. influenza viruses infect to % of the global population, resulting in~ , deaths annually [ ] . the viruses circulate in asia continuously and seed the temperate zones, beginning with oceania, north america, and europe, then later seeding into south america [ ] . diagnosis of influenza includes direct/ indirect immunofluorescent antibody staining for antigens in nasopharyngeal aspirates and pcr. a patient with compound heterozygous null mutations of the gene encoding irf , a transcription factor for amplifying ifn-α/β, was reported to have life-threatening influenza during primary infection [ ] . fatal influenza-associated encephalopathy in both chinese and japanese children has been reported to be associated with genetic variants of thermolabile carnitine palmitoyltransferase ii [ ] . patients with scid will have prolonged viral shedding [ ] . severe pandemic influenza a virus (h n ) infection has been associated with igg and igg subclass deficiency [ , ] . in addition, influenza infection can be more severe in pidd patients with underlying lung disease, such as bronchiectasis, and antibiotic coverage of chronic colonizing bacteria (such as pseudomonas) in this setting may be helpful. inactivated seasonal influenza vaccine should be given to pidd patients even those with humoral deficiencies as their t cell response to influenza could be normal and offer protective immunity against severe influenza [ , ] . antiviral drugs include neuraminidase inhibitors (oseltamivir and zanamir) and adamantanes (amantadine and rimantadine), but resistance to adamantanes is widespread. measles is a single-stranded, negative-sense, enveloped (nonsegmented) rna virus of the genus morbillivirus. measles is highly communicable, transmitted by droplets, and less commonly by airborne spread. patients present with fever, cough, coryza conjunctivitis rash, and koplik spots. complications include pneumonia, acute encephalitis, and subacute sclerosing panencephalitis (sspe) [ ] . diagnosis of measles includes serological tests, virus isolation, and pcr. in an outbreak, the clinical features may be sufficient for diagnosis. measles vaccine has caused severe measles in children with stat and ifn-α/β receptor deficiency [ , ] , demonstrating the importance of type interferon pathway in controlling measles. immune compromised of nearly any type is associated with severe disease and higher mortality [ ] . t cell deficiency states are the most strongly associated with the development of giant cell pneumonia and inclusion body encephalitis, the most feared complications of measles. sspe is a slow encephalitis due to persistence of replication defective measles virus in the cns. it is most frequently seen when young infants are infected with measles and - years later, sspe becomes evident. there are case reports supporting the immune compromised as increasing the risk of sspe [ ] . treatment of sspe with ribavirin has shown some improvement, but the prognosis in general with sspe is very grave. patients with cgd have defective memory b cell compartment, resulting in lower measle-specific antibody levels and antibody-secreting cell numbers, but severe disease has not been reported [ ] . pidd patients may harbor the virus latently for longer than usual, leading to complications at the time of transplant [ ] . specific antiviral therapy is lacking, but ribavirin has been given to severely ill and immunocompromised children. for measles post-exposure prophylaxis, intravenous immunoglobulin (ivig) is recommended for severely immunocompromised patients without evidence of measles immunity [ ] . this would likely include patients with scid and hypogammaglobulinemia who are not yet on regular ivig. measles vaccine, given in a two-dose regimen, has brought down incidence enormously worldwide and the who is planning for eradication globally. enteroviruses (evs) are among the most common viruses infecting humans worldwide. evs are small non-enveloped, single-stranded rna viruses of the picornaviridae family. human evs are categorized into seven species that include hundreds of serotypes, such as polioviruses (pv), coxsackie viruses a, and b (cv-a and b), echoviruses, and human rhinoviruses (hrvs). of these species, many important serotypes are known to infect human such as pv - , cv-a , cv-b , ev-a , ev-d , and hrv (table ) . non-polio enteroviruses (npevs) have a worldwide distribution. infants and young children have higher incidence of infection and a more severe course of illness than adults. the mode of transmission is mainly through fecal-oral and respiratory routes. infection occurs all around the year in tropical and subtropical regions, while in temperate climates the peak incidence of infection is during summer and fall months [ ] . npevs are associated with diverse clinical manifestations ranging from mild febrile illness to severe, potentially fatal conditions. most cases are asymptomatic or have mild symptoms including fever with or without rash; symptoms of hand, foot, and mouth disease; herpangina; acute hemorrhagic conjunctivitis; upper respiratory infection; and gastroenteritis. more severe symptoms occur in infants and young children [ , ] . acute flaccid paralysis [ ] , neonatal enteroviral sepsis [ ] , myocarditis/pericarditis [ , ] , hepatitis, pancreatitis, pneumonia, and atypical hemolytic uremic syndrome [ ] are severe manifestations seen in immunocompetent people. chronic infections have been seen in immunocompromised patients [ ] . each virus may produce one or more of the aforementioned manifestations; however, some serotypes are often associated with particular features ( table ) . the definitive diagnosis of npev infection relies on pcr or virus isolation from the cerebrospinal fluid, blood, stools, urine, or throat swab [ , ] . treatment of npevs is mainly supportive since most infections are self-limited. high doses of intravenous immunoglobulin (ivig) are recommended in patients with severe symptoms. the efficacy of some new antiviral drugs (pleconaril, vapendavir, and pocapavir) is still under investigation [ ] . no vaccine has been licensed yet for npevs. however, phase clinical trials of inactivated monovalent ev-a vaccines manufactured in china showed high efficacy against ev-a in infants and young children [ ] . patients with a variety of pidds are unusually susceptible to ev [ ] . the most susceptible groups are patients with primary antibody deficiency such as xla, cvid, and hyper-igm syndrome (higms) as well as those having scid and major histocompatibility class ii deficiency [ , ] . the most severe form of infection has been described in patients with xla due to the profound deficiency of immunoglobulins essential for viral neutralization during infection. affected patients usually present with indolent but relentlessly progressive non-necrotizing meningoencephalitis. regression of cognitive skills, flaccid quadriplegia, and deafness has been described. the reported non-neurologic presentations in xla include septicemia, dermatomyositis-like disease, hepatitis, and/or arthritis [ , ] . the incidence of npev meningoencephalitis in large registries of xla cases is - % [ ] . unpublished data from the kuwait national pidd registry, which includes pidd patients, showed that nine patients had documented npev infections and two died from these infections. the two deaths were seen in scid patients (personal communication with prof. waleed al-herz, md). in addition, npev meningoencephalitis and/or septicemia were reported in few cases with either primary b cell deficiency such as b cell linker (blink) protein deficiency [ ] or acquired b cell deficiency following the administration of anti-cd (rituximab) [ , ] . in all reports, better outcome was attributed to the early administration of high doses of ivig during npev viremia [ ] . npev infection in pidd diseases remains a major threat to patients. also, the possible prolonged viral excretion and the emergence of resistant strains runs the risk of spreading infection to the surrounding community. oral polio vaccine (opv) consists of a mixture of three live attenuated poliovirus serotypes. opv induces production of neutralizing antibodies against all three serotypes, in addition to a local intestinal immune response. opv can result in vaccine-associated paralysis (vap) secondary to reversion of the vaccine strain to the neurovirulent wild-type strain. an example for such an event was demonstrated by the - outbreak in the dominican republic and haiti [ ] , believed to be driven at least in part by undervaccination of the population, which allowed the spread of the reverted vaccine strain [ ] . although rare, patients with pidd have a higher risk to develop vap. reports have shown that pidd patients with antibody deficiency can have prolonged viral replication which can persist for years and therefore theoretically increase the risk for a spontaneous reversion within the immunodeficient host [ ] [ ] [ ] [ ] . cases of vap were shown in patients with antibody deficiency and combined immunodeficiency syndromes [ , , ] . therefore, opv is contraindicated in patients with pidd, and this contraindication extends to their household contacts [ ] . beyond the obvious risk for the pidd patient, prolonged virus shedding also increase the risk for spreading vaccine-derived paralytic strain in the general population. bacterial infections have molded human behavior and altered societies over human history. today, largely ignored due to antibiotic susceptibility, they continue to cause misery and disease around the world. three infections are highlighted, and additional commonly encountered infections are listed in table . pertussis is a respiratory infection caused by bordetella pertussis that begins after a -to -day incubation period as a minor upper respiratory infection that progresses with cough. initially intermittent, it evolves into paroxysmal coughing spells usually followed by vomiting in infants and young children. it lasts to weeks and can have many complications such as syncope, weight loss, rib fracture, and pneumonia. infants under months are more severely affected, developing pneumonia, pulmonary hypertension, hypoxia, subdural bleeding, and seizures. death can occur, especially in young infants [ , ] . adults typically have a prolonged cough with fewer complications [ ] . it is transmitted via aerosolized droplets during close contact. people are most contagious during the catarrhal stage and the first half of the paroxysmal phase, totaling to weeks [ ] . the introduction of whole-cell pertussis vaccine (dpt) in the s in the usa reduced the incidence of the disease from , cases to around cases per year in the s. a resurgence in was associated with the substitution of the whole-cell vaccine by the acellular pertussis vaccine (dtap) [ ] . new strategies such as boosters with acellular pertussis for adolescents and adults with tdap and use of tdap during pregnancy seem to be effective in partially reducing the incidence of the disease [ ] ; however, pertussis cases in the usa remain higher than the s. the lack of persistence of antibody in the adult population means adults not only represent a reservoir for the disease but also do not provide sufficient titers to immunoglobulin products prepared from adult plasma pools. a relatively recent requirement in some countries is vaccination of adults every years to maintain immunity. this should, over time, improve titers in immunoglobulin products. culture of specimens obtained by nasopharyngeal swabs is the gold standard of laboratory diagnosis due to the % specificity, but polymerase chain reaction (pcr) is gaining prominence due to its higher sensitivity and speed of results; serodiagnosis can be used in the late stages of the disease [ ] . filamentous hemagglutinin (fha) and pertussis toxin (pt) antibodies were detected at peak measurements in pidd patients on regular ivig, although some of them had pt antibodies below the protective level as trough measurements [ ] . severe pertussis cases have not been reported in pidd patients, but severe disease has been seen in malignancies [ ] . antimicrobials such as azithromycin, erythromycin, and clarithromycin, if given during the catarrhal stage, may ameliorate the disease and shorten the contagious period. to avoid cases of pertussis, it is also worth emphasizing the importance of good vaccine coverage rate among the whole population, but especially among healthcare workers and family members of patients with pidd. neisseria meningitidis the onset of neisserial meningitis is associated with sore throat, headache, drowsiness, fever, irritability, and neck stiffness [ , ] . purpuric lesions are very characteristic. this pathogen can also present with sepsis which has a % mortality rate as opposed to % mortality with a meningitic presentation. this bacterium can also cause a chronic condition referred to as chronic meningococcemia. this condition is characterized by intermittent fevers lasting week to - months [ ] . a non-purpuric rash is common which may evolve into purpura. arthritis, similar to that seen with gonococcus, is common. meningococcal disease primarily affects children under years of age. n. meningitidis is a global pathogen [ ] . there are serogroups, but the majority of invasive meningococcal infections are caused by organisms from the a, b, c, x, y, or w serogroups. the annual number of invasive disease cases worldwide is estimated to be at least . million, with , deaths related to invasive meningococcal disease. serogroups b and c are responsible for most infections in europe. serogroup a has historically been the major organism in africa; mass vaccination has led to some improvement, but the emergence of group x disease is worrisome. the hajj in the middle east has seen epidemics of w- , and vaccination is now required for hajj travelers. b and c serogroups are the most common through the americas. n. meningitidis cases occur at a rate of about case per , people throughout the world [ ] , but across the sahel of africa and in china, epidemics can lead to case rates of per , [ ] . the bacterium is a natural human commensal, with carriage rates of about %. diagnosis can be by clinical examination in epidemics or by gram stain and culture. complement-deficient individuals have an increased risk of neisserial disease, but not necessarily increased mortality. hiv is associated with increased disease, suggesting that t cells are also important for defense. thirdgeneration cephalosporins are typically used for treatment. penicillin, ampicillin, aztreonam, and chloramphenicol are alternatives. there is great inter-individual variability in the development of tb disease. roughly, % of infected individuals develop clinical disease within years of infection (mostly during childhood). about % became latently infected without clinical disease, and the remaining to % develop pulmonary tb later in life, either from reactivation of latent infection or reinfection. molecular epidemiology studies from high burden areas suggest more disease results from recent transmission than from reactivation of latent tb, particularly in people living with hiv [ ] . acquired or inherited host factors may at least partially account for the variable disease course, resulting in increased susceptibility to mycobacteria infections [ ] . pidd associated with tb and ntm infections include t cell deficiencies, gata deficiency, cgd, anhidrotic ectodermal dysplasia with immunodeficiency, x-linked (xl) recessive cd ligand deficiency, autosomal recessive (ar) stat deficiency, ar irf deficiency, and ar tyk deficiency. in addition, a group of disorders with a strong susceptibility to ntm, named mendelian susceptibility to mycobacterial diseases (msmd), have been recognized since the s. these are rare inborn errors of ifn-γ signaling pathway that present with isolated predisposition to infections caused by weakly virulent mycobacteria such as bcg vaccine and environmental ntm, in otherwise healthy patients. the genetic defects involve impairment in the production of interferons (ar il rβ , ar il p , autosomal dominant (ad) irf , ar isg , xl recessive nemo) or response to interferons (ifn-γr, ad stat , ad irf , cgd) [ ] . an acquired form exits: adults with ntm infection in thailand and taiwan were found to have high-titer anti-interferongamma antibody [ ] . these individuals from southeast asia were found to have hla-drb * / : and dqb * : / : . patients suspected of having pulmonary tb should have acid-fast bacilli (afb) smear microscopy and culture performed in three sputum samples. pcr for mtb can be performed [ ] . the use of rapid tests facilitates early diagnosis, and the who has recently recommended their use. the only recommended rapid test for detection of tb with and without rifampicin resistance is the xpert mtb/rif assay (cepheid, sunnyvale, ca). the who recommends the xpert test for those suspected of having drug-resistant tb or in hiv; however, culture is still the mainstay and is not replaced by the xpert test. tb skin testing (mantoux testing) uses a purified protein derivative injected under the skin. its advantages are that it can be used for large-scale screening and it is cost effective. skin testing does have several disadvantages when used as a diagnostic test. reading the induration requires training and immunodeficiencies can alter the magnitude of the induration. immunosuppressed patients (hiv, organ transplant) are considered positive if the induration is ≥ mm. some immunodeficiencies may completely ablate the response. other causes of false-negative tests are malnutrition, concurrent infection, recent live viral vaccine administration, renal failure, malignancy, medical stress, very elderly, young infants, or with a very recent infection with mtb. conversely, the results may be falsely positive if bcg has been administered. interferon gamma release assays can be used in any setting where skin testing would be done but are considered superior in settings where the patient has had bcg vaccination and, in some cases, where skin testing has been sown to have high false-negative rates. in general, tb treatment for patients with impaired immune response, including pidd, hiv infection, and immunosuppressive therapy, is based on the standard -month regimen consisting of a -month intensive phase of isoniazid, rifampin, pyrazinamide, and ethambutol, followed by months of isoniazid and rifampin. decisions regarding treatment duration can be individualized, taking into account disease severity, organs involved, and response to treatment. significant pharmacological interaction can occur between rifampin-based mtb regimens and immunosuppressive drugs, such as calcineurin inhibitors or rapamycin, requiring strict monitoring of drug plasma concentrations [ ] . therapy for ntm is complex with highly variable drug resistance patterns and a need for biological augmentation to effectively clear the organism. aspergillus fumigatus (see above), cryptococcus gattii, histoplasma capsulatum, coccidioides immitis (or c. posadasii), blastomyces dermatitidis, paracoccidioides brasiliensis, emmonsia pasteuriana, and penicillium (talaromyces) marneffei are environmental fungi that are endemic in certain parts of the world (table ). with the exception of penicillium marneffei and emmonsia pasteuriana that only cause disease in profoundly immune compromised individuals, these fungi can cause infection in healthy individuals, ranging from mild, self-limited pulmonary disease to infection that requires antifungal therapy for eradication. on the other hand, patients with acquired defects in cell-mediated immunity such as those infected with hiv, and patients with specific monogenic disorders, particularly those involving the il- / ifn-γ/stat signaling pathways, scid, and gata depends on underlying state of immunosuppression and magnitude of environmental exposure icl idiopathic cd lymphocytopenia, aids acquired immune deficiency syndrome, stat signal transducer and activator of transcription , gata gata binding protein , scid severe combined immunodeficiency, cvid common variable immune deficiency, pidd primary immunodeficiency, il rb , interleukin- receptor subunit beta , ifngr gamma interferon receptor deficiency, are at high risk of developing life-threatening disseminated infections by these endemic fungi following environmental exposure [ , [ ] [ ] [ ] [ ] [ ] . diagnosis relies on culture of the corresponding fungus, histopathological demonstration of the fungus-specific characteristic morphologies, and/or surrogate serological and fungal antigen tests. treatment of clinical disease (as opposed to colonization) typically involves an initial induction phase with amphotericin b, followed by long-term azole maintenance therapy and secondary prophylaxis, and prognosis varies significantly depending on the fungal pathogen and underlying pidd. melioidosis is caused by b. pseudomallei, a gram-negative bacteria found in soil and water, in tropical climates of southeast asia and northern australia [ , ] . melioidosis is an emerging, potentially fatal disease ( % mortality). b. pseudomallei can be transmitted by inhalation, ingestion, or direct contact (through open skin) with contaminated soil or water. animals (sheep, goats, swine, horses, cats, dogs, and cattle) are also susceptible to infection and cases of zoonotic transmission through direct contact of skin lesions with infected animal meat or milk have been described [ ] . b. pseudomallei infections are endemic in northern australia and southeast asia. approximately % of reported infections occur during the rainy seasons. cases have also been reported in south pacific, africa, india, and the middle east. in temperate areas, infection is extremely rare and is predominantly imported by travellers or immigrants [ ] . the incubation period of melioidosis is variable from day to years, although common symptoms develop between and weeks after exposure. melioidosis presents most frequently in adults - years of age, but can occur in all ages, with one study reporting % of cases occurred in children [ ] . in australia, the average annual incidence in - was reported as . cases per , people [ ] . the incidence in indigenous australians was higher at . cases per , . a case-cluster in an australian community was associated with post-cyclonic flooding. a recent review suggests that b. pseudomallei is increasingly prevalent in the americas, with a mortality rate of % [ ] . infection in healthy individuals is uncommon, and more than % of cases occur in the setting of underlying conditions such as chronic renal disease, diabetes, chronic lung disease, and alcoholism. a recent review of melioidosis in travelers found that % of cases were acquired in thailand. symptoms usually started at days (range - days) after leaving the endemic area. traveller infections were less often associated with predisposing risk factors ( . %), diabetes mellitus being the most common ( %). melioidosis in travelers had lower mortality ( %) than infection in autochthonous cases in southeast asia [ ] . pneumonia (~ - %) is the most common presentation in adults. there is usually high fever, headache, anorexia, and myalgia. b. pseudomallei infection may also present as localized skin infection, septicemia, or disseminated infection. localized infection results in an ulcer, nodule, or skin abscess. this usually occurs from the bacterium breaching through a break in the skin. patients with renal disease and diabetes are more susceptible to sepsis. in disseminated infection, abscesses may develop in the liver, spleen, lung, and prostate. in children, primary cutaneous melioidosis is the commonest presentation ( %). bacteremia is less common in children than in adults, but brainstem encephalitis has been reported [ ] . difficulties in laboratory diagnosis of melioidosis may delay treatment and affect disease outcomes [ ] . diagnosis of melioidosis is primarily by isolation of the organism. identification of b. pseudomallei can be difficult in clinical microbiology laboratories, especially in non-endemic areas where clinical suspicion is low. although various serological tests have been developed, they are generally unstandardized bin house^assays with low sensitivities and specificities. pcr assays have been applied to clinical and environmental specimens but are not widely available and sensitivity remains to be evaluated. cases of melioidosis have been reported in patients with acquired or inherited immune deficiency. melioidosis was the presenting complaint in several patients with cgd. bacteremic melioidosis was recently reported in two patients with prolonged neutropenia, who succumbed despite appropriate antibiotics [ ] . it is likely that there is increased susceptibility in situations where innate or adaptive immunity is compromised. treatment is with intravenous antimicrobial therapy for - days, followed by - months of oral antimicrobial therapy. intravenous therapy with ceftazidime or meropenem is usually effective. oral therapy may continue with trimethoprim-sulfamethoxazole or doxycycline. free-living amoebas (fla) are protozoa found worldwide that do not require hosts to survive. fla do not employ vectors for transmission and are not well adapted to parasitism in humans. however, there are four genera/species that can cause human disease: naegleria (n. fowleri), acanthamoeba (multiple species), balamuthia (b. mandrillaris), and sappinia (s. pedata). all of these amoebae are capable of inducing cns disease in humans, but acanthamoeba species also cause various extra-cns infections, especially in immunocompromised hosts. the fla that are pathogenic in humans are reviewed below. naegleria are a diverse group of fla flagellate protozoans with a large number of distinct species. only one species, n. fowleri, has been shown to cause infection in humans. n. fowleri has a multi-stage life cycle with amoeboid and trophozoite-infective forms as well as a cyst form [ ] . n. fowleri is found commonly in warm freshwater around the world including lakes, rivers, and hot springs. humans become can become infected when swimming or diving in contaminated water. in rare circumstances, infections have also been attributed to exposure from contaminated tap water sources when utilized for religious cleansing of the nose or irrigation of the sinuses. thus, tap water should not be used for nasal and sinus irrigation. it is not possible to become infected from drinking contaminated water or from contact with an infected person, and the amoeba is not found in salt water. after entry to the nasal cavity, the amoeba travels through the cribiform plate to the olfactory bulbs and migrates to the cerebellum, resulting in primary amoebic meningoencephalitis (pam), a rapidly fatal brain infection characterized by the destruction of brain tissue. in its initial presentation, pam can mimic bacterial meningitis, further delaying accurate diagnosis and initiation of therapies that may save the patient. overall, n. fowleri infections are rare. worldwide, most cases are reported in the usa, australia, and europe; however, in developing countries, it is suspected that a large number of cases go unreported. between and , there were only infections reported in the usa with of the cases attributed to contaminated recreational water, infections following nasal irrigation with contaminated tap water, and case where a person was infected following use of a backyard slipn-slide utilizing contaminated tap water [ ] . the fatality rate associated with n. fowleri infection is over %, and between and , only of the infected persons in the usa have survived infection. initial symptoms of pam start to days after infection and can include headache, fever, nausea, or vomiting [ ] . progressive symptoms can include stiff neck, confusion, lack of attention, loss of balance, seizures, and hallucinations. cardiac arrhythmias have also been observed. the infection progresses rapidly after initial onset and causes death within to days after exposure (mean of . days). since infection often progresses rapidly to death, there is often insufficient time to mount a robust immune response. however, both the innate (neutrophils, macrophages, and complement system) and the adaptive (both t and b cells) arms of the immune system have been shown to participate in the immune response to n. fowleri [ ] . patients with pam have csf with elevated pressure that is often cloudy or hemorrhagic, with neutrophil-predominant pleiocytosis, elevated protein levels, and very low glucose. wet mounts from centrifuged csf will show motile mono-nucleated trophozoites measuring~ - μm in size. additionally, trophozoites can be identified with giemsa and wright stains of csf smears combined with an enflagellation test [ ] . confirmation can be achieved via a variety of timeconsuming methods including an immunofluorescence assay [ ] , culture of csf [ ] , or pcr-based methods [ ] . the optimal therapy for n. fowleri pam is still debated. the use of intravenous amphotericin b and fluconazole followed by oral administration of rifampin resulted in survival of a year-old child with pam [ ] . another child was shown to survive following intravenous and intrathecal amphotericin b and miconazole as well as oral rifampin [ ] . most recently, an adolescent girl was successfully treated with the combination of azithromycin, rifampin, fluconazole, and miltefosine [ ] . prevention is critical for this highly fatal infection and warning pidd patients not to use tap water for nasal irrigation is important. since its original description in , over cases of b. mandrillaris infections have been described worldwidewith most cases occurring in south america and the usa. balamuthia are found in soil, and acquisition of disease has been associated with agricultural activities, dirt-biking, gardening, and swimming in contaminated water sources. b. mandrillaris is thought to enter the body of the host through breaks in the skin and or via inhalation. the organism is believed to access the cns through hematogenous spread, resulting in a chronic, insidious, but often fatal granulomatous amoebic encephalitis (gae), which has been documented in both immunocompetent and immunocompromised hosts [ , ] . the incubation period from exposure to development of clinical symptoms is not well established and experts believe that this may occur between months and years. finally, an alternative mode of transmission via solid organ transplantation has also gained attention [ ] [ ] [ ] . in many cases, gae is diagnosed post-mortem, due to delayed diagnosis or unawareness of the clinical entity. following infection by b. mandrillaris, two clinical patterns of presentation have been described. in the first pattern, patients initially develop a skin lesion that may resemble a painless plaque that may evolve into subcutaneous nodules and rarely ulcerations [ ] . these patients may develop neurologic manifestations weeks to months later. histopathologic examination of these lesions typically reveals granulomatous reactions in the reticular dermis, associated with lymphocytic and plasma cell infiltrates as well as multinucleated giant cells, without distinct epidermal changes. skin lesions will harbor trophozoites, but these are scarce and often easily overlooked as they resemble histiocytes. it is believed that early diagnosis of b. mandrillaris infections in those presenting with skin lesions may prevent subsequent development of cns disease, but there have also been cases in which patients presenting with skin lesions have progressed to developing gae despite treatment. in the second pattern, patients present with cns involvement without a previously recognized skin lesion. patients presenting with gae may initially display fever, malaise, headache, nausea and vomiting, and frank lethargy. later, these symptoms evolve into visual abnormalities, cranial nerve palsies, seizures, focal paresis; as intracranial pressure builds, coma, and eventually death with tonsilar or uncal herniation ensue within - weeks [ ] . upon infection with b. mandrillaris, brain endothelial cells produce the proinflammatory cytokine il- , thereby initiating an inflammatory response [ ] . moreover, the amoebic trophozoites infiltrate blood vessel walls. degradative enzymes, vessel wall infiltration, and the host inflammatory responses result in tissue necrosis and infarctions in the cerebral hemispheres, cerebellum, and the brainstem. in a mouse model of b. mandrillaris infection, cd + t cells were shown to be protective [ ] , suggesting that patients with lowered number or dysfunction in cd + t cells may be more susceptible to disease by this amoeba. however, b. mandrillaris infections have been described in a variety of human hosts [ ] , ranging from the young, healthy, and presumably immunocompetent to the elderly, and those with hiv, chronic corticosteroid exposure, on post-transplant immunosuppression and even patients with cvid. as such, further research is necessary to fully delineate the susceptibility of pidd patients. in patients who develop the characteristic skin lesions, recognition, testing, and treatment for b. mandrillaris may prevent subsequent gae. as such, obtaining tissue and looking for granulomas and trophozoites is quite helpful. skin biopsies can be stained via immunofluorescence or immunoperoxidase techniques to identify b. mandrillaris [ ] . additionally, a pcr technique that identifies mitochondrial s ribosomal rna from b. mandrillaris is also available through the cdc [ ] . in patients in whom the diagnosis is confirmed via skin biopsy, wide resection and medical treatment appears to prevent development of cns disease in at least a proportion of patients. in patients presenting with cns involvement, lumbar punctures reveal csf with lymphocytic pleiocytosis, low-to-normal glucose, and mildly to significantly elevated protein levels. trophozoites are not typically found in the csf, but pcr analysis may be performed. ct or mr imaging may show multiple nodules with ring enhancement; some of these nodules may also contain focal areas of hemorrhage. biopsies of brain tissues typically reveal granulomas and foamy macrophages and multinucleated giant cells surrounded by lymphocytic infiltrates. additionally, there will be areas of necrosis filled with neutrophils, multinucleated giant cells, and lymphocytes, with balamuthia trophozoites and cysts interspersed with macrophages [ ] . as with the skin biopsies, immunofluorescent and immunoperoxidase stains may aid diagnosis and should be performed. unfortunately, the optimal medical management of cns disease is unknown. in the usa, a few patients have been successfully treated with a combination of fluconazole, flucytosine, pentamidine, a macrolide antibiotic (either clarithromycin or azithromycin), and one of the following agents: liposomal amphotericin b, miltefosine, sulfadiazine, or thoridazine [ ] [ ] [ ] ; others in peru have been treated successfully with fluconazole (or itraconazole), albendazole, and miltefosine [ ] . based on these case reports, most experts recommend treatment with a combination of medications (along with partial or complete resection of nodules) for a prolonged period of time to prevent further deterioration and death [ ] [ ] [ ] [ ] . acanthamoeba spp. the genus acanthamoeba contains at least morphologically distinct species that live in a diverse array of habitats, including soil, salt, brackish, and fresh water. acanthamoeba spp. have also been found in humidifiers, heating and cooling unit components, jacuzzis, hot water tanks, bathrooms and drains, eye wash stations and dentistry irrigation systems, and more. acanthamoeba spp. have been isolated from reptiles, birds, and other non-human mammals, suggesting a broad distribution in the environment. acanthamoeba trophozoites feed on bacteria, but have also recently been shown to harbor a number of bacteria (including legionella and burkholderia spp., e. coli, listeria monocytogenes, vibrio cholerae, mycobacteria spp., chlamydophila, and others) and at least one virus (mimivirus) as endosymbionts. acanthamoeba infections in humans can present in a variety of ways. of primary importance are cns infections. like b. mandrillaris, acanthamoeba spp. can induce gae (described above). there is a high predilection for gae in those with hiv/aids, patients on chemotherapy, and those receiving broad spectrum antibiotics [ ] . acanthamoeba are rarely found in csf, but some case reports indicate isolation of amoebae by culturing csf on bacterized agar plates. similar to gae seen with b. mandrillaris, cns histopathology may reveal edema, multiple areas of necrosis and hemorrhage, and occasional findings of angitis and blood vessel cuffing by inflammatory cells, as well as occasional trophozoites or cysts. cns disease treatment is not standardized, but several patients have been successfully treated with pentamidine, fluconazole, flucytosine, sulfadiazine, as well as miltefosine. acanthamoeba can rarely cause cutaneous infections; these lesions, like gae, are also predominantly seen in immunocompromised hosts. these lesions can start as nodules or papules on the lower extremities and develop into necrotic ulcers. histopathologic examination may reveal granulomatous dermal lesions in immunocompetent hosts, with histiocytes, as well as neutrophils and plasmacytes; trophozoites are typically visible [ , ] . the optimal management of cutaneous disease is not known, but typically involves combinational therapy with topical (e.g., chlorhexidine, gluconate, or ketoconazole) and systemic (miltefosine, sulfadiazine, flucytosine, liposomal amphotericin b, azole antifungals, etc.) drugs. additionally, nasopharyngeal and sinus infections have been seen in people with severe compromise in immunity [ , ] . patients typically present with purulent nasal discharge, and examination may reveal erosion of the nasal septum. nasopharyngeal disease can present concomitantly with cutaneous disease. treatment of nasopharyngeal or sinus disease is difficult and involves surgical debridement and combinations of systemic drugs. disseminated disease is also seen in immunocompromised hosts and typically involves concomitant pulmonary and cutaneous disease in the presence or absence of cns infection. keratitis readily occurs in immunocompetent hosts-with the major risk factor being contact lens wearing without proper adherence to recommended cleansing protocols. this infection less commonly presents as a result of direct inoculation with trauma. one of the most common reasons for contact lens wearers to acquire disease is due to the use of non-sterile tap water in preparing contact lens saline solutions [ ] , although contaminated solutions from manufacturers have also been identified. patients will have pain and photophobia. physical exam reveals conjunctival injection and epithelial abnormalities (including pseudodendritic lesions) and stromal infiltrates [ ] . the proper diagnosis can be made by staining corneal scrapings with calcofluor or wright-giemsa stains and examined by confocal microscopy, culture, or pcr analysis. prompt therapy with a combination of polyhexamethylene biguanide (or biguanide-chlorhexidine) and propamidine or hexamidine [ , ] is indicated, but misdiagnosis and delayed therapy are common. more severe cases may also require debridement. the use of topical steroids before administration of combinational therapy may result in worse outcomes and should be avoided; however, if scleritis ensues, it may be necessary to use immunosuppressants to reduce the need for enucleation. severe and/or refractory cases may result in the need for cornea transplantation. phenotypes seen in pidd like hsv- and candida [ ] . patients with ifnar deficiency seem highly susceptible to cns disease caused by mmr vaccine, an otherwise extremely rare phenomenon [ ] . recently, a case of noroviral cns disease was described associated with a novel, yet unpublished pidd, suggesting that some pidds may lead to susceptibility of the cns to viruses that normally do not exhibit neurotropism (casanova jl, personal communication) . this again favors metagenomic approaches in the study of cns sequelae in pidd patients. in pidds, the cns is also more vulnerable to virally induced immunodysregulation [ ] . conditions like primary hemophagocytic lymphohistiocytosis (hlh) may present as isolated cns disease or relapse only in the cns [ ] [ ] [ ] . almost % all human malignancies are associated with chronic infections by hbv, hcv, hpv, ebv, hhv /kshv, htlv-i, hiv- , hiv- , jcv, merkel cell polyomavirus (mcpv), helicobacter pylori, schistosomes, or liver flukes [ ] . accordingly, pidd patients' malignancies are often associated with chronic infections. mcpv-associated merkel cell carcinoma has now been described in gata and tmc (ever ) deficiencies as well as other forms of pidd [ , [ ] [ ] [ ] [ ] [ ] . large follow-up cohorts are needed to refute or confirm associations between novel pidds and malignancies, such as hyperactivating pik cd and ovarian dysgerminoma or gata deficiency and leiomyosarcoma [ , ] . recently, hymenolepis nana was found to have driven malignant transformation in an hiv patient. likely, other novel pidd-and pathogen-associated malignancies will be found in the future by those with an open and inquisitive mind [ ] . understanding the specific infection susceptibility for each pidd allows not only a better understanding of host defense, but also allows the clinician to collaborate with the microbiology laboratory to make definitive diagnoses and provide the best therapy. reviewing all of the infections for each pidd is not within the scope of this article, but there are several infections that are unique for specific pidds and require special attention from the microbiology laboratory. three examples are provided below. g. bethesdensis is a gram-negative bacterium that was identified to cause disease in patients with cgd in [ ] . g. bethesdensis is a member of the methylotroph group of bacteria, which are able to use single-carbon organic compounds as their only source of energy. they are widespread in the environment, but are rare human pathogens, and infections with g. bethesdensis have been limited thus far to patients with cgd. the organism was first detected in an adult patient with indolent and recurrent necrotizing lymphadenitis [ ] . subsequently, g. bethesdensis was isolated from nine patients with cgd, primarily causing lymphadenitis, but there have been two fatalities [ ] . treatment has been most effective with intravenous ceftriaxone. the microbiology laboratory should be alerted when there is concern for g. bethesdensis infection to allow for proper culture media. charcoal yeast extract (cye) agar and lowenstein jensen (lj) media are appropriate culture media. mycoplasma and ureaplasma spp. as molecular techniques are becoming more widely used to detect pathogens, the spectrum of infections that were previously only detected through serologic assays and research laboratories will increase. this is important especially for patients with pidd who have unique susceptibility to infection and may not have the ability to mount a serologic response. examples of infections in this group are those caused by mycoplasma and ureaplasma [ , ] . these pathogens have been known to cause osteoarticular infections for those with antibody deficiency, such as xla and cvid. recently, mycoplasma orale, typically an oral commensal, has been isolated from two patients with defects in the activated pi k delta syndrome, as chronic lymphadenitis in one and chronic splenic abscess in the other (sm holland personal communication). defects in pi kcd and pi kr are frequently associated with hypogammaglobulinemia and therefore would fit in the pattern of mycoplasma infections in those with humoral immunodeficiency. mycoplasma orale has also previously been reported as causing bone disease in a patient with cvid [ ] . in patients with xla, helicobacter, camplyobacter, and the related flexispira bacteria that are typically isolated to the gi tract can disseminate and often lead to chronic bacteremia, ulcers, and bone infections [ ] [ ] [ ] . xla patients have higher susceptibility than other humoral pidd and are thought to be due to the role that igm is playing in controlling the dissemination of these pathogens and potentially iga in providing mucosal immunity. these bacteria can be fastidious to grow, and therefore, when there is suspicion, identification needs collaboration with the microbiology laboratory. for instance, the blood culture media may allow growth (although with a longer incubation period), but then the organisms may need molecular techniques for identification, such as s sequencing, as they will not grow on the agar plates. treatment is often difficult, requiring combination antimicrobials for prolonged periods (such as year), and relapse is common. this review provides an important perspective for practicing immunologists, namely that we are a part of a global community as are our patients. this overview of emerging infections and infectious concerns for travelers serves as a foundation for practical considerations for clinicians and patients. using prevalence data, an estimation of the number of infected patients with pidd (table ) can be developed [ , , [ ] [ ] [ ] [ ] . thus, the concerns addressed in this review are not theoretical but 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management clinical presentation and medical management of melioidosis in children: a -year prospective study in the northern territory of australia and review of the literature a review of melioidosis cases in the americas melioidosis in travelers: review of the literature laboratory diagnosis of melioidosis: past, present and future fatal bacteremic melioidosis in patients with prolonged neutropenia pathogenic free-living amoebae: epidemiology and clinical review primary amoebic meningoencephalitis caused by naegleria fowleri: an old enemy presenting new challenges the immune response to naegleria fowleri amebae and pathogenesis of infection production of monoclonal antibodies to naegleria fowleri, agent of primary amebic meningoencephalitis pathogenic and opportunistic free-living amoebae: acanthamoeba spp., balamuthia mandrillaris, naegleria fowleri, and sappinia diploidea multiplex real-time pcr assay for simultaneous detection of acanthamoeba spp., balamuthia mandrillaris, and naegleria fowleri successful treatment of naegleria fowleri meningoencephalitis by using intravenous amphotericin b, fluconazole and rifampicin successful treatment of primary amebic meningoencephalitis successful treatment of an adolescent with naegleria fowleri primary amebic meningoencephalitis centers for disease c, prevention. notes from the field: transplant-transmitted balamuthia mandrillaris-arizona balamuthia mandrillaris transmitted through organ transplantation-mississippi transmission of balamuthia mandrillaris by organ transplantation balamuthia mandrillaris infection of the skin and central nervous system: an emerging disease of concern to many specialties in medicine infections with free-living amebae balamuthia mandrillaris exhibits metalloprotease activities resistance to intranasal infection with balamuthia mandrillaris amebae is tcell dependent balamuthia mandrillaris amoebic encephalitis: an emerging parasitic infection histopathologic spectrum and immunohistochemical diagnosis of amebic meningoencephalitis demonstration of balamuthia and acanthamoeba mitochondrial dna in sectioned archival brain and other tissues by the polymerase chain reaction successful treatment of balamuthia amoebic encephalitis: presentation of cases balamuthia mandrillaris meningoencephalitis: survival of a pediatric patient balamuthia mandrillaris brain abscess successfully treated with complete surgical excision and prolonged combination antimicrobial therapy neurosurgical intervention in the diagnosis and treatment of balamuthia mandrillaris encephalitis cutaneous acanthamebiasis infection in immunocompetent and immunocompromised patients tender ulceronecrotic nodules in a patient with leukemia. cutaneous acanthamebiasis parasitic sinusitis and otitis in patients infected with human immunodeficiency virus: report of five cases and review acanthamoeba: a rare primary cause of rhinosinusitis the epidemiology of acanthamoeba keratitis in the united states acanthamoeba sclerokeratitis. determining diagnostic criteria acanthamoeba keratitis update-incidence, molecular epidemiology and new drugs for treatment comparison of polyhexamethylene biguanide and chlorhexidine as monotherapy agents in the treatment of acanthamoeba keratitis primary immunodeficiency diseases: an update on the classification from the international union of immunological societies expert committee for primary immunodeficiency heterozygous stat gain-of-function mutations underlie an unexpectedly broad clinical phenotype clinical and immunologic phenotype associated with activated phosphoinositide -kinase delta syndrome : a cohort study clinical spectrum and features of activated phosphoinositide -kinase delta syndrome: a large patient cohort study the extended clinical phenotype of patients with dedicator of cytokinesis deficiency new clinical phenotypes of fungal infections in special hosts progressive multifocal leukoencephalopathy in primary immune deficiencies: stat gain of function and review of the literature kaposi sarcoma of childhood: inborn or acquired immunodeficiency to oncogenic hhv- . pediatr blood cancer immunologic defects in severe mucocutaneous hsv- infections: response to ifn-gamma therapy paracoccidioidomycosis and other unusual infections in brazilian patients hyper igm syndrome: a report from the usidnet registry genetic variation in schlafen genes in a patient with a recapitulation of the murine elektra phenotype warts and all: human papillomavirus in primary immunodeficiencies severe infectious diseases of childhood as monogenic inborn errors of immunity enrichment of rare variants in population isolates: single aicda mutation responsible for hyper-igm syndrome type in finland a homozygous card mutation in a family with susceptibility to fungal infections genetic, immunological, and clinical features of patients with bacterial and fungal infections due to inherited il- ra deficiency primary polyomavirus infection, not reactivation, as the cause of trichodysplasia spinulosa in immunocompromised patients tlr deficiency in herpes simplex encephalitis: high allelic heterogeneity and recurrence risk hemophagocytic lymphohistiocytosis with isolated central nervous system reactivation and optic nerve involvement isolated central nervous system hemophagocytic lymphohistiocytosis: case report predominant neurological manifestations seen in a patient with a biallelic perforin mutation (prf ; p.r w) the prevention of infection-associated cancers merkel cell carcinoma in a patient with gata deficiency: a novel association with primary immunodeficiency spontaneous regression of merkel cell carcinoma developed in a patient with epidermodysplasia verruciformis merkel cell polyomavirus detection in a patient with familial epidermodysplasia verruciformis merkel cell polyomavirus-positive merkel cell carcinoma in a patient with epidermodysplasia verruciformis merkel cell polyomavirus in merkel cell carcinoma from a brazilian epidermodysplasia verruciformis patient association of gata deficiency with severe primary epstein-barr virus (ebv) infection and ebv-associated cancers malignant transformation of hymenolepis nana in a human host granulibacter bethesdensis gen. nov., sp. nov., a distinctive pathogenic acetic acid bacterium in the family acetobacteraceae methylotroph infections and chronic granulomatous disease osteoarticular infectious complications in patients with primary immunodeficiencies increased susceptibility to mycoplasma infection in patients with hypogammaglobulinemia disseminated mycoplasma orale infection in a patient with common variable immunodeficiency syndrome relapsing campylobacter jejuni systemic infections in a child with x-linked agammaglobulinemia bacteremia and skin/bone infections in two patients with x-linked agammaglobulinemia caused by an unusual organism related to flexispira/helicobacter species successful approach to treatment of helicobacter bilis infection in x-linked agammaglobulinemia prevalence and morbidity of primary immunodeficiency diseases, united states epidemiological characteristics of spotted fever in israel over years complications of bacille calmette-guerin (bcg) vaccination and immunotherapy and their management acknowledgements the authors would like to thank thomas krell for information regarding ivig safety and david peden for recognizing global warming as central to medical knowledge. conflict of interest the authors declared that they have no conflict of interest. pidds display wide genetic and phenotypic heterogeneity [ ] . similar disease phenotypes may be caused by multiple genes, while patients' phenotypes caused by the same gene and even by the same mutations vary between individuals. importantly, after a novel pidd has been described, subsequent reports often reveal a wider variation in associated infections and cellular findings, often without clear genotype-phenotype correlations [ ] [ ] [ ] [ ] [ ] . variation may be caused by mechanisms such as other contributing genes or geographical variation in infectious exposures. geographic differences seem most pronounced in intracellular and often chronic infections. while the numbers of described pidd patients increase, at first seemingly rarely pidd-associated infections turn out to be found in a significant subset of pidd patients [ ] [ ] [ ] . for example, patients with cd l deficiency living in endemic areas display susceptibility to bartonellosis and paracoccidioidomycosis, infections not described in european and us cohorts [ , , ] .often, an infectious phenotype previously only described in secondary immunodeficiencies may reveal the possibility of an underlying primary immunodeficiency [ , , ] . increasing numbers of genetic defects causing early-onset, severe, and recurrent susceptibility to commonly circulating pathogens like pneumococci, tuberculosis, herpes simplex, and influenza viruses as well as endemic protozoans like trypanosomes and fungi are being recognized, and thus, infections with unusual pathogens require a high index of suspicion for pidd [ ] . in contrast, pidds may also manifest as suspected infection but sterile inflammation. for example, in inflammatory lesions like granulomas and necrotizing fasciitis where no clear pathogens are found, one needs to rule out aberrant host responses due to pidd [ ] .chronic viral and fungal infections may also display novel phenotypes never or rarely seen in secondary immunodefic i e n c i e s . i n f e c t i o n s l i k e d e r m a t o p h y t o s i s a n d phaeohyphomycosis deeply infiltrating the skin and lymph nodes, occasionally extending to bones and central nervous system (cns) as well as predisposition to primary cns candidiasis and extrapulmonary aspergillus slowly revealed the full phenotypic spectrum of card deficiency [ , , ] . chronic skin ulcers caused by hsv- and severe molluscum contagiosum suggest dock deficiency or gain-of-function mutations of stat [ , ] . chronic mucocutaneous candidiasis has revealed a large group of monogenic diseases (il ra, il rc, il f, stat (gof), rorx, act ), which may also be associated with recurrent bacterial infections or syndromic features [ ] . while novel diseases by newly described viruses are being discovered, one needs awareness to suspect these in pidd patients [ ] .interestingly, most novel forms of infectious disease in pidd patients have been described either in easily accessible sites like the skin or in immunologically privileged, normally sterile sites like the cns. this suggests that with the increasing use of invasive sampling and sensitive metagenomic approaches, we might find more novel infectious phenotypes. pathogens highly suggestive of certain pidds, like chronic enteroviral cns infections in xla patients are reviewed above. in hypomorphic mutations, cns seems to be especially vulnerable to chronically active and/or recurrent novel infectious bsmoldering^focal encephalitis lesions by pathogens key: cord- -efc msf authors: blumberg, lucille title: severe malaria: manifestations, diagnosis, chemotherapy, and management of severe malaria in adults date: journal: tropical and parasitic infections in the intensive care unit doi: . / - - - _ sha: doc_id: cord_uid: efc msf nan the burden of malaria is increasing, especially in sub-saharan africa, because of drug and insecticide resistance and social and environmental changes ( ). each year an estimated three to four hundred million people will contract malaria globally, resulting in five hundred thousand to two million deaths. ninety percent of the world's malaria, and at least % of malaria-related mortality, occurs in sub-saharan africa, primarily in young children ( ). malaria occurs in every country in sub-saharan africa, with the exception of lesotho, but transmission rates vary within regions and within countries. in parts of africa where endemicity of malaria is high and transmission stable, such as tanzania, malawi, and mozambique, severe malaria is mainly a disease of children under years of age and of pregnant women. it is less common in older children and adults because of the partial immunity acquired as a result of repeated infections. in areas of low endemicity severe malaria occurs in both adults and children. non-immune travellers to malaria areas are always at risk for severe disease ( , ). the majority of malaria cases in africa are due to plasmodium falciparum, the major species associated with mortality and morbidity. the development of parasite resistance to chemotherapeutic agents such as chloroquine has resulted in a significant increase in malaria morbidity and mortality. the demise of chloroquine, an affordable option in resource-poor countries, has major implications for malaria management ( ). in africa resources for management of severe malaria are limited and at least - % of patients with complications of disease will die. in a confidential inquiry into malaria deaths in an area of south africa with limited tertiary care facilities, major contributing factors were delays in diagnosis and initiation of adequate therapy, failure to administer the correct antimalarial at the correct dosage and frequency, inadequate monitoring of severity indicators in complicated cases, and the suboptimal management of complications ( ). key features of malaria are the adherence of infected red blood cells to the endothelium of small blood vessels compromising blood flow through tissues, and the production of pro-inflammatory cytokines ( ). factors that determine whether a patient develops mild or severe disease are complex and multifactorial and are related to both the parasite and the host. parasites causing severe malaria have a greater multiplication potential than those causing uncomplicated infections ( ). the effect of inoculum dose on severity is unclear and difficult to investigate. cyto-adherence of parasitised red cells may be influenced by the virulence of different strains of parasite ( ). the development of immunity to the clinical effects of malaria requires several years of continuous exposure. lack of this protective immunity would be expected to be the major factor determining the severity of a clinical attack of malaria. differences in hla antigens may play a role in host predisposition to severe disease. certain red blood cell abnormalities, including sickle-cell trait, protect against malaria disease. prevalence rates of these abnormalities are high in some parts of africa and may provide some protection against severe malaria ( ). plasma interleukin (il- , il- ) and tumour necrosis and the il- : il- ratio is significantly higher in patients who die than in survivors ( ). symptoms and signs of malaria may present as early as seven days, but more commonly an average of - days after being bitten by an infected mosquito. fever is prominent, but may be absent in some cases. some of the following symptoms may also appear: rigors, headache, myalgia, diarrhoea, vomiting and cough. physical signs may include fever, anaemia, jaundice, hepatosplenomegaly and a variety of cerebral signs. malaria should be suspected in any person presenting with any of the above symptoms or signs with a history of travel to, or residence in a malaria transmission area. presentation is very variable and may mimic other diseases, including influenza, hepatitis, meningitis, septicaemia, typhoid, tickbite fever, viral haemorrhagic fever, trypanosomiasis, hiv seroconversion illness, and relapsing fever ( ). p. falciparum infections may progress rapidly to a lethal, multi-system disease. the diagnosis of malaria is urgent, and complications can develop rapidly within hours of the onset of disease in any non-immune person but especially in young children and pregnant women ( ). the clinical manifestations of severe malaria depend on the age of the patient. in children, hypoglycaemia, convulsions, and severe anaemia are relatively common; acute renal failure, jaundice, and ards are more common in adults. cerebral malaria, shock and acidosis may occur at any age ( ). a number of clinical and laboratory criteria are used to define severe malaria, as shown in table ( , ). patient blood should be examined immediately to confirm or exclude the diagnosis of malaria. in the majority of cases of severe malaria, examination of correctly stained blood smears will reveal malaria parasites, however, a negative smear does not exclude the diagnosis, and repeat smears are indicated. some patients with severe malaria may have a negative smear due to sequestration of parasitised red blood cells, and a decision to treat with antimalarial chemotherapy should be considered if the index of suspicion is very high. in these cases it is imperative to continue to look for alternative diagnoses, especially trypanosomiasis, septicaemia and viral haemorrhagic fever. high levels of parasitaemia are generally predictive of severe malaria in nonimmune patients. importantly, the converse may not be true, with severe disease also occurring with low parasitaemias in the peripheral blood ( , ) . quantification is often inaccurate, peripheral parasitaemia may not reflect the total parasite load and sequestration in the organs, and levels of parasitaemia may vary cyclically. prognosis worsens considerably if p. falciparum schizonts are present in a blood smear, and if more than % of peripheral polymorphonuclear leucocytes contain visible malaria pigment ( ). commercial kits are available that rapidly detect parasite antigen or enzymes. the tests for p. falciparum are highly sensitive, but depend on correct usage, interpretation of results, and the quality of the particular test used. these tests can only be used for diagnosis of acute malaria infections, and not for follow-up, as the test may remain positive for several weeks, even after successful treatment ( ). in a febrile patient where there is no obvious cause of fever, and a recent history of visiting or living in a malaria area is not forthcoming, malaria should still be excluded, as infected mosquitoes have been documented to travel long distances in road, rail and air transport. mortality is high in this group of patients, because of missed diagnosis, but a finding of thrombocytopenia should always stimulate a search for possible malaria parasites ( ). patients should be treated urgently with the most effective treatment regimen available, in a facility with the highest level of care. the choice of chemotherapy for malaria is dependent on the severity of disease, the known or suspected resistance pattern of the parasite in the area where the malaria infection was acquired, the species of parasite, and patient profile (age, pregnancy, comorbidity, allergies, and medications, including any antimalarials recently administered). quinine, the drug of choice for the treatment of severe malaria in africa, is rapidly effective ( , ). in most parts of africa quinine resistance has not developed. in some parts of west africa however, foci of low-level resistance have been documented ( ). an initial loading dose of quinine to rapidly reach a therapeutic level is critical in the management of severe malaria and has a major impact on favourable outcome. the loading dose should be omitted if the patient has definitely received mefloquine, quinine, quinidine or halofantrine in the previous hours, mefloquine in the previous seven days, or mg/kg of quinine in the previous two days. if there is doubt, the loading dose of quinine should be given ( , , ) . the loading dose is given as quinine di-hydrochloride salt, mg/kg body weight diluted in - ml/kg body weight of dextrose water, by slow intravenous infusion over two to four hours. quinine must never be administered by bolus intravenous injection, as this is associated with cardiotoxicity. the loading dose is given strictly according to body weight. the disposition of quinine in very obese patients is not known. it has been suggested that there is a ceiling dose above which quinine should not be given, but there is no evidence to support this ( ). six to eight hours after starting the loading dose, a maintenance dose of quinine di-hydrochloride salt, l mg/kg diluted in - ml/kg body weight of a dextrose-containing solution should be commenced and infused over - hours. intravenous quinine should be administered every eight hours until the patient can take oral medication (usually by hours). for obese patients, the maintenance dose should be calculated according to ideal body weight ( ). males: ibw (kg) = . x height in cm - females: ibw (kg) = . x height in cm - . the dosage of oral quinine is l mg/kg/dose or mg/dose given three times a day. the total duration of quinine therapy is - days. additional drugs, tetracycline (usually as doxycycline mg twice a day x days), or clindamycin (l mg/kg twice a day x days) are recommended to improve cure rates ( , , ). these, however, do not add initial therapeutic benefit, may contribute to drug side effects, and should be introduced only once the patient is improving. quinine can be administered by deep intramuscular injection if intravenous infusion is not possible ( ). quinine has a narrow therapeutic window, although serious side effects are rare. the pharmacokinetic properties of quinine are altered considerably in malaria with a contraction in the volume of distribution and a reduction in clearance that is proportional to the severity of disease ( ). there is significant binding of quinine to acute phase reactants, notably glycoprotein, with reduction in the levels of free quinine. quinine toxicity is, therefore, relatively uncommon ( ). the most frequent side effect of quinine therapy is hypoglycaemia, especially in children and pregnant women ( , ) . although quinine may prolong the qtc-interval, hypotension, heart block, and ventricular arrhythmias are rare ( , , ) . convulsions and visual disturbances have been reported as idiosyncratic responses or with overdosage ( , ). doses should be reduced by - % after the third day of treatment to avoid accumulation of the drug in patients who remain seriously ill, especially those with evidence of renal failure ( ). the measurement of levels of free (not total) quinine may be helpful in patients with severe malaria and renal failure, but accessibility to this test is very limited. the precise level has not been defined but probably lies between . - mg/l ( ). quinidine is more active than quinine, but is also more cardiotoxic and more expensive, is not readily available, and consequently is not used for treating severe malaria in africa ( ). in the early 's chinese scientists identified artemisinin, a sesquiterpene lactone peroxide, as the principal active component of the traditional chinese malaria remedy, qinghaosu. artemisinin and two derivatives, artesunate and artemether are effective against multi-drug resistant p falciparum and clear sensitive parasites from the blood more rapidly than other antimalarial agents due to their broad stage specificity of anti-malarial action. despite administration to over million people, resistance has not emerged, and only rarely has treatment failure been reported. the drugs are well tolerated and despite neurotoxicity in animal studies, serious adverse reactions have included only a few case reports of anaphylaxis. the chemical structure and mode of action of these drugs distinguish them from other currently available antimalarial agents, and render them less vulnerable to cross-resistance. however, when used alone, unacceptably high recrudescence rates are seen ( , , , ). combination therapy, which includes an artemisinin, is the recommended malaria treatment policy to delay the emergence of drug resistance to sequential monotherapy, as well as to improve cure rates. drugs used in combination with the artemisinins include mefloquine, sulfadoxine pyrimethamine, amodiaquine and lumefantrine, and the choice depends on parasite resistance in the geographical area ( ). there are parenteral preparations of the artemisinins, either intramuscular (artemether, arteether, artesunate) or intravenous (artesunate). artemether and arteether are oil-based preparations and absorption from the intramuscular site may be compromised in severe malaria, leading to treatment failures ( ). artesunate is water-based, can be given intravenously, or intramuscularly from where it is well absorbed. although theoretically preferable, there are no large comparative trials to indicate whether artesunate is superior to artemether or quinine ( ). the use of parenteral artemisinins is limited by availability and manufacturing practices, which may not adhere to international standards. a meta-analysis of randomized clinical trials comparing the efficacy of artemether with quinine in the management of severe malaria demonstrated equality, but indicated a trend toward greater effectiveness of artemether in regions where there is recognised quinine resistance. artemether was superior to quinine in terms of overall serious adverse events ( , ). in patients with hyperparasitaemia there may be an advantage of the artemisins over quinine. in south-east asia, where multi-drug-resistant malaria is a major problem and quinine resistance has emerged, the artemisinin drugs are used as first-line therapy for severe malaria ( ). widespread, high-level chloroquine resistance precludes the use of chloroquine in the treatment of both uncomplicated and severe malaria in most parts of the world, including africa. sulfadoxine pyrimethamine, mefloquine and halofantrine are not indicated in the management of severe malaria ( ). anaemia may result from haemolysis or dyserythropoeisis ( ). severe anaemia is defined as a haemoglobin of less than g/dl, or haematocrit < %. severe anaemia is the most important manifestation of severe malaria in areas of high stable transmission and occurs predominantly in children. pregnant women may also present with profound degrees of anaemia. anaemia may manifest as shock, cardiac failure, hypoxia, or confusion and the rate at which anaemia develops is an important determinant of compensatory mechanisms. blood transfusion using packed cells should be considered in patients in whom the haemoglobin is g/dl or less, especially those with cardiovascular decompensation. fluid overload must be avoided. transfused blood has a reduced lifespan in malaria patients ( ). in many parts of the world cerebral malaria is the most common clinical presentation and cause of death in adults with severe malaria. the term cerebral malaria in many published studies is restricted to the syndrome in which altered consciousness associated with a malaria infection could not be attributed to convulsions, sedative drugs or hypoglycaemia alone or to a non-malarial cause. cerebral malaria may be part of multi-system pathology, in which case the outlook is much poorer than if disease was localised only to the central nervous system. clinically, the commonest neurological picture is of a symmetrical upper motor neuron lesion, mild neck stiffness is not uncommon, and muscle tone and tendon reflexes are variable. cerebral malaria can resemble bacterial or viral meningitis and a lumbar puncture should be considered in patients where the diagnosis is not clear. hypoglycaemia, metabolic disturbances, severe anaemia and hypoxia as a result of malaria can all present with signs of central nervous system dysfunction. generalised or focal convulsions may occur as a result of cerebral malaria, or in association with hypoglycaemia ( ). imaging of the brain commonly shows evidence of mild cerebral swelling. oedema is very unusual, and may be an agonal phenomenon ( , ). studies to date with dexamethasone or mannitol have not shown benefit and have been associated with prolongation of coma and gastro-instestinal haemorrhage ( ). anticonvulsants should only be used once convulsions occur, and should not be used prophylactically ( ). the use of ironchelating agents has not been shown to impact on mortality ( ). in adult patients who recover, neurological sequelae are uncommon. renal failure is an early complication of severe malaria in adults. hypovolaemia, sequestration of parasitised red cell in the renal vasculature, intravascular haemolysis and haemoglobinuria are implicated and may lead to acute tubular necrosis. renal failure is generally oligaemic and hypercatabolic. a serum creatinine of greater than or a rapidly rising creatinine and/or a urine output of < ml/day in an adult should be regarded as renal failure. a central venous catheter (cvp) should be inserted and dehydration should be corrected. the cvp should not be above cm of water. the indications for dialysis are the same as for patients with other diseases, but since renal failure in malaria occurs against a background of a hypercatabolic state and non-renal causes of acidosis frequently co-exist, early dialysis is recommended ( ). venovenous haemofiltration is the recommended mode of dialysis and is significantly more efficient than peritoneal dialysis ( ). quinine is not removed by dialysis and in patients with severe malaria and renal failure, the dosage of quinine should be reduced by half to one-third after days of full dosage administration. if the patient survives the acute phase of the disease and has no pre-existing underlying disease, recovery of renal function generally occurs within three weeks ( ). this is a grave complication of severe falciparum malaria in adults, and may present several days after commencing malaria chemotherapy. the cause of this often lethal complication is unknown in falciparum malaria. some cases show evidence of pulmonary oedema while others resemble acute respiratory distress syndrome. pregnant women are particularly at risk. latrogenic overadministration of fluids may contribute to the development of ards or pulmonary oedema and should be avoided. some patients may require ventilatory support ( , , , ) . although a raised indirect bilirubin due to haemolysis is a frequent finding in malaria, the clinical presence of jaundice or the finding of raised hepatic transaminases x normal) should alert the clinician to the probability of severe malaria. the presence of jaundice combined with renal failure and acidosis may indicate a grave prognosis ( ). dic is rare in patients with severe malaria although laboratory evidence of haemostatic abnormalities may be present without bleeding. moderate degrees of thrombocytopenia are noted in the majority of cases of uncomplicated malaria unassociated with other coagulation abnormalities and bleeding is uncommon. possible mechanisms of thrombocytopenia include sequestration in the spleen, decreased production, or reduced survival from intravascular lysis. platelet transfusion should be considered if the platelet count is less than or if there is evidence of bleeding. platelet counts should return to normal within a few days with effective malaria treatment. continuing thrombocytopenia may indicate failed antimalarial therapy, sepsis, or a drug reaction to quinine ( ) secondary bacterial infections may complicate malaria: aspiration pneumonia, urinary tract infections or nosocomial septicaemia. in a significant number of patients, especially children, septicaemia may complicate severe malaria very early. salmonella species and staphylococci are common causes of septicaemia. the syndrome is associated with high mortality. since the features of bacterial sepsis and malaria overlap, empiric treatment using a broad-spectrum antibiotic for gram-positive and gram-negative organisms is recommended ( ). metabolic acidosis is a consistent feature of severe malaria. lactic acidosis is a major cause of death from severe falciparum malaria. the pathophysiology of acidosis is multifactorial and results from tissue hypoxia and anaerobic glycolysis, liver dysfunction and impaired renal handling of bicarbonate. the presence of acidosis is an important predictor of poor outcome ( ). the management of acidosis includes correction of fluid balance, improvement in haemodynamic status, and haemodialysis ( ). the use of dichloracetate has been shown to be beneficial in animal models. the pathogenesis of this rare condition is unknown, and is seen in patients with g- -pd deficiency who receive oxidant drugs. it may also occur in patients without apparent g- -pd deficiency but who have severe malaria and are treated with quinine or artemisinin derivatives. intravascular haemolysis results in anaemia, and the passage of haemoglobinuria. a small minority will develop renal failure, the cause of which is unknown. in patients with malarial haemoglobinuria, quinine chemotherapy should be continued. supportive therapy includes blood transfusions for severe anaemia, maintaining adequate hydration, and renal dialysis where indicated ( , ). hypoglycaemia may result from impaired glycolysis or gluconeogenesis, or as a result of quinine-induced hyperinsulinaemia. it is a particular problem in pregnant women and patients on intravenous quinine. blood glucose should be monitored, as the signs may be very subtle. hypoglycaemia must be excluded in all patients with an altered mental state and in those who present with convulsions ( ). shock may occur as a result of hypovolaemia, massive blood loss from splenic rupture or gastrointestinal haemorrhage, bacterial septicaemia, hypoxia and severe metabolic acidosis. myocardial function is remarkably good in severe falciparum malaria and most patients have an elevated cardiac index ( ). hypovolaemia should be corrected with an appropriate intravenous infusion, usually . % saline initially, followed by a plasma expander. the central venous pressure should not be allowed to exceed cm. if hypotension persists, inotropes should be administered ( ). the placenta acts as a haven for parasites due to upregulation of adhesion receptors. the course of malaria in pregnancy is rapidly progressive and common complications are anaemia, hypoglycaemia and ards. the risk of severe disease extends into the immediate postpartum period. malaria may cause abortion, premature delivery and low birth-weight. the management remains the same as in non-pregnant patients, with emphasis on preventing and managing the complications mentioned. in particular, fluid restriction is important to prevent ards. quinine is the drug of choice but may be associated with intractable hypoglycaemia. the use of the artemisinin drugs is currently not indicated due to a lack of safety data, unless there is evidence of quinine resistance. there is no indication to terminate pregnancy. in areas of high malaria transmission, anaemia is the most common manifestation of severe disease and placental parasitaemia is associated with low birth-weight infants ( ). the non-falciparum malarias are not generally associated with severe disease due to a lack of sequestration of parasitised red cells. rarely plasmodium vivax has been associated with the development of ards and cerebral malaria ( , ) . mixed infections with falciparum malaria occur occasionally and should be managed as for falciparum malaria. malaria and human immunodeficiency virus (hiv) infections are common, widespread and overlapping problems in africa. any interaction between these two pandemics would be of great importance. this interaction could be in either direction, with malaria causing more rapid progression of hiv, and hiv-associated immunosuppression leading to an impaired immune response to malaria. greater parasite densities and rates of clinical malaria have been demonstrated in hiv-positive patients from uganda, an area of high malaria endemicity, where the majority of people would be expected to have developed some malaria immunity ( , ) . in a cohort study of non-immune patients with malaria in south africa, hiv-positive patients had an increased rate of severe malaria compared to hiv-negative patients, and the rate increased as cd + cell count decreased. hiv-positive patients had significantly increased rates of renal failure, severe anaemia and dic ( ). the efficacy of exchange transfusion as adjunctive therapy for severe malaria is controversial. no sufficiently powered, randomized, controlled study has been reported, although anecdotal case reports in the literature indicate benefits in selected groups of patients with hyperparasitaemia and organ failure ( , ). a meta-analysis of eight studies comparing survival rates associated with exchange transfusion to survival rates with antimalarial chemotherapy alone did not show improved survival rates in the former groups of patients. there were significant problems with the comparability of treatment groups in the studies reviewed, with higher levels of parasitaemia and more severe malaria in the group who received transfusions ( ). recent studies suggest that the benefits associated with exchange transfusion result from replacing the rigid, non-deformable parasitised and unparasitised red cells with fresh blood, and not from reducing parasite load or removal of toxins or cytokines ( ). requirements for exchange transfusion include the availability of pathogen-free compatible blood, facilities for adequate clinical monitoring, and a haemodynamically stable patient. exchange transfusion may be considered in a patient who is seriously ill and the parasitaemia exceeds %. exchange should still be considered with parasitaemia in the range of - %, if there are other signs of poor prognosis. there is no consensus of the volume of blood to be exchanged for a given parasitaemia and the volumes have varied from litres to litres. blood may be exchanged using a double-lumen catheter or alternatively via haemodialysis ( , ). successful red blood cell exchange using a cell separator has been reported ( ). in a study conducted in a well-established intensive-care unit in south africa, despite appropriate chemotherapy with quinine, and standard intensive-care support including inotropic agents, ventilatory support and haemodialysis where appropriate, mortality was . % in a group of patients ( adults and children). pregnancy was a major cause of unfavourable outcome. ards was the most important cause of death. high apache ii scores, high arterial lactate, and negative base excess in the first hours of admission correlated with mortality. admission haemoglobin, platelet count, level of parasitaemia and level of glasgow coma scores in the first hours were shown not be predictors of mortality. these parameters may be useful in the assessment of disease severity and in patient triage for icu admission ( ). marsh k. malaria disaster in africa age-dependent characteristics of protection v. susceptibility to plasmodium falciparum severe falciparum malaria impact of chloroqine resistance on malaria mortality confidential 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severe falciparum malaria the anaemia of falciparum malaria do patients with cerebral malaria have cerebral oedema? a computer tomography study magetic resonance imaging of the brain in patients with cerebral malaria dexamethsaone proves deleterious in cerebral malaria. a double-blind trial in comatose patients phenobarbitone prophylaxis in childhood cerebral malaria: final results of a randomized, controlled intervention study effect of iron chelation therapy on recovery from deep coma in children with cerebral malaria malaria and acute renal failure acute renal failure in patients with severe falciparum malaria pulmonary damage associated with falciparum malaria. a report of ten cases acute pulmonary oedema in falciparum malaria the pathophysiologic and prognostic significance of acidosis in severe adult malaria blackwater fever in southern vietnam: a prospective descriptive study of cases acute respiratory distress syndrome complicating plasmodium vivax malaria cerebral involvement in benign tertian malaria effect of hiv- and increasing immunosuppression on malaria parasitaemia and clinical episodes in adults in rural uganda: a cohort study fresh from the field: some controversies in tropical medicine and hygiene. hiv and malaria, do they interact? exchange blood transfusion in severe falciparum malaria: retrospective evaluation of patients treated with, compared to patients treated without, exchange transfusion exchange transfusion as an adjunct to the treatment of severe falciparum malaria: case report and review exchange transfusion as an adjunct therapy in severe plasmodium falciparum malaria: a meta-analysis what is the future of exchange transfusion in severe malaria? predictors of mortality in severe malaria: a two-year experience in a non-endemic area key: cord- -s d q ob authors: lau, yu-lung title: sars: future research and vaccine date: - - journal: paediatr respir rev doi: . /j.prrv. . . sha: doc_id: cord_uid: s d q ob severe acute respiratory syndrome (sars) is a new infectious disease of the st century that has pandemic potential. a novel coronavirus (cov) was identified as its aetiological agent and its genome was sequenced within months of the world health organisation issuing a global threat on sars. the high morbidity and mortality of this potentially pandemic infection demands a rapid research response to develop effective antiviral treatment and vaccine. this will depend on understanding the pathogenesis and immune response to sars cov. further understanding of the ecology of sars cov in human and animals will help prevent future cross species transmission. likewise for the super-spreading events, clarification of the underlying reasons will be important to prevent a large scale outbreak of sars. lastly it is of utmost importance that international research collaboration should be strengthened to deal with sars and any other emerging infectious disease that can seriously threaten our future. severe acute respiratory syndrome (sars), a newly emerged infectious disease of humans in the st century, appeared in guangdong province in southern china in november and spread to countries on five continents along international air travel routes, causing large scale outbreaks in hong kong, singapore and toronto in early . the world health organisation (who) issued a global alert on sars on march . with support of the who, authorities in affected regions implemented epidemiologic surveillance and adherence to infection-control procedures, including patient isolation and quarantine for contacts, which helped to contain the sars outbreak by mid-july . however, there were a total of sars cases and associated deaths. the aetiologic agent of sars was identified as a coronavirus (cov) [ ] [ ] [ ] [ ] and the genome sequence established it as a novel member of the family , . this novel cov has satisfied koch's postulates for causation by its consistent isolation from sars patients, viral isolation, reproduction of disease in non-human primates after inoculation and the presence of specific antibody response against the virus in both patients and experimentally infected primates . all of these remarkable findings were accomplished within a few months of the issue of the who global alert on sars, testifying to the rapid response of the collaboration of the international research effort to deal with such emerging pandemics. the origin of sars remains uncertain despite closely related covs that were recovered from civet cats and other animals in guangdong province, suggesting the sars-cov could have originated from such animals and implicating sars as a zoonotic disease. other members of the cov family can cause fatal diseases in poultry and laboratory rodents. the two previously known human covs cause only mild upper respiratory infections. despite the / sars epidemic being eventually controlled by case isolation, there is still neither an effective treatment for sars nor an efficacious vaccine to prevent infection. the high morbidity and mortality of sars, as well as the potential of reemergence, make it paramount to focus on future research to develop effective means to treat and prevent the disease should it reappear. indeed, sporadic reemergence of cases have been reported in guangdong province as well as from research laboratories summary severe acute respiratory syndrome (sars) is a new infectious disease of the st century that has pandemic potential. a novel coronavirus (cov) was identified as its aetiological agent and its genome was sequenced within months of the world health organisation issuing a global threat on sars. the high morbidity and mortality of this potentially pandemic infection demands a rapid research response to develop effective antiviral treatment and vaccine. this will depend on understanding the pathogenesis and immune response to sars cov. further understanding of the ecology of sars cov in human and animals will help prevent future cross species transmission. likewise for the super-spreading events, clarification of the underlying reasons will be important to prevent a large scale outbreak of sars. lastly it is of utmost importance that international research collaboration should be strengthened to deal with sars and any other emerging infectious disease that can seriously threaten our future. ß elsevier ltd. all rights reserved. in singapore, taiwan and beijing since the conclusion of the sars epidemic. on may , the national institute of allergy and infectious diseases (niaid) convened a colloquium entitled 'sars: developing a research response' on the national institute of health campus, with over participants that included physicians, scientists and policymakers from the united states, china, canada, europe and elsewhere, to coordinate a robust research response to the sars threat in each of the following five areas: ( ) clinical research; ( ) epidemiology; ( ) diagnostics; ( ) therapeutics; and ( ) vaccines. one year has gone by since this colloquium, with intense research activities conducted in many laboratories worldwide addressing those issues raised. despite over published papers in pubmed on various aspects of sars, many questions are still awaiting answers. this review will attempt to highlight some of the important issues that have only been partially addressed, pointing to areas of clinical interest rather than covering all aspects comprehensively. careful description of the clinical manifestations of sars, correlating with virologic and immunologic parameters, has suggested that there is an initial viral replicative phase of about days, possibly followed by an immunopathological phase. it has been hypothesised that the immunopathological response is triggered by the viral antigens, hence the most strategic treatment is to stop the viral replication at the initial phase of the disease so that the peak viral load and the subsequent damage is minimised. therefore, an effective antiviral treatment has a window of opportunity of several days after disease onset to modify the peak viral load, thereby, in theory, decreasing the morbidity and mortality. at present, the treatment is largely empirical and therefore controversial, ranging from supportive therapy without intervention to a combination of antivirals and steroids. a recent report on an open trial of a combination of a protease inhibitor and a nucleoside analogue against a historical control suggests a favourable clinical response using lopinavir/ritonavir and ribavirin. however, controversy remains as no controlled studies have been undertaken, and there is a need to set up international clinical trial networks to develop and perform clinical protocols should sars reappear. the future research priorities in sars therapeutics should focus on antiviral drug screening as no clinically proven candidates exist. a high-throughput screening assay has to be developed and applied to the existing drug candidate libraries. some in vitro activity against sars cov was observed with certain preparations of interferona as well as with glycyrrhizin. , in vivo activity has also been demonstrated with interferon-a in a non-human primate model but there have been no studies in humans. molecular studies of sars cov and other coronaviruses suggest several potential molecular targets for antiviral drugs. these include viral binding, fusion and other activities mediated by the glycoprotein spike on the cov surface, as well as the rna-dependent rna polymerase and the cysteine protease. the identification of angiotensin-converting enzyme (ace ) as a functional receptor for the sars cov has opened up possibilities of treatment strategies such as interference of binding or fusion of sars cov with target cells. , the sars cov has surface spike (s) proteins which contain the s and s domains; the ace , which is a metallopeptidase, binds the s domain of the sars cov s protein efficiently and anti-ace has been shown to block sars cov replication in vitro using vero e cells as an experimental model. a number of antibodies, peptides and small compounds can bind to ace and it is possible that some of these can be useful in the treatment of sars, either by blocking the s-protein-binding site or by inducing a conformation in ace that is not favourable to binding or fusion. a soluble form of the ace may slow viral replication in an infected individual. recently, a human monoclonal antibody r, against s domain of the sars cov, has been developed that can potently neutralise sars cov infection and efficiently inhibit syncytia formation through blocking of receptor binding. this monoclonal antibody can be used as an immediate treatment strategy for emergency prophylaxis and treatment of sars, while the more time-consuming development of vaccines and new drugs is underway. during the winter and spring months, the diagnostic challenges of differentiating sars from other respiratory infections can be great, making a rapid, simple and accurate diagnostic assay for sars cov an imperative public health tool to control any future sars outbreak as well as the initiation of treatment protocol. the first generation pcr assays for sars cov were far from satisfactory but realtime pcr, coupled with an improved rna extraction method, has allowed detection of viral rna in nasopharyngeal aspirates with % specificity. knowing where in the body the virus can be found at different stages of the disease might lead to new diagnostic strategies. for example, the report that sars cov can replicate in peripheral blood cells in sars patients shortly after onset indicates that blood could be used as an appropriate clinical specimen for diagnosis. , nevertheless, availability of a highly sensitive and specific direct detection method of sars cov in readily obtained clinical specimens such as nasopharyngeal secretions will be most ideal, obviating the need of the laborious steps of rna extraction. serologic assays are not useful for early diagnosis as igg antibodies do not appear for - days after onset of symptoms. it has been stated that igm antibodies typically appear earlier, but detection of igm antibodies does not appear to permit earlier diagnosis. , since a few sars patients have had late seroconversion, it is best to test the convalescent serum collected at least days and preferably days after onset of symptoms, to rule out sars. at present, the most widely used methods for detection of antibodies against sars cov are indirect immunofluorescence assay and elisa with cell-culture extract, which are difficult to standardise. therefore, recombinant-antigenbased elisa assays are being developed using highly immunogenic nucleocapsid protein of sars cov, which can be used for a large scale epidemiological study of seroprevalence. the characterisation of immune responses to the sars cov, including the impact of sars on immune function and any immunopathological responses the virus may trigger, is crucial in helping the development of new therapeutic strategies and vaccines. there is a rapid and generalised lymphopaenia in patients with sars during the acute phase of infection, which is in distinct contrast to the proliferative response seen in hiv-, cmv-or ebv-infected patients. in patients who recovered from sars, an equally rapid and dramatic restoration of t cell, b cell and nk cell counts was seen in peripheral blood. the mechanism through which the sars cov precipitates such lymphopaenia so rapidly is unclear but could be related to immunologic trigger of apoptosis of uninfected lymphocytes. elucidation of the underlying mechanism may help design treatment strategies. the initial increase in viral load in the first days of disease also suggests that the role of innate immunity as a first-line defence is important and may influence the subsequent disease progression. it is important to identify the host innate immune response genetics that are predictors of disease susceptibility and progression. this would help to improve prognostic capabilities and allow identification of patients likely to benefit from aggressive interventions. such information is also of value in the development of entry criteria for interventional studies. since immune response might play a role in sars pathogenesis, one must be cautious of the possibility of enhancement of the disease in immunised subjects. this has indeed occurred for experimental vaccine directed against feline infectious peritonitis virus, which is also a coronavirus. therefore caution has been urged on developing sars vaccines. however, given the urgent need for a safe and effective sars vaccine, multiple strategies for vaccine development have been pursued simultaneously. at least candidate sars vaccines are at different phases of development. [ ] [ ] [ ] [ ] [ ] these include the development of live-attenuated and inactivated virus vaccine, in addition to other strategies that elicit strong t cell responses, such as dna-based vaccine and engineered adenovirus vectors, as well as those that elicit production of neutralising antibodies. subunit vaccines based on s protein fragments and peptides are also being developed. sars cov s protein, expressed by attenuated vaccinia virus, has been shown to immunise mice protectively. similarly, a dna vaccine encoding the sars cov s protein can induce t cell and neutralising antibody responses, as well as protective immunity, in a mouse model. viral replication can be reduced by more than six orders of magnitude in the lungs of mice vaccinated with these s plasmid dna expression vectors and protection is mediated by a humoral but not a t-cell-dependent immune mechanism. all the experimental testing of candidate sars vaccines would require an animal model that reproduces sars symptoms and pathology as in humans. however, no animal model described to date can reliably mimic the respiratory symptoms seen in humans with sars. even the macaques model, used initially to fulfil the last of koch's postulates in confirming sars cov as the etiologic agent, does not always reproduce the sars-like symptoms when infected with sars cov. this could be due to macaques not being inbred like mice. finding a consistent sars animal model which can be used for testing potential drugs and vaccines is certainly a top research priority. monkey, mouse and ferret have all been used as animal models for answering different research questions. the origin of sars cov and its ecological relationship with other animal cov will certainly be an important topic of research, as the animal reservoir of such cov will be the constant potential source of another sars outbreak. understanding of the ecology will help to implement measures to minimise transmission across species taking place again. nevertheless, the recent sporadic reemergence of sars was due to lapses of security in research laboratories in handling sars cov, testifying to the urgent need to maintain and monitor laboratory safety in research institutes that deal with such pathogens. another mystery in the sars epidemiology is the so-called super-spreading events, such as the community outbreak in the amoy garden housing complex of hong kong, which affected more than residents of this private housing estate. tentative evidence of airborne transmission of the sars cov has been suggested by an epidemiologic analysis of this outbreak, coupled with airflow simulations and experimental studies. better understanding of these super-spreading events will be crucial in preventing similar events from happening again. sporadic cases of sars may continue to appear but if vigorous public health measures can be maintained another major outbreak is unlikely, making it impossible to perform controlled clinical studies of either candidate antivirals or vaccines for sars cov. nevertheless, the global community needs to be prepared for such emerging and reemerging infectious diseases if the human race is to have a future. understand the pathogenesis and immune responses to sars cov. develop effective antiviral therapeutics and efficacious vaccines. develop rapid and accurate diagnostic tests for early diagnosis of sars cov infection. clarify the ecology of human and animal sars cov to prevent cross species transmission. understand the reasons for super-spreading events. the severe acute respiratory syndrome coronavirus as a possible cause of severe acute respiratory syndrome identification of a novel coronavirus in patients with severe acute respiratory syndrome a novel coronavirus associated with severe acute respiratory syndrome newly discovered coronavirus as the primary cause of severe acute respiratory syndrome characterization of a novel coronavirus associated with severe acute respiratory syndrome the genome sequence of the sars-associated coronavirus koch's postulates fulfilled for sars virus isolation and characterization of viruses related to the sars coronavirus from animals in southern china sars coronavirus: a new challenge for prevention and therapy sars research working group co-chairs.severe acute respiratory syndrome: developing a research response clinical progression and viral load in a community outbreak of coronavirus-associated sars pneumonia: a prospective study role of lopinavir/ritonavir in the treatment of sars: initial virological and clinical findings severe acute respiratory syndromerelated coronavirus is inhibited by interferon-alpha glycyrrhizin an active component of liquorice roots, and replication of sars-associated coronavirus pegylated interferon-a protects type pneumocytes against sars coronavirus infection in macaques angiotensin-converting enzyme is a functional receptor for the sars coronavirus expression cloning of functional receptor used by sars coronavirus novel peptide inhibitors of angiotensin-converting enzyme potent neutralization of severe acute respiratory syndrome (sars) coronavirus by a human mab to s protein that blocks receptor association early diagnosis of sars coronavirus infection by real time rt-pcr detection of sars coronavirus in patients with suspected sars sars-coronavirus replicates in mononuclear cells of peripheral blood (pbmcs) from sars patients quantitative analysis and prognostic implication of sars coronavirus rna in the plasma and serum of patients with severe acute respiratory syndrome relative rates of non-pneumonic sars coronavirus infection and sars coronavirus pneumonia significant changes of peripheral t lymphocyte subsets in patients with severe acute respiratory syndrome caution urged on sars vaccines generation and characterization of dna vaccines targeting the nucleocapsid protein of severe acute respiratory syndrome coronavirus effects of a sars-associated coronavirus vaccine in monkeys severe acute respiratory syndrome coronavirus spike protein expressed by attenuated vaccinia virus protectively immunizes mice a dna vaccine induces sars coronavirus neutralization and protective immunity in mice evidence of airborne transmission of the severe acute respiratory syndrome virus further reading key: cord- -gyq cpc authors: wang, chang‐zheng; hu, shun‐lin; wang, lin; li, min; li, huan‐tian title: early risk factors of the exacerbation of coronavirus disease pneumonia date: - - journal: j med virol doi: . /jmv. sha: doc_id: cord_uid: gyq cpc the purpose of this study was to investigate the early risk factors for the exacerbation of coronavirus disease (covid‐ ) pneumonia. restrospective analysis of clinical data of patients infected with sars‐cov‐ , including gender, age, comorbidities, symptoms, blood routine, clotting profile, biochemical examination, albumin, myocardial enzyme profile, inflammatory markers, and chest ct. all laboratory examination were measured within first hours after admission, and chest ct were performed before admission. ( . %) patients had a history of exposure to huanan seafood market in wuhan. fever and dry cough accounted for the highest percentage of all symptoms. male covid‐ patients were more likely to develop severe pneumonia. patients with severe and critical conditions are older and have higher rates of hypertension (p= . ) and coronary heart disease (p= . ). all severe and critical patients infected with sars‐cov‐ showed bilateral lung involvement and have more multiple lobes involvement than common patients (p< . ). severe and critical patients showed higher wbc count (p= . ), neu count (p= . ), neu% (p= . ), pct (p= . ), crp (p= . ), pt (p= . ), d‐dimer (p= . ), ast (p= . ), and lower lym count (p= . ), lym% (p= . ), alb (p< . ). logistic regression analysis showed neu count is a independent risk factor for deterioration, with the threshold of . × ( )·l(‐ ). we concluded that the laboratory independent risk factor for the progression of covid‐ pneumonia is neu count. in addition, covid‐ patients with bilateral lung involvement or multiple lobes involvement should be taken seriously and actively treated to prevent deterioration of the disease. this article is protected by copyright. all rights reserved. coronavirus disease (covid- ) is an emerging infectious disease with significant morbidity and mortality, which is caused by severe acute respiratory syndrome coronavirus (sars-cov- ) . in january , the who emergency committee announced "a public health emergency of international concern". as of march , , sars-cov- has swept through countries worldwide, causing , infections and , deaths. at present, the virus is spreading rapidly in many countries, with outbreaks in italy, iran, spain, germany and south korea causing the most concern. the main manifestations of the disease are fever, dry cough and fatigue . a small number of patients have symptoms such as stuffy nose, runny nose, sore throat, myalgia and diarrhea . severe patients often develop dyspnea and/or hypoxemia one week after onset, and may even rapidly progress to acute respiratory distress syndrome (ards), septic shock, hard-to-correct metabolic acidosis, bleeding and coagulation dysfunction, and multiple organ failure . to date, the worldwide mortality rate for patients infected with sars-cov- is about . %. early identification of patients with possible deterioration of the disease is particularly important for controlling the proportion of severe patients and reduce the mortality. therefore in this study, we aimed to analyze the early clinical features of covid- and investigate the early risk factors for exacerbation. the clinical data of patients diagnosed by covid- were collected, including common patients in department of infectious diseases from january , to february , and severe and critical patients in intensive care unit (icu) from january , to february , . all patients were collected continuously. the collection and use of relevant information can fully protect the privacy of patients and the study conformed to ethical norms. the diagnosis of covid- referred to the "diagnosis and treatment of novel coronavirus pneumonia (trial version )" issued by the national health commission of the people's republic of china . the confirmed patient should satisfy sputum, pharynx swab or lower respiratory tract secretions and other specimens to receive real-time qpcr to show positive viral nucleic acid . influenza virus, adenovirus, respiratory syncytial virus and other known viral infections and mycoplasma pneumoniae infection were excluded in all patients. at present, the clinical classification of the disease is: common type: with fever, respiratory symptoms, imaging manifestations of pneumonia; severe type, one of the following: . respiratory distress, rr≥ times/min; . in resting state, oxygen saturation is less than %; . partial arterial oxygen pressure (pao )/oxygen absorption concentration (fio )≤ mmhg ( mmhg = . kpa); . chest imaging showed that the lesion significantly progressed to > % within to hours; critical type: one of the following: . respiratory failure, requiring mechanical ventilation; . shock; . combined with other organ failure, intensive care unit is required . all patients underwent ct examination before admission. ge discovery and siemens somatom sensation were used for spiral ct scanning. the patient was supine and scanned at the end of inspiration with a conventional dose scan ranging from the tip of the lung to the base of the lung. the thin-slice ct image was evaluated by two radiologists who did not know the results of nucleic acid. were measured for all enrolled patients within first hours after admission. the normal distribution measurement data were presented as the mean ± standard deviation (sd), and student's t-test were employed to compare the mean of two groups. the non-normal distribution measurement data were described as median (quartile spacing), and the mann-whitney rank-sum test was used for the nonparametric analysis. the counting data were compared by chi-square test or fisher's exact test. multiple logistic regression was used for the regression analysis. statistical significance was set as p< . . spss . was used for statistical analysis. a total of patients were included in this study, including common patients and severe and critical patients. among them, ( . %) were males and ( . %) were females, and males were more likely to develop severe pneumonia ( vs , p= . ). in addition, severe and critical patients were older than common patients ( . ± . vs . ± . , p= . ). ( . %) patients had a history of exposure to huanan seafood market in wuhan. the most common symptoms at onset were fever ( . %) and dry cough ( . %); while the less common symptoms were fatigue or muscle aches ( . %), expectorant ( . %), dizziness or headeche ( . %), dyspnea ( . %), and diarrhea ( . %). there were no statistically significant differences in symptoms between the two groups. in addition, in terms of comorbidities, compared with common patients, the proportion of severe and critical patients with hypertension ( . % vs . %, p= . ) and coronary heart disease ( . % vs . %, p= . ) were higher. however, there were no statistically significant differences in other comorbidities and vital signs between the two groups (table ) . all patients presented abnormal chest ct findings before admission. of the patients, ( . %) had bilateral involvement. all severe and critical patients showed bilateral lung involvement. in addition, the rate of bilateral involvement was higher in severe and critical patients than in common patients ( . % vs . %, p= . ). typical features of chest ct images were ground-glass changes, grid-form shadow and paving stone sign. other manifestations included multiple patches, infiltration and even consolidation. the distribution characteristics of lesions in all included patients were shown in table . the lesions involved one lobe ( vs cases), two lobes ( vs cases), three lobes ( vs cases), four lobes ( vs cases) and bilateral whole lung ( vs case). apparently severe and critical patients have more lobes damage than common patients (p＜ . ). the results of blood routine showed that wbc count ( . × ·l - vs . × ·l - , p= . ), neu% ( . % vs . %, p= . ), and neu count ( . × ·l - vs . × ·l - , p= . ) in severe and critical patients were significantly higher than those in common patients. furthermore, the proportion of wbc count (p= . ) and neu count (p= . ) above the normal upper limit was significantly higher in severe and critical patients. however on the contrary, lym% ( . % vs %, p= . ) and lym count ( . × ·l - vs . × ·l - , p= . ) were lower in severe and critical patients. besides, the proportion of severely ill patients with lym counts less than × ·l - is lower than that of common patients ( . % vs . %, p= . ). there were no significant difference in hgb ( . g/l vs g/l, p= . ) and plt count ( × ·l - vs × ·l - , p= . ) between the two groups. in terms of clotting profile, pt ( . s vs . s, p= . ) and d-dimer ( ug·ml - vs ug·ml - , p= . ) were significantly elevated in severe and critical patients, while the level of aptt ( . s vs . s, p= . ) did not differ obviously. in addition, crp ( . mg·l - vs . mg·l - , p= . ) and pct ( . ng·ml - vs . ng·ml - , p= . ) were higher in severe and critical patients. for biochemical indicators, the values of ast ( iu·l - vs iu·l - , p= . ) in severe and critical patients were significantly higher than those in common patients, and iu·l - may be the threshold. besides, the level of alb ( g·l - vs . g·l - , p＜ . ) was significantly lower in common patients. although the value of creatinine in the severe group was significantly higher than that in the common group (p= . ), there was no significant difference when threshold was set at umol·l - (p= . ). lastly, there were no significant differences in other biochemical indicators and myocardial injury markers (table ) . multivariate logistic regression analysis between neu, lym, pct, crp, alb, ast and severe and critical covid- found that neu is a independent risk factor for deterioration, with the threshold of . × ·l - (p= . )( table ). since sars-cov- was found in wuhan city, hubei province in december , it has spread rapidly all over the country and the world . the virus transmission through fecal-oral, prolonged exposure in closed field, aerosol transmission in high concentration environment, and indirect transmission of conjunctiva are not excluded . the clinical features are fever, dry cough, persistent high fever in severe cases, and acute respiratory distress syndrome, sepsis, bleeding and coagulation dysfunction in critical cases . the detection of viral nucleic acid in respiratory tract or blood samples is the main method for the diagnosis of covid- , which can be combined with epidemiology and imaging to improve the accuracy . the treatment principle of the disease is, on the basis of symptomatic treatment, to actively prevent and cure complications, to prevent secondary infection, and to provide timely organ function support . there are no specific drugs for this infected disease, while some antiviral drugs that may be effective are recommended in china, such as interferon-alpha, lopiravir/ritonavir, ribavirin, chloroquine phosphate and abidor in this study, we collected a cohort of patients with viral rna-confirmed sars-cov- infection. the ratio of male to female was : , indicating that men and women are generally susceptible to sars-cov- . however compared with female patients, male patients accounted for a higher proportion of severe and critical cases. older patients were at higher risk for disease progression. in addition, clinical data suggested that severe and critical patients often incorporated more basic diseases, such as hypertension and coronary heart disease. this suggests that covid- patients with hypertension and coronary heart disease may require icu care later. chest ct before admission suggested that covid- patients with bilateral involvement or multiple lobes involvement were more likely to deteriorate to severe pneumonia. we observed greater increase in neu count, pct level, crp level and greater decrease in lymphocyte count in critically ill patients. this suggests that disease progression in covid- patients may be associated with bacterial infection and impaired cellular immunity. in addition, ast was significantly increased and albumin levels were significantly reduced in severe and critical patients, suggesting that sars-cov- virus may damage liver cells and impair the synthesis function of the liver. logistic regression analysis showed that neu is a independent risk factor for deterioration, with the threshold of . × ·l - , suggesting that combination of bacterial infections may be the most important factor in worsening the condition. in conclusion, a significant number of patients infected with sars-cov- may progress to severe illness or even death. in early stage of covid- , we can predict a patient's risk of progression by combining laboratory tests with chest ct. we concluded that the laboratory independent risk factor for the progression of covid- pneumonia is neu count, with the threshold of . × ·l - . in addition, covid- patients with bilateral lung involvement or multiple lobes involvement should be taken seriously and actively treated to prevent deterioration of the disease. the author declare no conflict of interests. writing: cw. data collection: sh, lw, and ml. data analysis: cw and hl. challenges to the system of reserve medical supplies for public health emergencies: reflections on the outbreak of the severe acute respiratory syndrome coronavirus (sars-cov- ) epidemic in china clinical characteristics of coronavirus disease in china clinical features of patients infected with novel coronavirus in wuhan national health commission of the people's republic of china. diagnosis and treatment of novel coronavirus pneumonia laboratory readiness and response for novel coronavirus ( -ncov) in expert laboratories in eu/eea countries world health organization. question and answer on coronaviruses long-term safety and durable antiretroviral activity of lopinavir/ritonavir in treatment-naï patients: -year follow up study a systematic review of therapeutic agents for the treatment of the middle east respiratory syndrome coronavirus (mers-cov) combination therapy with lopinavir/ritonavir, ribavirin and interferon-α for middle east respiratory syndrome broad-spectrum antiviral gs- inhibits both epidemic and zoonotic coronaviruses coronavirus susceptibility to the antiviral remdesivir (gs- ) is mediated by the viral polymerase and the proofreading exoribonuclease comparative therapeutic efficacy of remdesivir and combination lopinavir, ritonavir, and interferon beta against mers-cov first case of novel coronavirus in the united states convalescent plasma as a potential therapy for covid- alb present neutrophil, lymphocyte, procalcitonin, c-reactive protein, aspertate aminotransferase, albumin, respectively key: cord- -cu tl authors: dondorp, arjen m.; hoang, mai nguyen thi; mer, mervyn; dünser, martin w.; mohanty, sanjib; nakibuuka, jane; schultz, marcus j.; thwaites, c. louise; wills, bridget title: management of severe malaria and severe dengue in resource-limited settings date: - - journal: sepsis management in resource-limited settings doi: . / - - - - _ sha: doc_id: cord_uid: cu tl this chapter summarizes recommendations on important aspects of the management of patients with severe malaria and severe dengue. severe falciparum malaria requires rapid parasitological diagnosis by microscopy or rapid diagnostic test (rct) and prompt initiation of parenteral artesunate. fluid bolus therapy should be avoided in patients without hypotensive shock, and we suggest initial ( h) crystalloid fluid therapy of – ml/kg/h, which may subsequently be reduced to ml/kg/h in patients receiving additional fluids, e.g., through enteral tube feeding. in the minority of those patients presenting with hypotensive shock, we suggest fluid bolus therapy ( ml/kg) with an isotonic crystalloid and early initiation of vasopressor support. enteral feeding in non-intubated adult patients with cerebral malaria can start after h, to avoid aspiration pneumonia. there are insufficient data to suggest this in pediatric cerebral malaria. the diagnosis of severe dengue is commonly with a combined dengue antigen (ns ) and antibody rdt. no antiviral treatment is currently available. dengue shock results from capillary leakage, although hemorrhage or depression of myocardial contractility can contribute. the world health organization guidelines recommend restoration of the circulation guided by pulse pressure, capillary refill time, hematocrit, and urine output. large (> ml/kg) rapid (< min) fluid boluses should be avoided, but prompt fluid administration with crystalloids is essential and should be restricted as soon as the critical phase is over to avoid pulmonary edema. corticosteroids are not recommended, neither is platelet transfusion for thrombocytopenia in the absence of active bleeding or other risk factors. sepsis in resource-limited settings will often have different etiologies to those in western settings, including severe malaria, severe dengue, viral hemorrhagic fevers, melioidosis, typhus, and leptospirosis. the surviving sepsis campaign (ssc) guidelines [ ] are mainly based on evidence from studies on bacterial sepsis. these guidelines are widely applicable, but there are also exceptions. we here focus on disease-specific recommendations for the management of severe falciparum malaria and severe dengue. an international team with extensive practical experience in resource-limited intensive care units (icus) identified key questions concerning the ssc's management recommendations on these diseases. pertinent evidence from resource-limited settings was evaluated using the grading of recommendations assessment, development, and evaluation (grade) tools. severe falciparum malaria is a multiorgan disease caused by plasmodium falciparum transmitted by anopheles mosquitoes. the highest transmission and disease burden is in sub-saharan africa, where severe malaria is largely a pediatric disease, as older children and adults become partly immune. in asia and south america, all age groups may be affected. independent of age, the presenting symptoms with the strongest prognostic significance are coma (cerebral malaria), metabolic (lactic) acidosis, and renal dysfunction. acute respiratory distress syndrome is a common and often fatal complication in adult patients with severe malaria. hypotension occurs infrequently (~ % of cases), and should raise a suspicion of concomitant bacterial sepsis. one of the main pathophysiologic differences of severe falciparum malaria compared to bacterial sepsis is microcirculatory impairment caused by sequestration of parasite-infected erythrocytes, red cell rigidity, and red cell clumping. severe dengue is caused by dengue virus transmitted by aedes mosquitoes. approximately - % of patients will develop severe manifestations. the defining feature is a vasculopathy with increased capillary permeability, causing plasma leakage, reduced intravascular volume, and if severe life-threatening hypovolemic shock [ ] . this "critical phase" typically starts during the period of defervescence and lasts for approximately h. bleeding complications and organ involvement of the brain, liver, kidney, and heart may be additional features and occur more frequently in adult cases [ ] . recommendations and suggestions are summarized in table . . severe malaria is an old disease, and historically, the guidance for fluid management has been to "keep them dry." this approach was subsequently challenged when it was recognized that severe malaria is a severe sepsis syndrome with signs of tissue hypoperfusion and thus might benefit from fluid bolus therapy. the ssc guidelines recommend in patients with sepsis-induced tissue hypoperfusion and suspicion of hypovolemia an initial fluid challenge of minimal ml/kg of crystalloids, to be completed within h, of which a portion may be albumin equivalent; this applies to patients with hypotension or a plasma lactate ≥ mmol/l [ ] . it was shown by various techniques that both children and adults with severe falciparum malaria are intravascular dehydrated [ ] [ ] [ ] although this was debated by some [ ] . small trials in african children with severe malaria suggested a benefit from fluid bolus therapy, in particular with albumin [ ] [ ] [ ] [ ] , as recently reviewed [ ] . however, a subsequent large trial on fluid bolus therapy in african children with severe infections and compensated shock, of which % had falciparum malaria, showed overall a % increase in mortality with fluid bolus therapy ( ml/kg or ml/kg with either saline or albumin). in the children with severe p. falciparum malaria, mortality in the bolus groups was % higher (rr . [ . - . ]) than without fluid bolus therapy [ ] . in the same study, febrile patients with hypotensive ("decompensated") shock were randomized between and ml/kg fluid bolus therapy with either saline or albumin; % of the children ( of ) in the albumin bolus group and % ( of ) in the saline-bolus group died (p = . ). in asian studies in adult severe malaria, rapid fluid resuscitation did not improve metabolic we recommend not to use fluid bolus therapy in normotensive patients with severe falciparum malaria ( a). we suggest that patients receive maintenance isotonic crystalloid fluid therapy ( - ml/kg/h), which may subsequently be reduced to ml/kg/h in patients receiving additional fluids, e.g., through enteral tube feeding ( d). we suggest that in patients with hypotensive shock, fluid bolus therapy ( ml/kg) with isotonic crystalloids be commenced (ungraded) and, if available, early initiation of vasopressor medication (ungraded) timing of enteral feeding in cerebral malaria we suggest initiating enteral feeding in non-intubated adult patients with cerebral malaria after h, in order to limit the possibility of aspiration pneumonia ( b). there are insufficient data to make this recommendation for children with cerebral malaria permissive hypercapnia in ventilated cerebral malaria we suggest not to use a strategy of permissive hypercapnia to achieve ventilation with low tidal volumes in patients with cerebral malaria, because of the high incidence of brain swelling in these patients (ungraded) fluid management in severe dengue we recommend that fluid resuscitation in severe dengue is executed promptly and guided by pulse pressure, capillary refill time, hematocrit, and urine output according to who guidelines and that fluid therapy should be restricted as soon as the critical phase of the disease is over to avoid pulmonary edema ( c). we recommend that rapid administration of large fluid boluses should be avoided, unless the patient is hypotensive ( d). we recommend that in dengue patients with compensated shock, colloid fluids are not used ( a) use of corticosteroids in severe dengue we recommend not to use corticosteroids in the treatment of severe dengue ( b) we recommend not to use prophylactic platelet transfusion for thrombocytopenia in the absence of active bleeding complications or other risk factors (uncontrolled arterial hypertension, recent stroke, head trauma or surgery, continuation of an anticoagulant treatment, existing hemorrhagic diathesis) ( b) acidosis [ , ] , and transpulmonary thermodilution-guided rapid fluid resuscitation resulted in pulmonary edema in / ( %) patients [ ] . one observational study showed no deterioration in renal function or plasma lactate with maintenance fluid therapy between . and . ml/kg/h [ ] . a recent systematic review concluded that fluid bolus therapy with either crystalloid or albumen is not beneficial in severe falciparum malaria [ ] . we recommend not to use fluid bolus therapy in normotensive patients with severe falciparum malaria ( a). we suggest not to use colloid therapy, including albumin % ( c). in normotensive patients, we suggest initial crystalloid fluid therapy of - ml/kg/h ( d). in patients receiving enteral fluids, e.g., through enteral tube feeding, we suggest that this can be reduced to ml/kg/h ( d). this is slightly more conservative than the recommendation in the management guidelines for severe malaria issued by the world health organization, recommending - ml/ kg/h [ ] . there are no data on the benefit of balanced fluids over normal saline. we suggest fluid bolus therapy ( ml/kg) with an isotonic crystalline in patients with hypotensive shock and, if available, early start of vasopressive medication (ungraded). hypotensive shock in a patient with severe malaria could indicate concomitant bacterial sepsis, and be evaluated and treated accordingly. the ssc guidelines suggest administering oral or enteral (if necessary) feeds, as tolerated, rather than either complete fasting or provision of only intravenous glucose within the first h after a diagnosis of severe sepsis/septic shock (grade c) [ ] . early enteral feeding is thought to preserve gut integrity and function, maintain bile secretion and secretory iga, maintain gut-associated lymphoid tissue (galt) resulting in reduced translocation, improve splanchnic blood flow, and act prophylactically against stress ulceration. in patients with severe malaria, malnutrition is common, as is concomitant invasive bacterial infection [ ] . therefore, the recommendation for early start of enteral feeding seems valid for patients with severe malaria, including intubated patients with cerebral malaria. however, in resourcelimited settings, endotracheal intubation of comatose patient is often not practiced, and there might be an increased risk of aspiration pneumonia. we could identify one randomized clinical trial on the timing of enteral feeding in patients with cerebral malaria [ ] . this trial in (mainly) adult bangladeshi patients with cerebral malaria who were not on mechanical ventilation, and thus had an unprotected airway, showed that early (< h) enteral feeding was associated with aspiration pneumonia in / ( %) versus / with late start after h (p = . ). this despite proper positioning of patients, and pre-feed inspection of gastric retention. no difference in the incidence of hypoglycemia was observed. we suggest starting enteral feeding in non-intubated adult patients with cerebral malaria after h ( b). there are insufficient data on pediatric patients with cerebral malaria from african settings. acute respiratory distress syndrome (ards), or pulmonary malaria, is a feared complication of severe falciparum malaria and can also complicate the course of vivax malaria [ ] . the incidence of ards in adult patients with severe malaria is estimated to % and up to % in pregnant women; ards is thought to be rare in pediatric severe malaria [ ] . to protect the lung from the damaging effects of mechanical ventilation, the ssc recommends targeting a tidal volume of ml/kg predicted body weight in patients with sepsis-induced acute respiratory distress syndrome (ards), that plateau pressures be measured in patients with ards and that the initial upper limit goal for plateau pressures in a passively inflated lung be < cm h o [ ] . there are no randomized clinical trials to evaluate this recommendation specifically for ards in the context of severe malaria. however, given the large benefit of this ventilation strategy in patients with other causes of ards, this recommendation should also be valid in severe malaria. the ssc guidelines also suggest that to facilitate the use of a lung protective ventilatory strategy, permissive hypercapnia can be used. it should be noted that availability of blood gas or endtidal co monitoring is limited in resource-limited settings, compromising its safe implementation. there are no randomized clinical trials on the use of permissive hypercapnia in mechanically ventilated patients with severe falciparum malaria. however, in cerebral malaria, brain swelling is common, caused by an increase in intracerebral blood volume including the sequestered parasitized red blood cell mass, vasogenic edema, and cytotoxic edema, and is more prominent in pediatric cases [ ] [ ] [ ] [ ] . because hypercapnia will further increase intracranial pressure, we suggest against the use of permissive hypercapnia to achieve the goal of low tidal volume ventilation in patients with cerebral malaria, as cerebral malaria is associated with brain swelling and variably increased intracranial pressure (ungraded). severe dengue can be defined as a sepsis syndrome. yet, important aspects of the pathophysiology of the circulatory changes are distinct from bacterial sepsis. dengue shock syndrome is characterized by a vasculopathy during the critical phase of the disease, with a plasma leak and hemoconcentration, causing important intravascular volume depletion [ ] . this initially leads to a compensated shock with signs of tissue hypoperfusion and a decreased pulse pressure with preserved systolic blood pressure. this can be followed by life-threatening hypotensive shock. hemorrhage, in particular from the gastrointestinal tract, and more rarely myocarditis, can contribute to circulatory shock. the onset is usually more gradual than with bacterial sepsis. management of patients with severe dengue relies largely on careful monitoring, including early recognition of vascular leakage and proper fluid replacement, combined with prompt but carefully guided volume resuscitation for patients who develop dengue shock syndrome. the ssc guidelines advocate fluid bolus therapy for patients with sepsis-induced tissue hypoperfusion and suspicion of hypovolemia [ ] , which might not be appropriate for patients with severe dengue and compensated shock. in addition, because of the prominent plasma leak, the use of colloids might be beneficial in dengue with hypotensive shock, as opposed to its use in patients with bacterial sepsis. the who guidelines for the management of patients with severe dengue distinguish patients with compensated shock from those with decompensated (hypotensive) shock [ , ] . in compensated shock, recommended initial fluid therapy is with isotonic crystalloid solutions at - ml/kg over h, which can be tapered every few hours if the patient improves guided by the pulse pressure, capillary refill time, hematocrit, and urine output. prudential fluid therapy is important throughout the disease, but in particular fluid administration should be restricted as soon as the critical phase of the disease is over to avoid pulmonary edema. in the same guidelines, it is recommended in patients with hypotensive shock, to resuscitate with crystalloid or colloid solution at ml/kg as a bolus given over min. no randomized clinical trials to support the who fluid resuscitation recommendations could be identified. fluid bolus therapy, and liberal fluid management more in general, was a risk factor for respiratory distress in a large prospective observational study in latin american and asian patients with dengue [ ] . a large prospective observational study in vietnamese children with laboratory-confirmed dengue shock syndrome practiced an initial fluid regimen of ringer's lactate solution at ml/kg over h, with colloid solutions reserved for children presenting with decompensated shock [ ] . the observed case fatality rate with this approach was / children ( . %). we recommend to follow the current who guidelines on fluid management in severe dengue/dengue shock syndrome ( c). we recommend that rapid (< min) administration of large (> ml/kg) fluid boluses should be avoided, unless the patient is hypotensive ( d). there are several randomized clinical trials comparing crystalloid with colloid fluid management for the treatment of patients with severe dengue and compensated shock. in a vietnamese trial, children with moderately severe dengue shock syndrome were randomized to fluid therapy with either ringer's lactate, % dextrose, or % hydroxyethyl starch in a : : ratio [ ] . the need for rescue resuscitation with a colloid or the proportion of children with shock recurrence (which carries a worse prognosis) was similar between treatment arms. an additional two other randomized trials did not show better outcome parameters with (more expensive) colloids over crystalloid fluids [ , ] . a quasi-randomized study from the philippines with alternate allocation of starch versus crystalloid fluids also did not show an additional benefit of colloid therapy [ ] . we recommend that in dengue patients with compensated shock, colloids are not used for initial resuscitation ( a). there is insufficient evidence to recommend fluid choice in severe dengue with hypotensive shock, but there is discussion among experts whether there is a role for colloids in severe dengue patients with hypotension, given the prominent role of capillary leak it its pathogenesis. since current evidence strongly suggests that all hydroxyethyl starches (hes) increase the risk of acute kidney injury and renal replacement therapy [ ] , we suggest not to use hes for fluid resuscitation in patients with severe dengue (ungraded). both humoral and cellular immune responses are thought to be implicated in the pathogenesis of vasculopathy, which is central in the pathogenesis of dengue shock syndrome [ ] . the risk for developing severe disease is increased in secondary heterotypic infections, in which antibody-dependent enhancement (ade) of infection and cross-reactive memory t cells are thought to play a role. these insights have led to the use of immunomodulatory therapy with corticosteroids in severe dengue infection. a cochrane review on patients with dengue shock syndrome identified four randomized or quasi-randomized trials comparing corticosteroids with no corticosteroids or placebo involving participants with dengue shock syndrome [ ] . corticosteroids did not reduce the number of deaths (rr . , % ci . - . ; participants, trials), the need for blood transfusion (rr . , . - . ; participants, trials), or the number of serious complications (convulsions and pulmonary hemorrhage, trial). the evidence was rated low quality as most studies were underpowered or lacked stringent randomization or allocation concealment. corticosteroids were administered after the onset of shock. a more recent vietnamese randomized trial in children with dengue fever evaluated early oral prednisolone therapy ( mg/kg versus . mg/kg versus placebo for days) [ ] . the use of oral prednisolone was not associated with prolongation of viremia and was considered safe. however, no reduction in the development of dengue shock syndrome or other complications was observed with early prednisolone therapy, although the trial was not sufficiently powered to assess efficacy. an additional analysis of the same trial focusing on immunological endpoints did not show an important attenuation of the host immune response with prednisolone treatment [ ] . an additional cochrane review of trials on the early use of corticosteroids in patients with dengue fever identified four studies (including the study discussed above), enrolling a total of children and adults, showing no benefit of corticosteroids regarding mortality or dengue complications, although the evidence was considered low to very low quality [ ] . with the current level of evidence, the use of corticosteroids is not recommended in the treatment of severe dengue ( b). bleeding is a feared complication of severe dengue infection. thrombocytopenia with a thrombocytopathy is invariably present in patients with severe dengue infection. however, vasculopathy is a central and important additional contributor to the bleeding risk [ ] . prophylactic transfusion of platelets is a common practice in dengue-endemic countries [ ] . platelet transfusion is not without risks, since it can cause allergic reactions and transmission of blood-borne pathogens. an open-label randomized study in patients with dengue and a platelet count below , /μl did not show decreased incidence of severe bleeding with prophylactic platelet transfusion [ ] . a non-randomized singaporean study in dengue patients with thrombocytopenia < , /μl, of whom were given prophylactic platelet transfusion, also did not show decreased bleeding episodes in the treatment group [ ] . an observational study from martinique during a dengue outbreak evaluated a conservative strategy to prophylactic platelet transfusion (only if platelets count < /μl, or in case of additional risk factors). a poor correlation between thrombocytopenia and the occurrence of severe bleeding during admission was observed, and the followed conservative transfusion strategy was considered safe [ ] . the who guidelines do not recommend prophylactic platelet transfusion in severe dengue. the results of the adult dengue platelet study (adept, clinicaltrials.gov: nct ), a prospective randomized open-label trial to examine the safety and efficacy of prophylactic platelet transfusion in singaporean adults with severe dengue-related thrombocytopenia (platelet count below , / μl but no bleeding), are pending. in resource-limited settings, the availability of safe pathogens vs. screened blood products can be limited, and platelet transfusion can have important cost implications, supporting restrictive use of platelet transfusion. we do not recommend platelet transfusion for thrombocytopenia in the absence of active bleeding complications or other risk factors such as the use of anticoagulants, existing hemorrhagic diathesis, uncontrolled arterial hypertension, recent stroke, head trauma or surgery ( c). in case of bleeding complications, we suggest transfusion of fresh-frozen plasma (or cryoprecipitate) and platelet concentrate (ungraded). although most recommendations in the ssc guidelines are also applicable for the management of severe malaria and severe dengue, there are some important exceptions, in particular regarding fluid management. open access this chapter is licensed under the terms of the creative commons attribution . international license (http://creativecommons.org/licenses/by/ . /), which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons license and indicate if changes were made. the images or other third party material in this chapter are included in the chapter's creative commons license, unless indicated otherwise in a credit line to the material. if material is not included in the chapter's creative commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. 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at y the new coronavirus (covid- ) was first detected in wuhan city of china in december . most patients infected with covid- had clinical presentations of dry cough, fever, dyspnea, chest pain, fatigue and malaise, pneumonia, and bilateral infiltration in chest ct. soon covid- was spread around the world and became a pandemic. now many patients around the world are suffering from this disease. patients with predisposing diseases are highly prone to covid- and manifesting severe infection especially with organ function damage such as acute respiratory distress syndrome, acute kidney injury, septic shock, ventilator-associated pneumonia, and death. till now many drugs have been considered in the treatment of covid- pneumonia, but pharmacotherapy in elderly patients and patients with pre-existing comorbidities is highly challenging. in this review, different potential drugs which have been considered in covid- treatment have been discussed in detail. also, challenges in the pharmacotherapy of covid- pneumonia in patients with the underlying disease have been considered based on pharmacokinetic and pharmacodynamic aspects of these drugs. wuhan city of china. the most common clinical signs and symptoms of these patients were dry coughs, fever, dyspnea, and bilateral infiltration in chest ct. all these patients were associated with wuhan's huanan seafood wholesale market which sells fish and other live animals such as bats, poultry, snakes, etc. the causative agent, new coronavirus, was first detected through a swab sample which was drawn from the throat of these patients . this new coronavirus was subsequently named severe acute respiratory syndrome coronavirus (sars-cov- ). soon this disease, which called coronavirus disease (covid- ) by world health organization (who), promptly spreads around the world , and to date over . million cases have been diagnosed with covid- and this disease became a pandemic. on the late january covid- chinese outbreak, were introduced as a public health emergency of international concern . so although previously coronaviruses were considered as a potential cause of the common cold now we know that they are more than just the common cold! most of the covid- infected patients have an average age of s, it is slightly more predominant in the male sex, approximately % of infected patients involved with severe disease were required to intensive care unit services and % of them were required to mechanical ventilation . a published report from italian patients revealed that covid- was predominant in men ( . % in male and . % in female), most of the patients (about %) were over years old, approximately % of all confirmed patients had mild disease, % had severe disease, % were in a critical situation, and rest of the patients showed few symptoms, unspecified symptoms or were completely asymptomatic . according to recently published researches, the most common clinical presentations in covid- patients were fever in % to % of patients, dry cough in to %, and fatigue or myalgia in to % of them. other signs and symptoms which have been reported include sore throat, headache, confusion, rhinorrhea, sneezing, ageusia, anosmia, chest pain, hypoxemia, pneumonia, hemoptysis, acute cardiac injury, neurologic complications , , and gastrointestinal presentations such as nausea, vomiting, diarrhea and abdominal pain , - . patients with underlying diseases are highly prone to present with severe infection especially with organ function damage such as acute respiratory distress syndrome (ards), acute kidney injury (aki), septic shock, and ventilator-associated pneumonia (vap) , . severe covid- could cause death due to huge alveolar damage and highly progressive respiratory failure . covid- particles could spread through the respiratory mucosa and fecal-oral route . the nucleic acid of the virus was detected in stool, saliva, and respiratory specimens . this virus could be transmitted between humans during the epidemic and then pandemic of covid- . human-to-human transmission could highly accelerate the spread of this virus around the world. this type of transmission among humans is restricted to close contact and through sneezing or coughing of the infected patients who are capable to spread the respiratory droplets. then these respiratory droplets could settle in oral mucosa and lung of the people who inhaled the contaminated air near (about feet) to the infected patients , . although some researches have been focused on the airborne transmission of this virus but this route of transmission has not been approved yet and further studies are required. researches revealed that covid- could also be transmitted through asymptomatic carriers with an incubation period of to days . in order to prevent spreading of this new virus: hands should be washed frequently, the face should not be touched with unwashed hands, regular surface disinfecting is required, social distancing from people with respiratory symptoms is essential, sneezing or coughing should be done into the elbow or soft tissue if available . based on the published reports, in most of the patients with covid- , the absolute value of lymphocytes was reduced, which indicated that this novel coronavirus (covid- ) acts more on lymphocytes especially t lymphocytes, just similar to sars coronavirus . it seems that covid- could induce a cytokine storm and activate immune responses which could be appeared as changes in the number of white blood cells and immune cells especially lymphocytes, the clinical outcome of such events would be respiratory distress syndrome, septic shock and finally end-organ damage . covid- could also affect the liver which could be presented as hypoproteinemia, elevated aminotransferases, and prolonged prothrombin time. hepatotoxicity could be attributed to the higher expression of angiotensin converting enzyme ii (ace ) in cholangiocytes, ace could act as an entry receptor for covid- . so it seems that this new virus can directly damage the intrahepatic bile ducts . pathological findings of a liver biopsy from a patient with covid- showed moderate micro-vesicular steatosis and also a mild portal and lobular activity which could be a result of direct sars-cov- liver damage or antiviral drug-induced hepatotoxicity . almost all covid- patients had abnormal lung ct when diagnosed. according to the recently published article, an average of . ± . segments were involved in patients and the number of involved lung segments was significantly higher in symptomatic patients group in comparison to asymptomatic ones. ct findings revealed that affected covid- patients could present as bilateral lung involvement, peripheral distribution, or diffuse distribution. the most common presentation in chest ct was ground-glass opacity pattern, consolidation, and ill-defined margins , . laboratory confirmation of steatosis could be performed by real-time reverse-transcription polymerase chain reaction (rrt-pcr) , . according to who approved laboratory testing for covid- diagnosis is based on nucleic acid amplification test (naat) such as rrt-pcr which could detect the sequence of the rna of covid- . governments need to appreciate people to obey social distancing and isolation. in some situations, quarantine of major cities is also suggestive. global health governance should apply the least restrictive measures for people according to the international health regulations (ihr) , . scientists around the world are looking for drugs that could be beneficial in covid- treatment. many drugs have been studied that are listed in table with the usual dosage ranges in adults and pediatrics. the latest guidelines for the prevention, diagnosis, and treatment of novel coronavirus-induced pneumonia, have been suggested antiviral agents containing: interferon alpha (ifn-α), lopinavir/ritonavir, chloroquine phosphate, ribavirin, and arbidol as potential options in covid- treatment . drugs that have been considered in covid- management have been classified as investigational drugs, drugs under clinical trials, and drugs that have received u.s. food and drug administration (fda) as shown in table . . although many previous studies emphasized the potential therapeutic effects of these drugs in covid- management, unfortunately some recent publications reported that the efficacy of chloroquine/hydroxychloroquine in covid- management is not consistent. also, their safety is still remaining a major concern for physicians and pharmacists, since chloroquine/hydroxychloroquine could cause qt prolongation and arrhythmia. since the possible risks of these drugs could overweigh their potential benefits and efficacy, united states food and drug administration (fda) no longer recommended these two drugs as potential options for covid- management . results of a systematic review on the efficacy of hydroxychloroquine or chloroquine on the prevention or treatment of covid- revealed that the available evidences on their benefits and risks are weak and controversial . results of another systematic review and meta-analysis on randomized clinical trials on administration of hydroxychloroquine in covid- management revealed that hydroxychloroquine administration (case group) was significantly associated with higher incidence of total adverse effects in comparison to placebo or no treatment (control group) in overall population of patients with covid- . so, the recruitment of chloroquine/hydroxychloroquine in covid- management is still controversial and further larger multi-center randomized clinical trials are required to evaluate their efficacy, safety, risk-benefit ratio, dose and duration of individualized pharmacotherapy. also, close patient monitoring, especially cardiac, ocular, and neurotoxicity assessments, are required and strongly recommended during drug administration . recently published studies revealed that chloroquine could highly reduce covid- replication . chloroquine is a weak base that could be entrapped in organelles that are membrane-enclosed and have low-ph, so interfering with their acidification process. therefore chloroquine could inhibit ph-dependent viral fusion and replication. also, it might inhibit viral assembly in endoplasmic reticulum-golgi intermediate like structures . another possible antiviral mechanism of chloroquine is its immunomodulatory effect through cell signaling pathways and regulating the action of proinflammatory cytokines that can enhance its antiviral effect synergistically , , . chloroquine/hydroxychloroquine could prevent from covid- -induced ards by attenuating the pro-inflammatory cytokines and receptors . hydroxychloroquine can enhance intracellular ph and avoid lysosomal activity in antigen presenting cells containing b cells, also they can avoid antigen processing and mhc-ii presentation to t cells. so, t cell activation could be reduced by the action of hydroxychloroquine. it can suppress the cytokine release syndrome (crs), which is a result of immune system over-activation, caused by covid- . according to this mechanism, hydroxychloroquine could alleviate symptoms of mild to severe covid- pneumonia . chloroquine has different adverse reactions such as cardiovascular adverse reactions . also the results of a systematic review on dermatologic adverse effects of hydroxychloroquine emphasized that the most common dermatologic reactions due to hydroxychloroquine administration were rash, sjs, toxic epidermal necrolysis (ten), pruritus, hyperpigmentation, and hair loss. these dermatologic reactions were mostly occurred after cumulative dosages of hydroxychloroquine . in overall, since hydroxychloroquine has lower tissue accumulation potential in comparison with chloroquine, it has fewer adverse drug reactions and would be better choice . contraindications in chloroquine use contains hypersensitivity to chloroquine ( -aminoquinolone compounds) and the presence of retinal or visual field changes . chloroquine and hydroxychloroquine have a narrow therapeutic index and poisoning could be occurred with cardiovascular features so it should be used with caution in patients with predisposing cardiovascular disease . long-term exposure to these drugs could induce cardiomyopathy . chloroquine in patients consuming heparin, prone the patients to risk of bleeding. also, chloroquine in patients with digitalization (using digoxin) could cause cardiac block . there is no dosage adjustment available for chloroquine or hydroxychloroquine in patients with hepatic failure but it should be used with caution . there is no dosage adjustments available for chloroquine in patients with renal failure from the manufacture's labeling but according to uptodate some clinicians use the following guideline : a) patients with gfr ≥ ml/min: no dosage adjustment is required. b) patients with gfr < ml/min: dosage should be reduced to %. c) patients with peritoneal-or hemodialysis: dosage should be reduced to %. required. there is no dosage adjustments available for hydroxychloroquine in renal failure but it should be used with caution . although some studies showed a low risk of congenital abnormalities in patients receiving chloroquine during pregnancy because of the lack of a pattern in these congenital defects, the possible association is unlikely and it seems that the benefits of its use are higher than risks . hydroxychloroquine use during pregnancy could not be accompanied by risks for fetuses, especially in low doses. but patient monitoring during pregnancy is required . in general, since chloroquine may induce severe side effects during fetal development, so hydroxychloroquine would be a better option in pregnant women with covid- infection because of its safety profile during pregnancy . according to the american academy of pediatrics, chloroquine is compatible with breastfeeding. although it could be excreted into the milk, this amount was not considered harmful for nursing infants . according to the american academy of pediatrics, hydroxychloroquine is compatible with breastfeeding. small amounts of hydroxychloroquine could be excreted to the milk, but because of the slow elimination rate and the possibility of drug accumulation and toxicity, breastfeeding during hydroxychloroquine therapy should be done with caution . umifenovir is a broad-spectrum antiviral agent which is effective against enveloped and nonenveloped rna or dna viruses especially against influenza virus type a and b, respiratory syncytial virus, sars-cov, adenovirus, hepatitis c virus (hcv), etc. umifenovir was first developed in russia and now its usage is more common in russia and china and is less common in western countries. its possible antiviral mechanism is the inhibition of viral fusion with targeted membrane and preventing from the viral entrance to targeted cells . umifenovir has a dual pharmacologic action: first is its beneficial effect on respiratory viruses such as the covid- virus and the second is its immune-stimulating function which can activate serum interferon and phagocytes. since , umifenovir was patented for its beneficial effect in the treatment of severe acute respiratory distress (sars) coronavirus-induced atypical pneumonia . results revealed that umifenovir can induce direct viricidal effect so it would be a promising direct-acting antiviral (daa) agent. umifenovir could affect critical stages of viral life cycles such as cell attachment, cell internalization, viral replication, assembly, and budding so it also would be a promising host targeting agent (hta). its dual pharmacologic function is related to its potential interaction with both cell membranes and with cellular and viral lipids and proteins . the most important adverse reactions associated with umifenovir are diarrhea, nausea, vomiting, dizziness, confusion, and elevated liver enzymes (serum aminotransferases) . umifenovir is an indole derivative with poor water solubility which could affect its bioavailability and pharmacokinetics. after oral administration of umifenovir, it could rapidly distribute to organs and tissues, maximum plasma concentration (c max ) was achieved after to . hours. in the russian population, it had elimination half-life (t ½) of to hours, but t ½ was shorter in the chinese population. after multiple-dose administration of umifenovir, little drug accumulation could be predictable. the main site of drug metabolization is the liver. umifenovir could undergo several metabolism pathways such as oxidation at the s site, ndemethylation, glucuronidation, and conjugation at -hydroxy moiety. the potential antiviral effects of umifenovir metabolites are unknown until now . since the major site of umifenovir metabolization is in the liver, so it should be used with caution in patients with predisposing liver diseases. animal data revealed that umifenovir therapy couldn't induce embryo-toxic effects during pregnancy. according to these results umifenovir would be a promising safe and well-tolerated antiviral agent in pregnancy with a wide therapeutic index in administration for a few days up to one month . ribavirin is a nucleoside antihepaciviral agent (anti-hcv) which has been suggested for covid- treatment. ribavirin is a direct-acting antiviral (daa) agent . ribavirin is a nucleoside analog that has antiviral action against a variety of rna and dna viruses. the potential antiviral activity of ribavirin is inhibition of inosine monophosphate dehydrogenase (impdh) cellular protein and therefore intracellular gtp would be diminished which inhibits rna replication of viral genomes, so viral growth might be stopped. another possible antiviral activity of ribavirin is its immunomodulatory effects by suppression of il- . ribavirin also could inhibit rna polymerase activity and therefore inhibition of rna fragments' initiation and elongation, so viral protein synthesis could be inhibited. the ribavirin is contraindicated in patients with hypersensitivity to ribavirin, pregnant women and their partner, patients with severe renal failure, patients with severe hepatic failure, and patients with major hemoglobinopathies such as sickle cell anemia and major thalassemia . ribavirin distribution could significantly prolonged in erythrocytes for about to days, which is responsible for ribavirin-induced anemia . ribavirin has hepatic metabolism. its oral bioavailability (f) is about %. ribavirin elimination half-life (t ½) in the normal population is hours but in patients with pre-existing chronic hepatitis c infection, half-life could be increased to hours. so because of its prolonged half-life and potential overdose toxicity, ribavirin is contraindicated in patients with hepatic failure (child-pugh class b and c). time to peak level (t max ) after oral administration is between to hours. ribavirin excretion could take place through both urine and feces routes. because of its renal elimination, dose adjustment in patients with underlying kidney disease is highly essential. according to the previous pharmacokinetic/pharmacodynamic study, bayesian therapeutic drug monitoring would be a suitable approach to control ribavirin-induced anemia [ ] . one of the most important side effects of ribavirin is hemolytic anemia which could worsen cardiac disease in patients with underlying cardiac diseases and it could induce fatal or non-fatal myocardial infarction in them. so ribavirin should be avoided in patients with a history of unstable or severe cardiac diseases. ribavirin is contraindicated in patients with hepatic decompensation (child-pugh class b and c). ribavirin dosage adjustment in patients with renal failure highly depends on different formulations which are available. these data are shown in table . in children, if serum creatinine level rises over mg/dl during administration, ribavirin should be discontinued promptly . ribavirin has teratogenic and mutagenic effects. so it is a high-risk drug in pregnancy according to animal data. also because of its half-life of hours in multiple-dose drug therapy and the possibility of drug accumulation in tissue compartments for up to months, ribavirin administration is contraindicated in pregnant women and also in men who are pregnant women's partner. it was suggested that pregnancy should be avoided during ribavirin therapy and at least months after completion of therapy in women or men . ribavirin because of the prolonged plasma elimination half-life and molecular weight of da, potentially would have toxicity in nursing infants but there are no human data available . ribavirin may precipitate hematologic adverse effects of organ transplantation regimen such as immunosuppressive agents (mycophenolate mofetil, azathioprine, mtor inhibitors), trimethoprim/sulfamethoxazole, and valganciclovir. these hematologic adverse reactions would also worsen hematologic reactions related to covid- . so close patient monitoring is essential. . also, lopinavir would be a promising drug of choice in children with covid- . lopinavir/ritonavir are protease inhibitor, anti-retroviral agents. the potential antiviral mechanism of lopinavir/ritonavir is inhibition of viral protease, which is a critical enzyme in viral maturation and infectivity. low dose ritonavir in combination with lopinavir act as a pharmacokinetic enhancer by inhibition of lopinavir inactivation metabolism . the since lopinavir undergoes hepatic metabolism through cyp a enzyme, potential drug-drug interactions could occur with all drugs that are strong inhibitors or inducers of cyp a enzyme and p-glycoprotein inhibitors. also, ritonavir has hepatic metabolism via cyp a and cyp d . ritonavir has serious and life-threatening drug interactions with sedative-hypnotic agents, antiarrhythmic drugs, and ergot alkaloid agents because of the effect of ritonavir on their hepatic metabolism through cyp a and cyp d . concurrent use of these agents with ritonavir is absolutely contraindicated and should be avoided . lopinavir/ritonavir is contraindicated in patients with a history of hypersensitivity reactions such as toxic epidermal necrolysis (ten), stevens-johnson syndrome, angioedema, etc. to lopinavir/ritonavir or its components . absorption and oral bioavailability are highly affected by fasting or fed state, its absorption, bioavailability, and peak level (c max ) could be significantly increased with food. but food can delay t max from hours in fasting state to hours in the fed state. ritonavir also has small renal elimination so dosage adjustment is not required in patients with underlying kidney disease . lopinavir/ritonavir have major interactions with drugs used in cardiovascular diseases such as anti-coagulating agents (anti-factor xa inhibitors), none dihydropyridine calcium channel blockers, digoxin, antiarrhythmic agents such as amiodarone, etc. so close patient monitoring is required in covid- patients with pre-existing cardiovascular disease who are planned to treat with lopinavir/ritonavir and sometimes alternative drugs might be considered . in patients with mild to severe hepatic failure, there is no dosage adjustments available but lopinavir has primary liver metabolism and its auc will be increase by about %, so it should be used with caution . there is no dosage adjustments based on renal function according to the manufacture's labeling . results revealed that embryo-fetal risk of lopinavir/ritonavir is low, so it is compatible with pregnancy and should not be stopped during pregnancy . lopinavir and ritonavir with a molecular weight of and da respectively and their lipid solubility nature, are good candidates for excretion to milk during lactation period but their high plasma protein binding could limit this excretion. a comprehensive data is not available yet. it has been recommended that breastfeeding during lopinavir/ritonavir therapy is better to be avoided especially in developed countries . immunosuppressive agents are critical drugs in patients undergone solid organ transplantation. lopinavir/ritonavir cannot be administered in combination with immunosuppressive agents because of the occurrence of strong drug interactions. lopinavir/ritonavir can enhance the plasma level of immunosuppressive agents such as calcineurin inhibitors and mtor inhibitors. so if co-administration is essential, immunosuppressive agents dose reduction and therapeutic drug monitoring, to maintain optimum immunosuppressive plasma level, is highly recommended. it has been suggested that during covid- treatment, calcineurin inhibitors (ex. cyclosporine and tacrolimus) and mtor inhibitors (ex. sirolimus and everolimus) could be discontinued and replaced with lopinavir/ritonavir but it seems that the benefit of this drug discontinuation could not overlay the risk of allograft transplant rejection . tocilizumab is an interleukin- (il- ) inhibitor which is a disease modifying anti-rheumatic agent . a small retrospective observational study on covid- pneumonia patients receiving tocilizumab revealed that this drug would have potential benefits in these patients such as since an active immune response against respiratory viruses such as covid- is highly dependent on cytotoxic t cells' action, so in patients with total t cell count of fewer than cells/µl, aggressive intervention is essential. one of the possible approaches in enhancing t cell count in these patients is the administration of tocilizomab, because there is a reverse relationship between t cell count and the number of cytokines such as il- . ifn-α is a broad-spectrum antiviral agent that is commonly used in hepatitis management. after corticosteroids are a double-edged sword, they can inhibit our immune response and so the clearance of covid- could be delayed, but on the other hand, they can suppress our inflammatory response which is highly responsive to the lung damage and ards during viral convalescent plasma or immunoglobulins would be a promising therapy in covid- patients. previous results revealed that convalescent plasma therapy during viral infection outbreaks, it has been recommended that patients with covid- who are suffering from refractory hypoxemia should be managed with extracorporeal membrane oxygenation (ecmo in some countries such as switzerland, tocilizumab has been considered in patients with multiorgan failure and inotropic support . ace has a critical role in cardiac and immune systems. ace is related to heart function and it could be a potential cause of hypertension and diabetes mellitus development . since ace is a functional receptor for covid- , in patients with underlying cardiovascular diseases, clinical symptoms of covid- are more severe and fatal than the general population because, in cardiovascular diseases, ace secretion might be enhanced. administration of reninangiotensin-aldosterone system inhibitors, such as angiotensin converting enzyme inhibitors (aceis) and angiotensin receptor blockers (arbs) could enhance ace level. thiazolidinediones and ibuprofen might also enhance the ace level pneumonia but arbs, through blockade of angiotensin receptors, may have beneficial effects in these patients. most of the hepatic metabolites of drugs considered in covid- treatment such as chloroquine, hydroxychloroquine, and lopinavir/ritonavir would be found in urine due to renal elimination. so in patients with chronic kidney disease (ckd), the accumulation of drug metabolites would be expected if administered in routinely recommended doses for the normal population. therefore, individualized dose adjustment based on kidney function is required for each drug as mentioned above . a small study on covid- patients with end-stage renal disease (esrd) who undergone hemodialysis, revealed that the number of total t cells (cytotoxic and helper t cells), natural killer (nk) cells, and inflammatory cytokines were significantly lower than these levels in non-hemodialysis patients with covid- . this study revealed that esrd patients with hemodialysis who infected with covid- had a good prognosis and they had mild symptoms of pneumonia, it might be related to the fewer number of inflammatory cytokines and reduced immune function which can avoid crs but further studies are required to confirm this hypothesis . according to the recent studies, liver abnormalities (such as elevated ast and alt serum levels) have been occurred after and during infection with covid- . these abnormalities would be related to viral infection pathogenesis and direct liver injury or it may be drug-induced . almost all of the potential drugs in covid- treatment containing chloroquine, hydroxychloroquine, ribavirin, and lopinavir/ritonavir have hepatic metabolism. so injury to the liver because of pre-existing liver disease or acute hepatic failure would impair drug metabolism and therefore drug accumulation and enhancement in plasma level, which can lead to drug toxicity. in these patients, frequent liver function monitoring is essential to achieve an optimal serum drug level . also, dosage adjustment for each drug should be done individually according to the patients' liver function as mentioned above. the impact of chronic liver diseases such as chronic viral hepatitis, alcoholic and non-alcoholic liver diseases on occurrence of liver injury related to covid- infection, still is not clear. it seems that in patients with underlying liver disease, with the immunocompromised condition, who infected with covid- , more intensive and individualized pharmacotherapy is required. further studies would be also helpful to explain the exact role of pre-existing liver diseases in covid- prognosis . the new coronavirus (covid- ) was first detected in wuhan city of china in december . soon this coronavirus disease (covid- ) spreads around the world and became a pandemic. now many patients around the world are suffering from this disease. patients with underlying diseases are highly prone to severe covid- pneumonia. till now many drugs have been considered in the treatment of covid- pneumonia, but pharmacotherapy in patients with pre-existing comorbidities is highly challenging. in this review, different potential drugs which have been considered in covid- treatment have been discussed in detail. also, challenges in the pharmacotherapy of covid- pneumonia in patients with underlying disease especially heart diseases have been considered based on pharmacokinetic and pharmacodynamic aspects of drugs. patients with covid- who have cardiac diseases such as coronary heart disease and those who are aceis consumers should be highly considered in treatment options. also, patients with liver and kidney disease and those with organ transplants should be considered to avoid the occurrence of drug overdose toxicities and potential drug-drug interactions. funding: this work was not funded. coronavirus infections-more than 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a letter to editor liver injury in covid- : management and challenges. the lancet gastroenterology & hepatology ethical approval: not required. p.g. and s.m. were contributed equally in data gathering, writing-original draft, reviewing, and revising the final version of this manuscript. key: cord- -qw atrx authors: bhattacharyya, rajat; iyer, prasad; phua, ghee chee; lee, jan hau title: the interplay between coagulation and inflammation pathways in covid- -associated respiratory failure: a narrative review date: - - journal: pulm ther doi: . /s - - - sha: doc_id: cord_uid: qw atrx the novel coronavirus disease (covid- ) pandemic has caused an unprecedented worldwide socio-economic and health impact. there is increasing evidence that a combination of inflammation and hypercoagulable state are the main mechanisms of respiratory failure in these patients. this narrative review aims to summarize currently available evidence on the complex interplay of immune dysregulation, hypercoagulability, and thrombosis in the pathogenesis of respiratory failure in covid- disease. in addition, we will describe the experience of anticoagulation and anti-inflammatory strategies that have been tested. profound suppression of the adaptive and hyperactivity of innate immune systems with macrophage activation appears to be a prominent feature in this infection. immune dysregulation together with endotheliitis and severe hypercoagulability results in thromboinflammation and microvascular thrombosis in the pulmonary vasculature leading to severe respiratory distress. currently, some guidelines recommend the use of prophylactic low molecular weight heparin in all hospitalized patients, with intermediate dose prophylaxis in those needing intensive care, and the use of therapeutic anticoagulation in patients with proven or suspected thrombosis. strong recommendations cannot be made until this approach is validated by trial results. to target the inflammatory cascade, low-dose dexamethasone appears to be helpful in moderate to severe cases and trials with anti-interleukin agents (e.g., tocilizumab, anakinra, siltuximab) and non-steroidal anti-inflammatory drugs are showing early promising results. potential newer agents (e.g., janus kinase inhibitor such as ruxolitinib, baricitinib, fedratinib) are likely to be investigated in clinical trials. unfortunately, current trials are mostly examining these agents in isolation and there may be a significant delay before evidence-based practice can be implemented. it is plausible that a combination of anti-viral drugs together with anti-inflammatory and anti-coagulation medicines will be the most successful strategy in managing severely affected patients with covid- . suppression of the adaptive and hyperactivity of innate immune systems with macrophage activation appears to be a prominent feature in this infection. immune dysregulation together with endotheliitis and severe hypercoagulability results in thromboinflammation and microvascular thrombosis in the pulmonary vasculature leading to severe respiratory distress. currently, some guidelines recommend the use of prophylactic low molecular weight heparin in all hospitalized patients, with intermediate dose prophylaxis in those needing intensive care, and the use of therapeutic anticoagulation in patients with proven or suspected thrombosis. strong recommendations cannot be made until this approach is validated by trial results. to target the inflammatory cascade, low-dose dexamethasone appears to be helpful in moderate to severe cases and trials with anti-interleukin agents (e.g., tocilizumab, anakinra, siltuximab) and non-steroidal anti-inflammatory drugs are showing early promising results. potential newer agents (e.g., janus kinase inhibitor such as ruxolitinib, baricitinib, fedratinib) are likely to be investigated in clinical trials. unfortunately, current trials are mostly examining these agents in isolation and there may be a significant delay before evidence-based practice can be implemented. it is plausible that a combination of anti-viral drugs together with anti-inflammatory and anti-coagulation medicines will be the most successful strategy in a novel coronavirus infection caused by the severe acute respiratory syndrome coronavirus (sars-cov- ), now called coronavirus disease- , has led to a global pandemic in . it has affected , , people worldwide with , deaths directly attributed to the virus at the time of writing this paper (june , ) , and the rate of new infections continues to increase at a very fast pace [ ] . while most of those infected have mild cases of the disease, up to % can have a severe clinical picture (i.e., dyspnea, tachypnea, hypoxemia and/or lung infiltrates on radiological imaging), and approximately % of the infected patients will require admission to the intensive care unit (icu) for respiratory failure, septic shock, and/ or multiple organ failure [ ] . the severe respiratory failure caused by covid- infection results in acute hypoxemia, which is associated with severe ventilation/ perfusion (v/q) mismatch and overt intrapulmonary shunting [ , ] . however, the exact mechanism of respiratory failure remains unclear and there is evidence that pulmonary microvascular thrombosis due to thrombo-inflammation may be a major factor contributing to respiratory decompensation in these patients [ ] . this narrative review aims to summarize the current available evidence on the interplay between hypercoagulability, thrombo-inflammation, and pulmonary microvascular thrombosis in covid- infection resulting in respiratory failure and how this information can be used to design clinical trials to optimize patient outcomes. we begin with a concise review of the unique properties of sars-cov- that impacts the coagulation and inflammation pathways before proceeding with an in-depth discussion on the current available medical literature. we will also discuss anticoagulation and other pharmacological strategies described for these patients that pertain to the role of the coagulation pathway in the prevention and management of covid- -induced respiratory failure. for the purpose of this review, we performed a literature search on pubmed using the following keywords: covid- , coronavirus, sars-cov- , thrombosis, and respiratory failure. this review is based on previously conducted studies and does not contain any studies with human participants or animals performed by any of the authors. the virulence of sars-cov- sars-cov- is a novel betacoronavirus [large ribonucleic acid (rna) virus] that is similar to the earlier sars-cov virus that caused the severe acute respiratory distress (sars) outbreak in [ ] . multiple spike glycoproteins (s) protrude from the viral surface and give it a halo-like appearance (hence corona). the spike s protein enables the virus to engage with its target cell receptor, angiotensin-converting enzyme (ace ) [ ] . the host's transmembrane protease serine (tmprss ) primes the spike s protein of the virus to facilitate its internalization into the cell [ ] (fig. ) . after binding to the ace receptor, the virus enters the cell through endocytosis, releases ribonucleic acid (rna) into the cytosol, replicates by using the cell machinery, and finally, by the process of exocytosis, is excreted from the cell [ ] . ace is mostly expressed in the airway and type ii pneumocytes explaining the tropism of the virus to the respiratory system. not surprisingly, one of the major features of covid- infection is involvement of the respiratory system. importantly, ace receptors are also expressed by vascular endothelial cells [ ] . while most patients will have symptoms of respiratory tract infection and start to improve within a week, sudden clinical deterioration - days after initial symptom onset and development of severe respiratory failure often combined with multiorgan failure is commonly recognized [ ] . the group at highest risk of deterioration includes the elderly population, people with pre-existing chronic health conditions (e.g., cancer, chronic kidney disease, chronic obstructive pulmonary disease, immunocompromised state, obesity, cardiovascular comorbidities, sickle cell disease and type diabetes mellitus) [ ] . in addition, patients from certain ethnicity groups (e.g., afro-americans, south asians, and minority ethnic groups) and blood groups (e.g., blood group a with p . gene cluster) have been fig. indirect and direct injury to the lungs by the severe acute respiratory syndrome coronavirus involving the interplay between coagulation and inflammation pathways. ace angiotensin-converting enzyme , crp c-reactive protein, esr erythrocyte sedimentation rate, ldh lactate dehydrogenase, nets neutrophil extracellular traps, sars-cov- severe acute respiratory syndrome coronavirus , tmprss transmembrane protease serine shown to be at higher risk of worse outcomes [ ] [ ] [ ] (fig. ) . current literature suggests that a unique pattern of immune dysfunction is responsible for this disease progression in high-risk patients. suppression of lymphocytes by the virus leads to a complicated immune mechanism with activated macrophages which releases cytokines (fig. ). this creates a cytokine storm [mostly interleukin il- b and il- ], promoting expression of adhesion molecules for endothelial activation, inflammatory cell infiltration, and vascular inflammation. endothelial cells release pro-inflammatory cytokines that contribute to propagation of microcirculatory lesions. the dysfunctional endothelium then becomes proadhesive and pro-coagulant [ ] . presence of viral elements within endothelial cells causes endotheliitis and accumulation of inflammatory cells with increasing cell death, leading to impairment of microcirculatory function in different vascular beds and their consequent clinical effects [ ] . a study that examined the serum of covid- patients found elevated markers of neutrophil extracellular traps (nets) [ ] . nets are microbicidal proteins, extracellular webs of chromatin, and oxidant enzymes that are released by neutrophils to contain infections. nets have potential to cause inflammation and microvascular thrombosis in the lungs, causing respiratory failure. highly specific markers of nets such as myeloperoxidase (mpo)-deoxyribonucleic acid (dna), and citrullinated histone h (cit-h ) were found to be elevated in this study. these collective findings suggest a unique ability of the virus to cause intense inflammatory reaction that makes the virus lethal. severe hypercoagulability with respiratory failure is a recognized feature of covid- patients admitted in intensive care units (icu) [ ] . a combination of profound inflammation fig. currently known clinical and laboratory biomarkers of severity to predict disease progression, combining with timely antiviral, anti-inflammatory, and anticoagulation intervention to optimize outcome. chd chronic heart disease, cld chronic lung disease, ckd chronic kidney disease, doacs direct oral anticoagulants, fdps fibrinogen degradation products, htn hypertension, ifn interferon, jak janus kinase, ldh lactate dehydrogenase, lmwh low molecular weight heparin, nsaids nonsteroidal anti-inflammatory drugs, pt prothrombin time, tnf tumor necrosis factor, vw ag von willebrand antigen and microvascular thrombosis appears to be responsible for the clinical picture that leads to progressive multi-organ failure in a small percentage of patients, ultimately causing fatalities. initial response to the viral infection is by the activation of the innate immune system with neutrophils, macrophages, and cytokines. this results in elevation of acute-phase reactants like c reactive protein (crp), erythrocyte sedimentation rate (esr), and ferritin [ , ] . this can be considered as the first cytokine wave or hypercytokinemia, which is essential in controlling viral infection. dysregulated type interferon response has been demonstrated in mice models of sars-cov infection [ ] . it is postulated that similar suppressed type interferon response is present in covid- infection, which when combined with suppression of the adaptive immune system [t,b and natural killer (nk) lymphocytes] creates an immunodeficient state that results in poor and delayed viral clearance [ ] . initial hypercytokinemia, suppressed type interferon response, and the inability of the adaptive immune system to clear the virus sets the stage for hyper activation of the innate immune system and cytokine storm in some patients. cytokine dysregulation is a recognized feature of this infection. studies in hospitalized patients have shown that plasma interleukin beta(il b), il- receptor antagonist (il ra), il , il , il , il , granulocyte colony-stimulating factor (gcsf), granulocyte monocyte colony-stimulating factor (gmcsf), interferon gamma (ifnc), interferon gamma-induced protein (ip ), monocyte chemoattractant protein (mcp ), macrophage inflammatory protein alpha (mip a), macrophage inflammatory protein beta (mip b), platelet-derived growth factor (pdgf), tumor necrosis factor alpha (tnfa), and vascular endothelial growth factor (vegf) concentrations on admission were higher in patients with covid- infection compared to healthy adults [ ] . in the same study, plasma concentrations of il , il , il , gcsf, ip , mcp , mip a, and tnfa were found to be higher in patients needing icu admission compared to those who did not. disease severity has been found to positively correlate with serum levels of tnfa, il , and il , and there was also a strong negative correlation of levels of tnfa and il with the number of cd ?, cd ? t lymphocytes [ ] . in a study by diao et al., higher levels of the exhausted marker pd- were seen in covid- patients, suggesting not only that lymphocyte numbers are lower, but existing t cells are functionally exhausted. in a larger study of patients, of whom were classified as having severe disease, dysregulation of the immune system was seen with a high leucocyte count and high neutrophil-to-lymphocyte ratio [ ] . regulatory t cells that help control inflammation were also found to be low in this study, especially in patients with severe disease. these mechanisms partly explain the unabated progression of the inflammatory process, the ongoing cytokine storm, and the progressive tissue damage. the similarity of severe covid- disease to thrombotic microangiopathic anemia with elevated lactate dehydrogenase (ldh), d-dimer and bilirubin, decreased platelets, anemia, and renal and cardiac injury is suggestive of excessive complement activation. hence, complement inhibition as a therapeutic option has also been suggested [ ] . another multi-center study conducted in europe investigated immune dysregulation in patients ( with severe respiratory failure). clinical features suggestive of macrophage activation syndrome (mas) associated with low cd ? t-cells, cd ? b-cells, and nk cells were found in all the patients. sustained elevation of tnfa and il- was also found, representing a hyperactive monocyte macrophage system [ ] . in another study, lymphocyte subset analysis in covid- patients showed a statistically significant lower number of cd ? helper t cells, cd ? cytotoxic t cells, nk cells, and b cells when compared to healthy controls [ ] . in this study, patients with severe disease ( % cases) had significantly lower cd ? and cd ? t cells and b cells, although no significant differences were seen in the cd /cd ratio or nk cell numbers when compared to those with mild symptoms. these studies suggest that in this disease, a suppression of the adaptive immune system along with hyper activation of the innate immune system causes a disease phenomenon that is similar to mas/hemophagocytic lymphohistiocytosis (hlh). comparing the laboratory features, hyperferritinemia and high lactate dehydrogenase (ldh) levels are common in both covid- patients and patients with primary hlh. on the other hand, low fibrinogen and cytopenias due to bone marrow hemophagocytosis is not commonly found in covid- patients [ ] . it is likely that the pathophysiology of covid- overlaps with low-grade hlh, and further studies addressing this aspect will be important to evaluate the role of immunosuppression and hlh type therapy in these patients. the acronym covid- -associated coagulopathy (cac) is being used to describe the coagulation changes in infected patients [ ] . abnormal coagulation parameters including increased d-dimer, fibrin degradation products, and prolonged pro-thrombin time have been found to be associated with poor prognosis [ ] . in a study of patients with covid- disease, out of ( . %) non-survivors had disseminated intravascular coagulation (dic) as per the criteria set by the international society of thrombosis and haemostasis (isth). this was in contrast to only one ( . %) survivor fulfilling the criteria for dic during the period of hospitalization, indicating that non-survivors have a higher incidence of dic. described incidence of thrombocytopenia has been variable, reported as % in an initial case series of patients [ ] and . % in another series of patients [ ] . a meta-analysis analyzing pooled data from nine studies of patients including with severe disease has demonstrated that thrombocytopenia is prognostic of the severity of the disease. although there was notable heterogeneity between studies, severe thrombocytopenia was common in non-survivors [ ] and increased mortality has been associated with lower platelet counts [ ] . activated partial thromboplastin time (aptt)based clot waveform analysis (cwa) is an easily available form of global hemostatic assay and remarkably high cwa were noted despite prophylactic anticoagulation in patients admitted to the icu although there was no thrombosis or death in a study from singapore [ ] . a study on critically ill patients analyzed whole-blood thromboelastography (teg) which showed decreased r (reaction time) and k (kinetics time) values but increased k angle and ma (maximum amplitude) values, which is consistent with hypercoagulability. factor viii, von willebrand factor, fibrinogen, d-dimer, platelets, and protein c were increased in most patients, and antithrombin marginally decreased, while prothrombin time (pt) and aptt were mostly normal [ ] . the authors concluded that the above findings do not support features of acute dic but are consistent with severe hypercoagulability secondary to inflammation. another similar study in patients admitted to the icu with covid- showed fibrinolysis shutdown as evidenced by markedly raised d-dimer and complete absence of clot lysis in teg at min; this was predictive of high rate of venous thrombosis) and need for hemodialysis [ ] . this may suggest that functional coagulation assessment like teg may be more useful in assessing hypercoagulability rather than conventional coagulation tests in patients with covid- . a high rate of venous thrombosis has been observed even in anti-coagulated patients with this disease. in a french study of patients on mechanical ventilation, prophylactic anticoagulation was used in eight ( %) cases while therapeutic dose anticoagulation with low molecular weight heparin (lmwh) was started on admission in ( %) patients [ ] . all of these patients were screened by doppler ultrasound of the lower limbs. despite anticoagulation, high rates of deep vein thrombosis were noted in all patients in the prophylactic dose group and % in the therapeutic dose group that was associated with high rates of pulmonary embolism [eight ( %) in the whole cohort]. a larger study involving patients admitted in three hospitals in the netherlands and treated with prophylactic enoxaparin reported an incidence of % and % of deep vein or arterial thrombosis and pulmonary embolism, respectively [ ] . a follow-up of the same cohort of patients was published, which confirmed the high incidence of pulmonary embolism ( / , %) in those patients diagnosed with deep vein thrombosis; and pulmonary embolism was associated with higher risk of all-cause mortality [ ] . a high rate ( %) of lupus anticoagulant activity was identified in a study of patients when screened by a sensitive aptt test and direct russel viper venom test (drvvt). this may be a contributing factor in the development of thrombosis in some patients [ ] . peripheral arterial disease in the form of arteriosclerosis obliterans of lower extremity in combination with deep vein thrombosis has also been reported in severe infection, which confirms the presence of a severe hypercoagulable status in these patients [ ] . the pathophysiological mechanism of respiratory failure in covid- infection is not clear at present but there is an increasing body of evidence suggesting microthrombosis of the pulmonary vasculature is a key player (fig. ) . in covid- pneumonitis, histological changes showed pauci-inflammatory septal capillary injury with significant septal capillary fibrin deposition and neutrophil infiltration of alveolar septa [ ] . viral cytopathic and classical acute respiratory distress syndrome (ards) changes such as diffuse alveolar damage with hyaline membranes or hyperplasia of type ii pneumocyte were not prominent. deposits of terminal complement components c b- , c d, and mannose binding lectin-associated serine protease (masp) in the microvasculature was noted, suggesting activation of the alternative and lectin-based complement pathways. another study with lung autopsy findings from ten patients showed predominantly proliferative diffuse alveolar damage, epithelial viral cytopathic effects of small airway epithelium, but minimal lymphocytic infiltration [ ] . the lung injury observed in this study was secondary to formation of fibrinous thrombi in small pulmonary arteriole, endothelial tumefaction, and megakaryocyte aggregation in pulmonary capillaries, suggesting activation of coagulation cascade with microthrombus formation. the term ''microvascular covid- lung vessels obstructive thrombo-inflammatory syndrome (microclots)'' has been proposed to describe the thrombo-inflammatory lung pathology [ ] . the working hypothesis is that in individuals at risk of severe disease, an inflammatory reaction and pulmonary microvascular thrombosis leading to alveolar damage is initiated. because of the systemic endotheliitis, this thrombo-inflammatory syndrome may progress to involve a microvascular bed of other major organs contributing to multi-organ failure. it is increasingly recognized that in this infection, pulmonary thrombus develops due to a local inflammatory reaction. platelets interacting with the vascular wall, leukocytes, factor xiia, von willebrand factor and complement leads to an intense thrombo-inflammation, which is a direct consequence of vascular damage associated with viral infection [ ] . because of the unusually high incidence of thrombotic complications despite prophylactic anticoagulation, some centers recommend a higher dose for prophylactic anticoagulation (e.g., enoxaparin mg twice a day rather than the conventional dosing of mg once a day) [ ] . although there appears to be a consensus about treating all hospitalized patients with some form of anticoagulation, dosing strategies are not yet clear. the isth interim guidelines and some other expert opinions have proposed a day) . however, this approach of either standard or intermediate dose prophylaxis has been questioned and systemic anticoagulation with unfractionated heparin infusion is routinely used for critically ill patients in some centers [ ] . this approach is recommended because cac is considered to be an overwhelmingly thrombotic dic with pulmonary embolism being difficult to diagnose in patients who require mechanical ventilation. also, bleeding is noted to be rare even in sick patients, and hence the risk of therapeutic anticoagulation is not as great as other patients with sepsis-induced coagulopathy. hyperfibrinogenemia with fibrin deposition in the air spaces and lung parenchyma due to a hypofibrinolytic state is well recognized in these patients [ ] . fibrin deposition can occur even before appearance of symptoms of infection as identified in lung biopsy specimens of two patients with lung cancer [ ] . there is limited experience in the use of fibrinolytic therapy with tissue plasminogen activator (tpa). it has been used in off-label fashion in three patients with a noted improvement in oxygenation [increase in partial pressure of oxygen (pao )/ fraction of inspired oxygen (fio ) (p/f) ratio] [ ] . nebulized tpa may be effective with less risk of bleeding and this is currently being examined in a clinical trial (nct ). a recent review has recommended standard prophylactic dose of anticoagulation in hospitalized but well patients, intermediate dose prophylaxis to those admitted in icu and ards, and therapeutic anticoagulation to those with presumed or confirmed deep vein thrombosis [ ] . currently, there are trials registered with clinicaltrials.gov that are investigating anticoagulation in these patients [ ] (table ) . these are mainly adult trials with only one pediatric study. one registered study aims to look at antiplatelet agents to prevent cardiac complications. outcomes from these trials (table ) will be important for our understanding of the optimum anticoagulation strategy in this infection to prevent thrombosisassociated complications. glucocorticoids have been investigated in the treatment of covid- disease. unfortunately, the first randomized controlled trial of methylprednisolone against standard of care in beijing, china (nct ) had to be stopped because of lack of patients, with the pandemic ending in china. at the time of writing this review, reports but not the full peer-reviewed manuscript for the randomised evaluation of covid- therapy (recovery) conducted in the united kingdom provided preliminary data that low-dose dexamethasone reduced deaths by one-third in ventilated patients and by onefifth in other patients receiving oxygen, but made no difference to those with milder disease [ ] . there are several other ongoing trials of systemic steroids (nct , nct , nct , nct ) and some others looking at efficacy of inhaled steroids (nct , nct , nct ). similar to patients with sars and middle eastern respiratory syndrome (mers), the cytokine responses seen in patients with covid- have features of a cytokine storm. one study involving adult patients found significant correlation of increased baseline il levels with disease severity [ ] and this is supported by similar findings in another study [ ] . hence, some investigators have proposed using il as a biomarker for assessing disease severity and blocking il as a therapeutic strategy has been suggested [ ] . tocilizumab, a monoclonal antibody against the il receptor, has been used safely and effectively in patients with rheumatoid arthritis [ ] . a pilot trial of adding tocilizumab therapy in patients with severe covid- disease in combination with standard therapy of lopinavir, methylprednisolone, and oxygen therapy showed remarkable short-term clinical responses in out of patients within weeks of tocilizumab therapy [ ] . no adverse drug reactions attributable to tocilizumab were noted. following the success of this preliminary trial, a larger multicenter clinical trial is currently being undertaken (chictr ). another trial in belgium has been designed to observe the efficacy of the il blockers tocilizumab and siltuximab individually and in combination with the il blocker anakinra (nct ). indeed, il , il blockade, ifn-c inhibition, and tnfa inhibition are also amongst consideration for randomized control trials in patients with severe disease. high-dose intravenous immunoglobulin (ivig) has been used as an immunomodulator in patients with severe disease with good outcomes [ ] . there are several trials currently exploring the utility of ivig (nct , nct , nct , nct , and nct ). the role of non-steroidal anti-inflammatory drugs (nsaids) as therapy is extremely controversial. ibuprofen is a commonly used nsaid, which is used for fever control, usually with acetaminophen. however, ibuprofen increases the synthesis of ace enzyme and may potentially increase the risk of severity and fatality of covid infection [ ] . however, currently there is no definite evidence for or against the use of ibuprofen. the european medicine agency (ema) guidelines published on march , suggest using acetaminophen for fever but also had advised not to stop ibuprofen if the patient is on it for other reasons [ ] . currently, there are ongoing trials to assess the efficacy of inhaled ibuprofen (nct ) and also oral lipid formulation of ibuprofen (liberate trial, lipid ibuprofen versus standard of care for acute hypoxemic respiratory failure due to covid , nct ). the efficacy and safety of other nsaids are currently being evaluated, the details of which are beyond the scope of this article. combining anti-inflammatory agents with anti-viral agents is another attractive option. artificial intelligence (ai)-powered algorithm predictions have identified baricitinib, ruxolitinib, and fedratinib as potential candidates [ ] . these drugs belong to the janus kinase (jak) inhibitor class of medications that have been approved and used safely in conditions like myelofibrosis and rheumatoid arthritis. these drugs are powerful anti-inflammatory agents that target the jak-stat pathways and are likely to be effective against the deleterious consequences of elevated cytokines. of particular interest is baricitinib, which is predicted to interfere with the ability of the virus to enter and infect lung cells by receptor-mediated endocytosis [ ] . due to the favorable pharmacological and safety profile of baricitinib, it can be considered in combination with direct-acting anti-viral agents like lopinavir, remdesivir, and ritonavir to decrease the viral replication, infectivity, as well as to calm the cytokine responses. statins, another group of drugs that have anti-inflammatory and antithrombotic properties, have also been studied in covid- patients; a retrospective study showed a favorable recovery profile and lower all-cause mortality with the use of statins in hospitalized patients with covid- [ ] . convalescent plasma has historically been used to improve outcomes in a variety of viral epidemics [ ] . in theory, recovered covid- patients' plasma containing anti covid- neutralizing antibodies should provide passive immunity to patients with ongoing disease. however, a recent systematic review failed to demonstrate any benefit of this approach in patients hospitalized with covid- , in addition to uncertainty of the safety profile of convalescent plasma therapy [ ] . it has been a steep learning curve for the medical fraternity in learning how to manage critically ill patients with covid- . the existing knowledge of abnormalities in inflammatory and coagulation pathways offers opportunities for identifying biomarkers for severe disease, which combined with clinical risk factors will help in prompt identification of patients at risk of severe disease who require early and targeted intervention. better understanding of the pathophysiology suggests suppression of the adaptive immune system on one hand and a hyperactive innate immune system on the other as the two principle severity-driving mechanisms in covid- infection. this leads to a mas-like picture associated with a hypercoagulable state that causes both micro and macrovascular thrombosis in the pulmonary as well the systemic circulation. this understanding will help us develop treatment protocols targeting various inflammatory pathways as well as to optimize anticoagulation therapies. current anticoagulation practice suggests that all hospitalized patients should receive prophylactic anticoagulation, those with severe disease needing intensive care with respiratory compromise should have intermediate dose prophylaxis and therapeutic anticoagulation reserved for those with proven or suspected thrombosis. results of prospective trials will only answer if this strategy is optimum in preventing thrombotic complications and improving pulmonary and overall outcome. a multi-pronged approach combining anti-inflammatory drugs and anticoagulation with specific antiviral medications is likely the approach that will work in critically ill patients (fig. ) . many trials are currently trying to address efficacy of different antiviral, anti-inflammatory, and anticoagulation agents in isolation. unfortunately, these trials may take a significant amount of time for completion and analysis before the evidence-based recommendations can be produced. because of the enormous number of new patients over a short period, we have a unique opportunity to develop an international collaborative network and conduct large trials with many combination arms to answer the all-important question of which combination of antiviral, anti-inflammatory drugs with anticoagulation therapies are going to be the most effective in treating patients with severe disease. early intervention strategy is likely to be helpful in preventing disease progression and thereby reducing the morbidity and mortality from this pandemic. funding. no funding or sponsorship was received for this study or publication of this article. authorship. all named authors meet the international committee of medical journal editors (icmje) criteria for authorship for this article, take responsibility for the integrity of the work as a whole, and have given their approval for this version to be published. disclosures. rajat bhattacharyya, prasad iyer, ghee chee phua, and lee jan hau have nothing to disclose. compliance with ethics guidelines. this article is based on previously conducted studies and does not contain any studies with human participants or animals performed by any of the authors. data availability. data sharing is not applicable to this article, as no datasets were generated or analyzed during the current study open access. this article is licensed under a creative commons attribution-noncommercial . international license, which permits any non-commercial use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the creative commons licence, and indicate if changes were made. the images or other third party material in this article are included in the article's creative commons licence, unless indicated otherwise in a credit line to the material. if material is not included in the article's creative commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from 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immunoglobulin for people with covid- : a living systematic review key: cord- -o k ae authors: he, bing; wang, jun; wang, yudie; zhao, juan; huang, juan; tian, yu; yang, cheng; zhang, heng; zhang, mingxia; gu, lixing; zhou, xiaocui; zhou, jingjiao title: the metabolic changes and immune profiles in patients with covid- date: - - journal: front immunol doi: . /fimmu. . sha: doc_id: cord_uid: o k ae to explore the metabolic changes and immune profiles in patients with covid- , we analyzed the data of patients with mild and severe covid- as well as young children with covid- . of the leukocytes, % (iqr, – ) were lymphocytes [ . × ( )/l (iqr, . – . )], and monocytes were . × ( )/l (iqr, . – . ) in young children with covid- . in mild covid- patients, circulating monocytes were . × ( )/l (iqr, . – . ). twenty-one severe patients had low po( ) [ mmhg (iqr, – )] and so( ) [ % (iqr, – )] and high lactate dehydrogenase [ u/l (iqr, – )], cardiac troponin i [ . ng/ml (iqr, . – . )], and pro-bnp [ pg/ml (iqr, – , )]. serum d-dimer and fdp were . mg/l (iqr, . – . ) and . mg/l (iqr, . – . ), and a large number of rbc ( /μl (iqr, – ) was presented in urine, a cue of disseminated intravascular coagulation (dic) in severe patients. three patients had comorbidity with diabetes, and patients without diabetes also presented high blood glucose [ . mmol/l (iqr, . – . )]. fifteen of ( %) severe cases had urine glucose +, and nine of ( %) had urine ketone body +. the increased glucose was partially caused by reduced glucose consumption of cells. severe cases had extraordinarily low serum uric acid [ μmol/l (iqr, – )]. in the late stage of covid- , severe cases had extremely low cd (+) t cells and cd (+) t cells, but unusually high neutrophils [ . × ( )/l (iqr, . – . )], procalcitonin [ . ng/ml (iqr, . – . )], c-reactive protein [ mg/l (iqr, – )] and an extremely high level of interleukin- . four of ( %) severe cases had co-infection with fungi, and two of ( %) severe cases had bacterial infection. our findings suggest that, severe cases had acute respiratory distress syndrome (ards) i–iii, and metabolic disorders of glucose, lipid, uric acid, etc., even multiple organ dysfunction (mods) and dic. increased neutrophils and severe inflammatory responses were involved in ards, mods, and dic. with the dramatical decrease of t-lymphocytes, severe cases were susceptible to co-infect with bacteria and fungi in the late stage of covid- . in young children, extremely high lymphocytes and monocytes might be associated with the low morbidity of covid- . the significantly increased monocytes might play an important role in the recovery of patients with mild covid- . in december , an unknown viral pneumonia emerged in wuhan, china, and it then escalated into an unprecedented outbreak ( ) . chinese authorities have identified a new type of coronavirus named severe acute respiratory syndrome coronavirus (sars-cov- ) ( ). on february , , the infectious disease caused by this viral strain was officially named covid- (coronavirus disease ) by the world health organization (who) ( ) . by march , covid- had swept into at least countries and killed more than , people, and who officially announced a pandemic of covid- viral disease ( ) . as of june , , covid- cases have been confirmed in countries and territories, and the total was up to , , , including , deaths ( ) . so far, according to reported patients' data, some remarkable phenomena have been observed. first, only % of patients with covid- were infants and young children, and very few young patients have developed severe covid- ( ) . leukocytes are the main immune cells to fight against pathogens, and the total leukocyte count is higher in young children than in adults ( ) . moreover, the thymus gland of an infant is large and continues to grow throughout childhood. thus, the thymus produces more than enough matures tlymphocytes throughout the child's life ( ) . we explored whether the count and differential of leukocytes in infants and young children are associated with very low morbidity rates of covid- . second, from the epidemiology and clinical characteristics of covid- , % of patients were diagnosed as mild cases, and most mild cases can recover from covid- infection ( ) . so, it could be that specific leukocytes contributed to the recovery of patients with mild covid- . monocytes are important immune sentinel cells critical in the defense against viral infection in the blood. they achieve this via diverse mechanisms that include the detection of viruses, migration into infected tissues, differentiation into macrophages and dendritic cells, and pathogen clearance by phagocytosis and intracellular killing ( , ) . besides monocytes, the effect of lymphocytes on mild covid- cases is still unclear. in this study, mild patients have been examined to explore the potential roles of monocytes and lymphocytes in the recovery of patients with mild covid- . third, according to an analysis of nearly , confirmed cases, % of patients with covid- have been identified as severe cases and critically ill cases, involving severe pneumonia and metabolic disorders, developing into acute respiratory distress syndrome (ards), multiple organ dysfunctions (mods), and even septic shock and death ( , ) . some studies suggested that the immunopathogenesis after viral infection has been linked to the development of the disease into severe cases ( , ) . to explore the potential roles of immunopathogenesis in the progress of covid- infection, severe covid- patients have been investigated to explore how the immunopathogenesis was involved in ards and metabolic disorders, even mods, disseminated intravascular coagulation (dic), and death. in this study, we investigated mild cases and severe cases infected with sars-cov- , as well as healthy young children and adults. our multiple comparative analysis showed that not only is leukocyte composition different in healthy groups, these differences can also be found during various stages of sars-cov- infection. our study suggests that monocytes, neutrophils, and t-lymphocytes are associated with the onset and progress of covid- infection, and immunopathogenesis was involved in ards, metabolic disorders, and mods in severe cases. this study increases our understanding of the immune responses during covid- infection and provides support to develop novel, feasible, and effective treatments for covid- infection. research sources: covid- patients and healthy individuals covid- infection was rapidly endemic in wuhan, china, in january, . renmin hospital of wuhan university is at the very forefront of the fight against covid- . we collected the data of patients with covid- , including the clinical records, laboratory results and chest computed tomography (ct) scan images of mild and severe cases in the hospital. for comparison with covid- cases, the data of healthy adults and young children have been collected from the physical examination center of the hospitals. these healthy individuals have no significant medical condition and were in stable physical condition at that time. the data of patients with covid- and healthy persons have been all reviewed by a group of professional doctors from the hospitals, including basic features, nucleic acid tests, clinical data, laboratory results, co-infection with other pathogens, ct images, and other primary data. the study design has been approved by the ethics committee of the hospital. nasopharyngeal swab samples were collected from patients, and tested as soon as possible to increase the detection rate of sars-cov- . reverse transcription polymerase chain reaction (rt-pcr) kit (daan gene, shenzhen, china) was used to detect the conserved genes of sars-cov- , such as orf ab gene, n gene, and e gene with lightcycler system (roche, switzerland). if two or more of these three targeted genes has been detected as positive or one gene has been detected positive in two different samples from the same patient, the result is considered as positive for sars-cov- . meanwhile, the results can also be analyzed in conjunction with the patient's chest ct images. blood samples were collected from patients for laboratory tests. serum biochemical tests, including aspartate aminotransferase (ast), alanine aminotransferase (alt), creatine kinase (ck) and lactate dehydrogenase (ldh) were determined with cobas c testing system (roche, germany). procalcitonin (pct) and cardiac troponin i (ctn i) were analyzed by cl- i chemiluminescence immunoassay system (mindray, shenzhen, china). coagulation indicators were detecting with acl top hemostasis testing systems (werfen, usa). all the blood samples from healthy persons were used for comparison. to study the count and differential of lymphocytes, the blood samples from covid- patients were stained with cd , cd , cd , cd , cd , and cd antibodies (bd multi-test imk kit, usa) and were analyzed by bd facscanto ii flow cytometer (bd, usa). th /th kit (bd, usa) was used to quantitatively measure il- , il- , il- , tnf, and ifn-γ protein levels. to examine the effect of sars-cov- on the patients' humoral immune function, immunoglobulins (igm, igg, iga, and ige), complement (c ) and complement (c ) were tested (siemens healthineers, usa). c-reactive protein (crp) and interleukin (il- ) were measured for covid- patients (mindray, shenzhen, china). serum samples of patients were collected and tested for the igm of respiratory tract pathogens with pneumoslide igm kit (vircell, spain), including human respiratory syncytial virus, influenza a virus (subtypes h n and h n ), influenza b virus, parainfluenza virus / / , metapneumovirus, common coronavirus, epstein-barr virus, cytomegalovirus, rhinovirus, adenovirus, and bocavirus, as well as legionella pneumophila serum type i, mycoplasma pneumonia, and chlamydia pneumoniae. nasopharyngeal secretions were tested for nucleic acids of respiratory pathogens (health gene technologies, ningbo, china). sputum culture was performed to identify bacterial and fungal co-infection. the fungal examination was performed with fungus ( - )-β-d-glucan kit (dynamiker biotechnology, tianjin, china) and platelia aspergillus ag kit (bio-rad, usa). continuous measurements have been presented as median and interquartile range (iqr) and categorical variables as percentages. for assessing laboratory results, we also assessed whether the measurements were outside the normal range. unpaired t-test with welch's correction was used for comparison, and p < . and < . were considered statistically significant and highly statistically significant, respectively. graphpad prism . . (san diego, ca, usa) and spss . (ibm, armonk, ny, usa) were used for all analyses. patients with fever and/or cough were admitted to hospital after february , . chest ct images indicated multiple patchy, ground-glass opacity in the lungs ( figure a) . thirty-two patients were further diagnosed as infected with sars-cov- by real-time rt-pcr. there were men and women, and the median age of these mild cases was . the clinical characteristics of mild patients were presented in supplementary table s . compared with healthy adults, the count of leukocytes and neutrophils in mild covid- patients did not increase, but the median percentage and count of lymphocytes were % (iqr, - ) and . × /l (iqr, . - . ), respectively, which were significantly less than those of healthy adults, % (iqr, - ) and . × /l (iqr, . - . ), respectively (p < . ). interestingly, the median percentage and count of monocytes were . % (iqr, . - . ) and . × /l (iqr, . - . ), which were significantly higher than those of healthy adults . % (iqr, . - . ) and . × /l (iqr, . - . ) (p ≤ . ) ( table ). the significantly increased number of monocytes could play an important role in the recovery of patients with mild covid- . to investigate why infants and young children have low morbidity of covid- , we analyzed the clinical characteristics of young children with covid- , and collected the data of circulating leukocytes of young children with/without covid- . comparative analyses showed that young children have much higher leukocyte counts [ . × /l (iqr, . - . )] than adults. of note, % (iqr, - ) of leukocytes are lymphocytes [ . × /l, (iqr, . - . )] in young children. the median count of monocytes in young children is . × /l (iqr, . - . ), which is much higher than that of adults [ . × /l (iqr, . - . )] (p = . ). lymphocytes of young children with covid- was a little lower than those of healthy children, but remained at a high level [ . × /l (iqr, . - . )]. young children with covid- had a high level of monocytes [ . × /l (iqr, . - . )] as well ( table ) . such a high number of lymphocytes and monocytes has benefit to fight against sars-cov- , which might be associated with the low morbidity of covid- in young children. we collected and compared the data of severe cases and mild cases. chest ct images of severe cases indicated that there was critically diffuse ground-glass opacity in both lungs. the critically ill patient's bedside chest x-ray showed the lung texture enhanced and the translucency decreased, and multiple patchy shadows in both lungs. (d) serum il- concentration between mild patients (n = ) and severe patients (n = ). the normal range of il- is ≤ pg/ml. **p < . . (e) the analysis of th /th cytokine panel between mild patients (n = ) and severe patients (n = ). the normal range of il- , ifn-γ, il- , and il- are ≤ . pg/ml, pg/ml, . pg/ml, and . pg/ml, respectively. *p < . . a representative ct image is presented in figure b . in bedside chest x-ray results of the critically ill patients, the translucency of both lungs was diffusely decreased, and a large area of patchy shadow appeared with uneven density. tracheal intubation can be observed in the trachea and the heart shadow outline ( figure c) . the clinical characteristics of severe patients were presented in supplementary table s . these ct and x-ray images showed that the primary and most significant changes were in the lower respiratory tract of patients with severe covid- . among the respiratory indicators we measured, severe cases had lower partial pressure of oxygen (po ) and oxygen saturation (so ), mmhg (iqr, - ) and % (iqr, - ), respectively, and suffered from different degrees of ards, i to iii ( table ) . table ) . increased glucose and low uric acid in blood should be noted here. the level of blood glucose was . table ) . the total of leukocytes was . × /l (iqr, . - . ) in the peripheral blood of severe cases, which were much more than those in mild cases. compared with mild cases, severe cases had low levels of monocytes [ . × /l (iqr, . - . )]. however, the percentage and count of lymphocytes in severe cases were only % (iqr, - ) and . × /l (iqr, . - . ) respectively, which were significantly lower than those in mild cases ( table ) . the subsets of lymphocytes were examined by flow cytometry, including natural killer (nk) cells (cd + cd + ), b cells (cd + ), and t cells (cd + ). the results showed that severe cases had nk cells [ /µl (iqr, - )] and b cells [ /µl (iqr, - )], which was not a significant difference from the mild cases (p > . ). in addition, the functions of b cells and complements were tested, including igm, igg, iga, ige, c , and c , for both mild and severe covid- cases. for severe cases, the values of igm, c , and c were slightly lower than those in mild cases, but these values were still within the normal range. however, compared with mild cases, severe cases had much lower levels of cd + t cells and cd + t cells, /µl [iqr, - ] and /µl (iqr, - ), respectively. the decrease of cd + t cells was much more than that of cd + t cells, and the ratio of cd + t cells/cd + t cells increased by . (iqr, . - . ) ( table ). further examination of th /th cytokines also indicated that severe patients had normal levels of il- , and ifnγ, as well as il- in peripheral blood, but the level of il- in severe patients was times higher than normal (figure e ). in this study, the clinical course of severe cases was over weeks, and severe cases had a potentially high risk of co-infection with other pathogens due to critical exhaustion of cd + and cd + t cells. the respiratory tract pathogens were tested in severe cases, including viruses, legionella pneumophila, mycoplasma pneumoniae, and chlamydia pneumoniae, which were all negative. the fungal examinations, g assay and gm assay, were also performed in severe cases. the results of bacterial and fungal examinations indicated that four of ( %) severe cases had co-infection with fungi, and two of ( %) severe cases had co-infection with bacteria. a high number of white blood cells (wbc) [ /µl (iqr, ] was found in the urine of severe cases ( table ). further examinations showed that the median pct was . ng/ml (iqr, . - . ) in severe cases, a cue of potential sepsis/septic shock. among the inflammatory factors tested in severe cases, the median of crp was mg/l (iqr, - ), which was much higher than those in mild cases ( table ) . il- slightly increased in mild cases, but exceptionally high level of il- presented in severe cases, even times higher than normal in some critically ill cases (figure d) . the release of the inflammatory factors triggered by sars-cov- replication and/or co-infection with bacteria and fungi, played important roles in the progress of covid- infection. in the late stage of the disease in severe covid- cases, % (iqr, - ) of leukocytes were neutrophils [ . × /l (iqr, . - . )] ( table ) . previous studies showed that largely number of neutrophils triggered inflammatory responses and caused excessive organ injury in acute inflammatory disease, such as sepsis ( , ) . exceptionally high neutrophil numbers might be involved in severe inflammatory responses and might be associated with ards, mods, and even sepsis/septic shock, dic, and death during the late stage of severe covid- infection. in this study, we first analyzed the clinical features and leukocyte differential of mild covid- patients admitted to the hospital after february , . thirty-two mild cases, with a median age of years, had recovered from covid- infection. our data showed that compared with healthy adults, patients with mild covid- had lower lymphocytes in the acute stage, which was consistent with previous studies ( ) . however, mild covid- cases had high counts of circulating monocytes [ . × /l (iqr, . - . )]. in addition, mild patients had normal level of il- and il- in peripheral blood, but they had a - -fold increase of il- . monocytes/macrophages play very important roles in fighting against invading foreign viruses. literature from the past years has emphasized links among il- and innate immune response, such as mononuclear phagocytes ( , , ) . for patients with mild covid- , a high monocyte count and slight increase of il- might be helpful for eradicating the sars-cov- infection and were associated with recovery from covid- . based on the epidemiology and clinical characteristics of covid- , young children under six have the lowest morbidity rate, and very few young children with covid- develop severe cases ( , ) . according to our comparative analysis, young children under six have highly circulating monocytes, and % (iqr, - ) of leukocytes are lymphocytes had a high level of monocytes [ . × /l ( . - . )] as well. the intricate process of t-lymphocyte development in the thymus is essential in the formation and maintenance of the peripheral t-lymphocytes. the thymus of a young child is big, and has the function of maintaining the large amounts of t-lymphocytes in the peripheral blood ( , ) . extremely high levels of circulating lymphocytes and monocytes would benefit to fight against sars-cov- infection, which might be associated with the low morbidity of covid- in young children. to explore the metabolic changes and immune responses in the progress of covid- cases, we investigated patients with severe covid- infection. the median age of these patients was , and the clinical course was more than weeks. ct scan images showed multiple patchy ground-glass shadows in the left and right lungs. bedside chest radiography of critically ill patients indicated that the brightness of both lungs was decreased and multiple patchy shadows were observed. these clinical characteristics of severe cases are very similar to those reported in previous studies ( , ) . the severe covid- cases had ards i to iii, and had extremely high levels of ctni, ldh, and pro-bnp, a marker of severe cardiac dysfunction and even heart failure. besides that, an extraordinarily low level of serum uric acid [ µmol/l ( - )] was found in severe cases. uric acid is synthesized mainly in the liver and other tissues, which usually dissolves in the blood, and is removed from the body through urine. the extraordinarily low level of serum uric acid might indicate that potential liver and/or rental metabolism dysregulated in severe patients. among severe cases, three patients had the comorbidity of diabetes, and other patients also had very high blood glucose we further investigated immune responses in patients with severe covid- . first, different subpopulations of lymphocytes were investigated. the percentage and count of b cells and nk cells did not obviously change, which is consistent with the results from a previous report ( ) . the results of igm/igg/iga/ige, c and c also indicated that b cells and complements held normal functions. however, compared with mild cases, severe covid- cases had lower levels of cd + t cells [ /µl (iqr, - )] and an even more significant reduction in cd + t cells [only /µl (iqr, - )], which has a sharper drop than cd + t cell. we further analyzed th /th panel, in severe patients, th cytokines (il- and ifn-γ) were in the normal range, but il- , one of th cytokines, was about four times higher than normal. previous studies presented that in severe patients, cd + t cells and cd + t cells highly expressed the exhaustion markers, including nkg a, pd- , and tim- ( , ) . the dramatical decrease and functional exhaustion of cd + t cells and cd + t cells represents an important immunological characteristic of severe covid- infection. following the exhaustion of t cells, severe cases had high potential for co-infection with other pathogens. in this study, of ( %) severe patients had coinfection with fungi, and two of ( %) severe patients had bacterial co-infection. twenty-one severe cases had a high level of pct and crp, . ng/ml (iqr, . - . ) and mg/l (iqr, - ), respectively. il- was much higher than normal in severe cases, even times higher than normal in some critically ill cases. with sars-cov- replication and/or coinfection with bacteria and fungi, severe inflammatory responses played important roles in the progress of severe covid- infection. in the late stage of severe covid- , % (iqr, - ) of leukocytes were neutrophils [ . × /l (iqr, . - . ). a high number of wbc [ /µl (iqr, ] was presented in urine of severe patients. previous studies suggest that, in sepsis, a large number of neutrophil and the formation of neutrophil extracellular traps (net) triggered severe inflammatory responses and excessive tissue damage ( , , ) . the significant increase in neutrophils might be involved in severe inflammatory responses and mods, even dic and death in severe covid- patients. additionally, uric acid is the predominant anti-oxidant molecule in the plasma and respiratory tract, and is necessary for induction of type immune responses. uric acid plays a pivotal role in protecting against pathogen infections and autoimmune diseases ( , ) . whether the decrease of serum uric acid is associated with the inflammatory responses in severe covid- cases need to be explored. there are several limitations to this study. first, we investigated young children with covid- and adult cases, including mild cases and severe cases. more cases will need to be collected for comparative analysis of the difference between severe and critically ill patients. second, more inflammatory cytokines and chemokines will be analyzed for severe and critically ill patients and will be further evaluated for inflammatory storm mediated ards, dic, mods, and coagulation disorders. third, the mechanisms by which sars-cov- infection causes the reduction and functional exhaustion of cd + t cells and cd + t cells are still unclear. in-vitro and in-vivo experiments need to be performed to explore the mechanisms of t cell exhaustion. in summary, our findings suggest that extremely high level of lymphocytes and monocytes could help hamper sars-cov- replication, which might be associated with the low morbidity of covid- in infants and young children. a high number of monocytes would be helpful for removing sars-cov- and play an important role in the recovery of patients with mild covid- . in the late stage of the disease, severe cases suffered from ards, metabolic disorders, mods and coagulation disorders. with dramatical decrease of cd + t cells and cd + t cells, extraordinarily increased neutrophils and severe inflammatory responses are involved in ards, mods, and coagulation disorders and can even lead to dic and death in severe cases. whether the decrease of serum uric acid is associated with the inflammatory responses in severe covid- cases needs to be further explored. these findings can not only greatly improve our understanding of metabolic and immunological characteristics, but also provide a mechanistic basis for the prevention and treatment of covid- infection. all datasets generated for this study are included in the article/supplementary material. the studies involving human participants were reviewed and approved by renmin 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of hospitalized patients with novel coronavirus-infected pneumonia in wuhan longitudinal characteristics of lymphocyte responses and cytokine profiles in the peripheral blood of sars-cov- infected patients. medrxiv functional exhaustion of antiviral lymphocytes in covid- patients reduction and functional exhaustion of t cells in patients with coronavirus disease (covid- ) neutrophil extracellular traps in immunity and disease physiological functions and pathogenic potential of uric acid: a review barrier epithelial cells and the control of type immunity the supplementary material for this article can be found online at: https://www.frontiersin.org/articles/ . /fimmu. . /full#supplementary-material the authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.copyright © he, wang, wang, zhao, huang, tian, yang, zhang, zhang, gu, zhou and zhou. this is an open-access article distributed under the terms of the creative commons attribution license (cc by). the use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. no use, distribution or reproduction is permitted which does not comply with these terms. key: cord- -r qlbnc authors: xie, guo-hao; chen, qi-xing; cheng, bao-li; fang, xiang-ming title: defensins and sepsis date: - - journal: biomed res int doi: . / / sha: doc_id: cord_uid: r qlbnc sepsis is a leading cause of mortality and morbidity in the critical illness. multiple immune inflammatory processes take part in the pathogenesis of sepsis. defensins are endogenous antimicrobial peptides with three disulphide bonds created by six cysteine residues. besides the intrinsic microbicidal properties, defensins are active players which modulate both innate and adaptive immunity against various infections. defensins can recruit neutrophils, enhance phagocytosis, chemoattract t cells and dendritic cells, promote complement activation, and induce il- β production and pyrotosis. previous publications have documented that defensins play important roles in a series of immune inflammatory diseases including sepsis. this review aims to briefly summarize in vitro, in vivo, and genetic studies on defensins' effects as well as corresponding mechanisms within sepsis and highlights their promising findings which may be potential targets in future therapies of sepsis. sepsis, severe sepsis, and septic shock represent a continuum of clinical syndromes which are common complications observed in patients with infection, trauma, and major surgeries [ ] [ ] [ ] . these syndromes start with infection induced systemic inflammatory response syndrome (sirs) and evolve to sepsis induced acute organ dysfunction and cardiovascular collapse. epidemiology studies demonstrated that severe sepsis has a population prevalence of / in the united states and counts for - % of the intensive care unit (icu) patients [ ] [ ] [ ] . and severe sepsis has already been acknowledged as the first cause of death in noncoronary icus with a high mortality rate of approximately - % [ ] . in the past one or two decades, steady progresses in treatment of sepsis have been made due to the advanced supportive care in icu and the implementation of bundle therapies [ ] . however, searching for specific remedies and reliable predictors within the pathophysiological mechanisms of sepsis is still the emphasis of today's studies [ , ] . defensins are classified as a subfamily of cationic antimicrobial peptides, which are major components of the human innate immunity. they are small endogenous peptides with three disulphide bonds created by six cysteine residues. defensins are categorized into three subtypes, -, -, and -defensin, based on the spatial structure and the locations of three disulphide bonds within the peptide. in the past decade, cumulative evidences have suggested that defensins play an important role and may be a potential intervention target in sepsis. this review hereby will summarize in vitro, in vivo, and genetic studies on defensins' effects as well as corresponding mechanisms within sepsis and its sequential syndromes. defensins have broad spectrum antimicrobial activities against most pathogens in sepsis. [ ] [ ] [ ] . they can inhibit a large variety of gram-positive bacteria, gram-negative bacteria, and some species of fungi and viruses [ ] . the -defensins are mainly distributed in the epithelial cells of the respiratory system, digestive system, and genitourinary system [ ] [ ] [ ] . they can effectively kill a number of gram-negative bacteria, such as e. coli and p. aeruginosa, gram-positive bacteria, such as s. aureus and streptococcus pyogenes, and candida albicans. -defensin- even has bactericidal effect towards multiresistant s. aureus and vancomycin-resistant enterococcus [ , ] . the -defensins, which have a unique macrocyclic structure, are isolated from leukocytes from some species of monkey and have not been detected in humans [ ] . they are also reported to have antimicrobial activity against a spectrum of pathogens including e. coli, s. aureus, and c. albicans [ ] . also, they are found to have protective effect in a mouse model from a lethal pulmonary infection by a mouse adapted strain of sars-coronavirus [ ] . the classic mechanism of defensins' bactericidal effect is the "pore formation" theory. these positively charged antimicrobial peptides target negatively charged bacterial membrane components, such as lipopolysaccharides, teichoic acids, or phospholipids. then they form transmembrane pores, disrupt cell integrity, and lead to bacteria lysis [ , ] . recently, another mechanism has been reported that defensins kill bacteria by inhibiting the synthesis of bacterial cell wall through interaction with certain precursors such as lipid ii [ ] . defensins' bactericidal effect can be limited by high salt concentration of local environment where they encounter with the pathogens [ , ] . also, the antimicrobial action appears to be regulated by the redox response, as -defensin- become more potent after reduction of disulfide bridges by thioredoxin or a reducing environment [ , ] . defensins are also reported to have modulating effects on both innate and adaptive immune response. it is well known that hnp - participate in the host immune defense via multiple mechanisms, including enhancing macrophage phagocytosis, facilitating neutrophil recruitment, modulating complement activation, and chemoattracting immature t cells and dendritic cells [ , ] . in vitro studies showed that -defensins have potent chemotactic effects, leading to the recruitment and maturation of naive dendritic cells and memory t cells in the inflammatory sites and the triggering of specific immune response in the host [ ] . as the endogenous ligand of tlr- , -defensins interact with tlr- of the immune cells and regulate the expression of inflammatory mediators via the nf-b pathway [ ] . in vivo researches have revealed that the abnormal expression of -defensins is associated with sepsis and various infectious diseases, as levels ofdefensins in both plasma and bronchoalveolar lavage fluid in patients with pulmonary infections are elevated [ ] [ ] [ ] , transcription of -defensin- in leukocytes of severe septic patients is suppressed [ ] , expression of -defensins in burn wound is reduced [ ] , and impaired expression of -defensins is associated with inflammatory bowel diseases [ , ] . in a mouse model of acute lung injury, shu et al. expressed recombinant -defensin- in lung tissue via recombinant adenovirus to study its protective effect against p. aeruginosa infection. compared with control mice, they found considerably less p. aeruginosa in the transinfected lung tissue, as well as alleviated alveolar impairment, interstitial edema, and neutrophil infiltration [ , ] . in subsequent studies, mice transinfected by adenovirus with or withoutdefensin- genes received cecal ligation and puncture (clp) twice to generate sepsis models. the impact of -defensin- on the inflammatory response (e.g., the level of icam- expression), the severity of lung injury, and the sepsis outcome ( -day survival rate) were observed and evaluated. it was found that recombinant -defensin- could downregulate the expression of icam- in lung tissue h, h, and h after clp and significantly raised the -day survival rate in sepsis mice [ , ] . in the clinical setting, olbrich et al. found preterm neonates had lower level of -defensin- in cord blood when compared to term neonates [ ] . and among these preterm neonates, lower -defensin- level was associated with late-onset sepsis. these studies indicate that -defensin- may play an important role in the immune inflammatory response in sepsis and might influence the outcome of sepsis. among the -defensins, rhesus macaque -defensin (rtd), which has six subtypes, has been extensively studied. though not expressed in humans, rtds were reported to significantly reduce levels of tnf-, il- , il- , il- , mip , and so on, in human peripheral blood leukocytes that are preincubated with various toll-like receptor agonists [ ] . furthermore, in vivo study showed that subcutaneously administration of mg/kg rtd- could improve the survival rate and suppress the levels of a number of inflammatory cytokines and chemokines in two sepsis mouse models (received either intraperitoneal injection of e. coli or clp). although detailed mechanisms of the protective effect of rtd- have not been illuminated, the authors suggested that the interaction between rtd- and leukocyte is the critical determinant of tnf-blockade [ ] . the latter is a major proinflammatory cytokine and influences the consequent inflammatory cascades. these results indicate that -defensins may be a potential immune adjuvant in the treatment of sepsis, though they are not expressed in human. in sepsis and other inflammatory disorders, defensins are among a group of rapidly-released host endogenous molecules, which are capable of both recruiting and activating apcs and are also termed the alarmins. recently, in vitro studies have shown that alarmin hnp - have the ability to boost host inflammatory response by promoting macrophage il- production and pyroptosis via purinergic p x receptor [ ] . however, this effect is a double-edged sword in sepsis since it can promote pathogen elimination as well as mediate organ dysfunction such as acute lung injury [ , ] . in molecular genetics and molecular biology, knock-out animal model is one of the most convincing means to determine the role of a specific molecule in the physiopathology of a certain disease. however, as members of the defensin family have overlapped biological functions, the function of the knock-out gene in animal models may probably be compensated by other defensins. since the gene cluster coding for the entire defensin family cannot be fully knocked out using the present techniques of molecular biology and genetics as well as human defensins lack of absolute animal analogues, genetic association analysis is a good alternative that can effectively explore the relationship between genetic polymorphism and sepsis. in normal peripheral blood cells, mrna levels of both -defensin- and -defensin- raise remarkably when stimulated by lps or p. aeruginosa [ ] . however, the upregulation of -defensin- and -defensin- varies among individuals, resulting in interindividual differences in host defense capacity and hence influencing the clinical progression of sepsis. previous studies showed that single nucleotide polymorphism (snp) of -defensin- gene (defb ) correlates with chronic obstructive pulmonary disease, asthma, genetic allergy, hiv infection, and pseudomonas species infection in oral mucosa [ ] [ ] [ ] [ ] [ ] . since sepsis is a multifactorial disease caused by both environmental factors (pathogenic microbes) and host factors (comorbidities and genetic background), its occurrence and outcome are influenced with individual genetic background [ ] . chen et al. selected snps in the promote region of defb- (- a/g, - a/t, - a/g, - c/g, and - a/g) and one in its extron ( g/a) as candidate loci and studied patients with severe sepsis and healthy controls [ ] . distribution of alleles, gene types, and haplotypes associating with these loci were studied and compared between septic patients and controls, as well as between survivals and victims of severe sepsis. association analysis, logistic regression, and linkage disequilibrium study showed that - g allele was closely related with susceptibility to severe sepsis and poor outcome. and severe septic patients with haplotype - g/- g/- g had even poorer outcome, while individuals with haplotype - a/- c/- g were less susceptible to severe sepsis. the reason why - c/g is correlated with the occurrence and outcome of severe sepsis may attribute to the following points. it located in the untranslated region of defb and its polymorphism may result in changes in the space conformation of mrna, which would alter the stability of mrna and the efficiency of translation. and its impact on the protein function is more significant than nonsynonymous snp in coding region [ ] , as the quantity of protein would change dramatically. however, as any other genetic association analysis, defb − c/g may be only a surface marker of some unknown real genetic marker of sepsis in linkage disequilibrium. although these hypothesis need to be proved by further researches, the above-mentioned study indicated that -defensin- might be an influential factor in the process of immune defense and inflammation regulation in sepsis, and the locus of − c/g may be an important genetic warning indicator of susceptibility to severe sepsis and its outcomes. copy number variation (cnv) is a kind of genetic polymorphism that accounts for approximately % of human genomic dna. it refers to a large-scale duplication or deletion of certain dna sections, which causes a variation in the number of copies of one or more genes. previous publications reported that cnv is present in -defensin- gene (defb ), -defensin- gene (defb ), -defensin- gene (defb ), -defensin- gene (defa ), and -defensin- gene (defa ) [ , [ ] [ ] [ ] . and copy number of defb has a positive correlation with its mrna level [ , ] . recently, chen et al. screened severe sepsis and healthy controls for defa and defa [ ] . an average defa /defa copy number of per genome was observed in the studied population, with a range of to . the authors found that patients with high copy number of defa /defa were predisposed to severe sepsis and tended to have lower level of plasma hnp - as well as cytokines such as tnf-, il- , and il- . they further validated their findings in an independent cohort. these results indicated that cnvs in the defensin gene may be potential genetic markers for identifying high risk patients or providing individual treatment in sepsis. defensins are emerging therapeutic molecules against pathogens in sepsis because of their broad spectrum antimicrobial properties. in the past decade or two, a number of potent and salt insensitive defensins and their analogs have been screened, structurally modified, and synthesized. however, most of these studies are performed in vitro and not much is known about the in vivo roles of these molecules. in fact, chemoattracting and immunomodulating effects make defensins a double-edged sword in the pathogenesis of sepsis, which leads to facilitation of pathogen clearance as well as exacerbation of inflammation and injury of self-tissues. recently, several investigations showed that the chemoattractant and antimicrobial activities of defensins could be separated, which shed light on the design of defensin-derived pharmaceuticals [ ] . in addition, genetic studies help identify high risk patients with susceptibility to sepsis or its adverse outcome, which provides foundation for future individualized sepsis treatments that are targeting defensins. the pathophysiology and treatment of sepsis management of sepsis the immunopathogenesis of sepsis the epidemiology of sepsis in the united states from epidemiology of severe sepsis in critically ill surgical 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novel antibiotics targeting lipid ii defensins: antimicrobial peptides of innate immunity isolation and characterization of human -defensin- , a novel human inducible peptide antibiotic reduction of disulphide bonds unmasks potent antimicrobial activity of human -defensin cellmediated reduction of human -defensin : major role for mucosal thioredoxin amped up immunity: how antimicrobial peptides have multiple roles in immune defense multiple roles of antimicrobial defensins, cathelicidins, and eosinophil-derived neurotoxin in host defense increased levels of antimicrobial peptides in tracheal aspirates of newborn infants during infection elevated balf concentrations of -and -defensins in patients with pulmonary alveolar proteinosis expression and activity of -defensins and ll- in the developing human lung inducibility of the endogenous antibiotic peptide -defensin is impaired in patients with severe sepsis reduced antimicrobial peptide expression in human burn wounds a chromosome gene-cluster polymorphism with low human geta-defensin gene copy number predisposes to crohn disease of the colon hammarström, " -defensin- and - in intestinal epithelial cells display increased mrna expression in ulcerative colitis protection against pseudomonas aeruginosa pneumonia and sepsis-induced lung injury by overexpression of -defensin- in rats the protective effect of beta-defensin in acute lung injury induced by respiratory pseudomonas aeruginosa infection in rats the effect of pretreatment of recombinant beta-defensin on the icam- expression in lung of rats with acute lung injury association of human beta-defensin- serum levels and sepsis in preterm neonates rhesus macaque theta defensins suppress inflammatory cytokines and enhance survival in mouse models of bacteremic sepsis alarmin hnp- promotes pyroptosis and il- release through different roles of nlrp inflammasome via p x in lps-primed macrophages neutrophil alpha-defensins cause lung injury by disrupting the capillaryepithelial barrier genetic variants of human -defensin- and chronic obstructive pulmonary disease association of defensin beta- gene polymorphism with asthma a single-nucleotide polymorphism in the human beta-defensin gene as associated with hiv- infection in italian children single-nucleotide polymorphisms (snps) in humandefensin : high-throughput snp assays and association with candida carriage in type i diabetics and nondiabetic controls contribution of alpha-and beta-defensins to lung function decline and infection in smokers: an association study genomic polymorphisms in sepsis genomic variations within defb are associated with the susceptibility to and the fatal outcome of severe sepsis in chinese han population human catechol-o-methyltransferase haplotypes modulate protein expression by altering mrna secondary structure extensive normal copy number variation of a -defensin antimicrobialgene cluster copy number polymorphism and expression level variation of the humandefensin genes defa and defa screening of copy number polymorphisms in human -defensin genes using modified real-time quantitative pcr increased genomic copy number of defa /defa is associated with susceptibility to severe sepsis in chinese han population analysis and separation of residues important for the chemoattractant and antimicrobial activities of beta-defensin the authors declare that there is no conflict of interests regarding the publication of this paper. this work was supported by the national natural science foundation of china (no. ) and the research project of the department of education of zhejiang province (no. y ). key: cord- -h izji g authors: wei, yuan-yuan; wang, rui-rui; zhang, da-wei; tu, you- hui; chen, chang- shan; ji, shuang; li, chun-xi; li, xiu-yong; zhou, meng-xi; cao, wen- sheng; han, ming- feng; fei, guang-he title: risk factors for severe covid- : evidence from hospitalized patients in anhui, china date: - - journal: j infect doi: . /j.jinf. . . sha: doc_id: cord_uid: h izji g nan dysfunction ( , ( ) ( ) ( ) . due to a previous lack of in-depth research on the characteristics of the disease, the mortality of severe illness is high( ). it is very important to analyse the clinical characteristics of covid- in international regions and identify risk factors to reduce the incidence of severe and critical illness in the early stage. in this letter, we present discrepancies of patients with different disease severities and risk factors for severe covid- by comparing and analysing epidemiological and clinical data of confirmed patients in anhui, china. in the present study, the rate of severely ill patients was as high as . %. comparisons of demographics and clinical characteristics between severe and non-severe patients are shown in table . the mean age was . years in severe patients and . years in non-severe patients, with a significant difference (p= . ). there were males ( . %) and females ( . %); however, there was no significant differences in sex between the two groups. among patients, had fever ( . %), had cough ( . %) and had shortness of breath ( . %). the prevalence of shortness of breath was . % in severe patients, which was significantly greater than the . % prevalence in non-severe patients (p< . ). there were patients ( . %) with comorbidities, of which had multiple comorbidities ( . %). among patients with diabetes, severe cases were significantly more common than in non-severe patients (p< . ). compared to non-severe patients, fingertip oxygen saturation decreased significantly in severe patients (p< . ), which predisposed patients with chronic obstructive pulmonary disease to acute exacerbation. since sars-cov- affected multiple organs by binding angiotensin converting enzyme (ace ) receptor( ) and many severe covid- patients had comorbidities, multidisciplinary team (mdt) consultation played an important role in reducing the mortality of severe infection. there were significant differences in the use of mechanical ventilation, glucocorticoids and immunoglobulin between severe and non-severe patients (all p< . ). table presents comparisons of laboratory parameters between severe and non-severe patients. lymphocyte, cd and cd cell counts were decreased significantly in severe patients compared to non-severe patients (p= . , . and . ), suggesting that t lymphocytes were seriously destroyed. the increased level of c-reactive protein (crp) in severe patients was significantly higher than that in non-severe patients (p= . ). interleukin- (il- ) levels increased in patients ( . %); the increase was more significant in severe patients than in non-severe (p= . ). by clearing or blocking inflammatory factors( ), artificial liver therapy and tocilizumab, a monoclonal antibody of il- receptor, may prevent serious injuries in the lungs in severe patients. the lactate dehydrogenase (ldh) concentration was higher and the albumin concentration was lower in severe patients, with significant differences (p= . and p< . ). in severe patients, the fibrinogen concentration was significantly higher (p= . ) than in non-severe patients, suggesting that severe patients were more likely to experience myocardial infarction or sudden death. between the two groups, there was a significant difference in the neutrophil to lymphocyte ratio (nlr), a predictor for severe infection( ) (p= . ). other laboratory parameters that changed in covid- patients were not significantly different between the two groups (all p> . ). there are still no specific therapies for covid- ( ); nevertheless, assessing risk factors and symptomatic treatment in the early stage of the disease can improve the prognosis. of patients, . % received initial antiviral therapy using lopinavir and ritonavir ( mg/ mg, bid) for no more than days and/or atomized inhalation of interferon-α except for a -year-old patient, who was given interferon-α only, and . % of patients received mg methylprednisolone for to days. a total of . % of patients were supported with immunoglobulins, and . % of patients were managed with suitable oxygen therapy. all severe patients received mdt consultation, and three critical patients were treated with artificial liver therapy every other day, three times consecutively. finally, all patients in our study, both severe or non-severe, improved and were discharged without death. the clinical characteristics of covid- patients are summarized in tables and . the similarities and differences between severe and non-severe patients in this letter suggested that elderly patients with multiple comorbidities, hypoxia, decreased cd and cd cell counts and increased levels of crp and il- are all closely associated with disease severity and prognosis, which should be assessed seriously during diagnosis and treatment. a rapid decline in t lymphocytes and significant increases in the levels of inflammatory factors, including crp and il- , can be clinical warnings of severe infection. mdt consultation and artificial liver therapy are very effective methods for severe patients with covid- . with these integrated prevention and treatment strategies, there will be a good prognosis for covid- . a. mono-factor logistic regression analysis was used to analyse risk factors for severe illness using r software; b. multi-factor logistic regression analysis was used to analyse independent risk factors for severe illness using r software. notes: p< . indicates a risk factor in figure a and an independent risk factor in figure b for severe patients. coronavirus -ncov: a brief perspective from the front line we thank the medical workers fighting against covid- for their hard work and sincere responses to our information requests. funding. this work was funded by the key technology research and development program of anhui province (no: h ). key: cord- -mhs yext authors: simadibrata, daniel martin; pandhita, bashar adi wahyu; ananta, muammar emir; tango, tamara title: platelet-to-lymphocyte ratio, a novel biomarker to predict the severity of covid- patients: a systematic review and meta-analysis date: - - journal: j intensive care soc doi: . / sha: doc_id: cord_uid: mhs yext platelet-to-lymphocyte ratio (plr), a novel inflammatory marker, has been suggested to predict the severity of covid- patients. this systematic review aims to evaluate the association between plr levels on admission and the severity of covid- patients. a systematic literature search was done on july to identify peer-reviewed studies, preprints, and grey literatures. research articles comparing the plr value on admission in adult patients with covid- with varying degrees of severity were included in the analysis. the following keywords were used for the search: “covid- ”, “plr”, “severity”, and “mortality”. a total of seven studies were included in the meta-analysis, six of which were conducted in china. from a total of participants included, ( . %) had severe diseases; and those in the severe group were generally older and had underlying diseases compared to the non-severe group. in comparison to non-severe patients, the meta-analysis showed that severe covid- patients had higher plr levels on admission (smd . ; %ci . - . ; i( ) = %). high plr levels on admission were associated with severe covid- cases. therefore, the on-admission plr level is a novel, cost-effective, and readily available biomarker with a promising prognostic role for determining the severity of covid- patients. coronavirus disease (covid- ) is a disease caused by the severe acute respiratory syndrome coronavirus- (sars-cov- ), a virus thought to start as a zoonotic infection in wuhan in late december . the disease was declared by the world health organization (who) as a pandemic on march and has infected more than countries worldwide. as of july , a total number of cases and deaths attributed to covid- were recorded, only months after its emergence. covid- is known for being infectious and simultaneously manifesting in different organs aside from the pulmonary system. [ ] [ ] [ ] patients infected with covid- present a wide range of clinical conditions -ranging from asymptomatic infections, minimal symptoms to fatal respiratory distress. although the majority of covid- cases were classified as mild, involving flu-like symptoms to mild pneumonia, up to % of mild/moderate cases progressed to acute respiratory distress syndrome (ards). additionally, patients with relatively normal clinical conditions can rapidly deteriorate and worsen within a few days, making clinical presentation an unreliable prognostic predictor of covid- . thus, a more objective indicator is required to accurately assess and stratify the prognosis of covid- patients upon admission to healthcare services. immunological studies have shown that high levels of proinflammatory cytokines, known as a cytokine storm, are the hallmark characteristic of severe covid- cases. this extreme elevation of cytokines causes a massive proinflammatory response resulting in multiple organ dysfunction syndrome (mods) and ards, which subsequently leads to mortality in covid- patients. therefore, in theory, inflammatory markers can be used to assess the severity and mortality risk of covid- patients. platelet-to-lymphocyte ratio (plr) is a novel marker of inflammation, which is inexpensive and readily available in clinical settings. plr has been used in various diseases, such as cardiovascular diseases and autoimmune diseases, as a predictor of inflammation and mortality. , due to the rapid involvement of inflammatory processes in covid- , severe covid- patients have demonstrated elevated plr levels on admission. , this suggests the potential use of this inflammatory marker to determine the prognosis of covid- patients, especially in resource-limited settings. therefore, this systematic review aims to review the prognostic value of plr levels on admission to determine the severity and mortality of covid- patients. this systematic review was written in compliance with the preferred reporting items for systematic reviews and meta-analyses (prisma) checklist (table s ). prior to the writing of this systematic review, a protocol was formulated and registered in the international prospective register of systematic reviews (prospero) on june (crd ). we included cohort studies evaluating the difference in plr levels on admission in adults (> years old) with confirmed covid- (diagnosed using rt-pcr) categorized based on disease severity (severe and non-severe patients), and/or mortality (survivor and non-survivor). case series, correspondences, review articles, non-research articles, and letters to the editor were excluded from the study. we only included papers written and published in english. otherwise, no other exclusion criteria were applied. the severity of the disease was defined by the who-china joint mission on covid- report. severe covid- was defined as patients that met any of the following criteria: respiratory frequency ! x/ minute, blood oxygen saturation %, pao /fio ratio < , and/or lung infiltrates > % of the lung field within - hours. the keywords used in the search strategy were derived from the following key concepts "covid- ", "platelet-to-lymphocyte", "severity", and "mortality"; and were adapted to the respective databases (table s ) . a systematic literature search was finalized on july to identify peer-reviewed papers published in four databases (ovid medline, embase, scopus, and the cochrane library). additionally, manual handsearching was done for preprints in two databases (medrxiv and ssrn), and for grey literatures in two databases (who covid- global research database and center for disease control and prevention covid- research article). we also performed forward and backward tracing of references from relevant articles to identify additional papers missed from database searching. all articles retrieved from the searches were exported to endnote x reference manager. after deduplication of articles, publications were screened based on its titles and abstracts, and the remaining publications were screened according to the full text. this study selection process was carried out by two independent reviewers (dms and baw). any disagreement regarding the study selection was resolved by the inclusion of a third party. all included articles' data were extracted by two independent reviewers (dms and baw). the risk of bias assessment was done using the newcastle ottawa scale (nos). a standardized data extraction form was used to gather relevant data from the selected articles. data extracted included: first author and publication year, publication type, country of study, study design, baseline characteristics of patients (age, gender, and underlying diseases), and the outcome of the study (plr values). for the quantitative analysis, we exported quantitative data from all eligible studies to review manager software . (cochrane collaboration) and performed a meta-analysis. we extrapolated the mean and standard deviation of studies with non-normal data using the available median and interquartile range (iqr), according to hozo et al. the inverse variance method was used to obtain the effect estimate in the form of a standardized mean difference (smd) and the % confidence interval (ci). the statistical heterogeneity between the studies was assessed using cochrane chi-square and i . if there was significant heterogeneity (i > %), the random-effects model was used to calculate the pooled effect size; otherwise, the fixed-effects model was used. funnel plot analysis was used to qualitatively evaluate the risk of publication bias by comparing the smd with the standard error of the natural log of smd. we performed a sensitivity analysis by omitting one study at a time to identify the source of heterogeneity. all p-values were two-tailed and statistical significance was considered if p < . . the literature search from electronic databases resulted in publications, four of which were identified through manual handsearching of relevant literature ( figure ). following de-duplication, articles were screened for its titles and abstracts. the remaining ten articles were reviewed for its fulltext, and three articles were excluded, leaving a total of seven peer-reviewed articles included in this systematic review. from a total of seven selected studies, all were peer-reviewed and were retrospective observational studies. overall, six studies were done in china , , - and only one was done outside of china, in turkey. all studies compared the on-admission plr values between severe and nonsevere covid- patients, except for one that compared the plr values between severe and moderate covid- patients (table ) . unfortunately, no study explored the prognostic value of plr on admission to predict mortality. the quality of the included studies is shown in table s . four studies had seven nos stars, and the other three had six nos stars. in brief, all studies were of acceptable quality and eligible for inclusion in the meta-analysis. a total of participants were included from all the studies, ( . %) of which had severe disease. although four studies did not report the baseline comorbidity characteristics of covid- patients, in comparison to non-severe covid- a pooled effect size meta-analysis was conducted using the random-effects model (n ¼ ; severe-¼ , non-severe ¼ ) (figure ) . overall, patients with severe covid- had a higher plr value on admission compared to non-severe covid- (smd . ; %ci . - . ). there was significant heterogeneity among the studies (i ¼ %, p ¼ . ). the funnel plot was visually asymmetrical and indicated a potential risk of publication bias ( figure s ). sensitivity analysis by sequentially removing one study at a time did not significantly change the heterogeneity among the studies and the overall pooled effect size. however, the exclusion of the study by yang et al. resulted in no significant heterogeneity between the studies (i ¼ %, p ¼ . ) ( figure s ). the pooled effect size meta-analysis using the fixedeffects model showed a significantly higher plr value on admission in severe covid- patients than non-severe covid- patients (smd . ; %ci . - . ). as of july , covid- has infected approximately million people worldwide, thousand of whom died. more concerning, those with severe covid- rapidly deteriorate to critical cases, which involve multiorgan failure, leading to death. thus, there is an urgent need for healthcare providers to develop readily available biological markers to predict the severity and mortality of covid- at the early stage of the disease to provide the most optimal management. plr was initially suggested as an excellent candidate marker for determining the severity and mortality of covid- . first, plr is an established marker of inflammation. inflammation plays a considerable role in the pathophysiology of covid- , with cytokine storm as a hallmark condition in severe disease and poorer prognosis. thus, elevated plr value suggests an overactive inflammatory response and subsequently, worse prognosis. second, plr is sensitive to natural and acquired immune response. third, plr is an inexpensive and readily available measurement that can be used in resource-limited settings. therefore, our systematic review aims to review the validity of the plr level on admission as a prognostic indicator in covid- patients. our meta-analysis, which included a total of covid- patients, showed that high plr value was associated with severe covid- . six out of the seven included studies demonstrated similar results with increased plr on admission found in severe cases of covid- compared to those with mild or moderate diseases. this suggests that elevated plr on admission among severe covid- patients reflects a higher degree of the cytokine storm. this evidence can be useful for providing specialized treatment to patients with severe covid- , as they might require more prolonged hospital admissions. although plr on admission was generally shown to be increased in severe vs. non-severe covid- , qu et al. reported a reduced plr on admission in severe compared to non-severe covid- despite no statistical significance. inconsistencies between the results of different studies could be problematic, especially for daily clinical applications. however, we noted the rather small sample size of the study. only three severe cases of covid- were included in the analysis compared to non-severe cases, which could lead to this contradictory result. furthermore, the follow-up analysis of plr in the study showed elevated levels in severe covid- cases upon hospitalization. to date, there is no universal laboratory reference value for plr, especially for covid- patients. of all the included studies, only two studies attempted to determine the optimal cut-off plr value. yang et al. reported the optimal cut-off plr value as with auc of . , specificity of %, and sensitivity of %. meanwhile, sun et al. suggested a cut-off plr value of . with auc of . , specificity of . %, and sensitivity of . %. this discrepancy warrants the need for further research to determine the most appropriate plr cut-off value in determining the severity of covid- patients. as plr involves a comparison between the absolute platelet and absolute lymphocyte count, a comprehensive understanding of the role of platelets and lymphocytes in covid- is important. previous meta-analyses have reported that thrombocytopenia and lymphopenia were both associated with severe covid- cases. [ ] [ ] [ ] however, until the writing of this manuscript, the underlying mechanism by which high plr levels were reported in severe covid- cases remains unclear. a plausible explanation could be that the decrease in absolute lymphocyte counts was much more significant than the decrease in platelet counts, thus increasing the plr value. three hypotheses may explain the underlying decrease in platelet count in covid- patients. first, the cytokine storm triggered by sars-cov- can decrease the synthesis of platelets by destroying bone marrow progenitor cells. sars-cov- is also postulated to directly affect the production of platelets in the bone marrow. second, sars-cov- can induce the generation of autoantibody and immune complex, which may trigger the destruction of platelets. third, platelets activated during lung injury could be aggregated and be overactively involved in microthrombus formation. on the other hand, the underlying mechanism for the decreased absolute lymphocyte count is that sars-cov- triggers pyroptosis in lymphocytes through the activation of nlrp inflammasome. furthermore, another hypothesis points out the role of the proinflammatory cytokine il- that utilizes the lymphocytes; hence, decreased lymphocyte counts are associated with poor prognosis in covid- patients. we also observed a significant heterogeneity among the included studies in this meta-analysis. the possible reasons for the high heterogeneity could be due to the distinct interstudy baseline characteristics of the subjects, different number, and proportion of patients with comorbidities as well as the proportion of severe and non-severe cases. fesih, qu, and sun excluded patients with other comorbidities, such as chronic lung diseases, hematological diseases, and liver diseases , , while gong did not provide any data on the comorbidities of the patients. another reason could be due to the small number of included studies in this meta-analysis, which could reduce the accuracy of the heterogeneity analysis. we performed a sensitivity analysis by sequentially omitting one study at a time and determined that yang et al. was the source of heterogeneity. however, with the removal of yang et al., the pooled analysis still showed a statistically significant higher plr value on admission in severe covid- in comparison to non-severe covid- patients. this meta-analysis is not without limitations. we acknowledge that only including articles written and published in english would disregard those written in other languages and present with a geographical bias. moreover, most of the included studies were from china, whereas the majority of confirmed cases and deaths were located in the usa and europe. the variability in plr values between different populations could limit the relevance of this finding. based on the funnel plot, we also identified a potential risk of publication bias. in addition, the limited data presented by the included studies did not allow further stratification of the severe group into severe and critically ill patients. therefore, further research still needs to be conducted to determine an optimal cut-off value for plr value to predict severity in covid- . our meta-analysis showed that plr could be used as a novel, cost-effective, and readily available biomarker in determining the severity of covid- patients. our finding implies that elevated plr levels on admission can be utilized as a prognostic indicator of severity in covid- patients, especially in resource-limited settings, where there is an urgent need to effectively allocate medical resources and divert attention to patients with poorer prognosis. however, further studies are needed to determine the cut-off value of plr with the most optimal sensitivity and specificity prior to adaptation in clinical practice. dms and bawp: idea formulation, article draft writing, data collection and analysis, interpretation of the data, critical review of the writing; tt: article draft writing, data collection, interpretation of the data, critical review of the writing; mea: article draft writing, critical review of the writing. all authors contributed substantially to the writing of this manuscript and have consented to this submission. the epidemiology and pathogenesis of coronavirus disease (covid- ) outbreak world health organization cardiovascular complications in covid- covid- can present with a rash and be mistaken for dengue characteristics of ocular findings of patients with coronavirus disease (covid- ) in hubei province, china clinical management of adult coronavirus infection disease (covid- ) positive in the setting of low and medium intensity of care: a short practical review covid- pathophysiology: a review association of coronavirus disease (covid- ) with myocardial injury and mortality the platelet-to-lymphocyte ratio as an inflammatory marker in rheumatic diseases platelet-to-lymphocyte ratio is associated with prognosis in patients with coronavirus disease- the diagnostic and predictive role of nlr, d-nlr and plr in covid- patients report of the who-china joint mission on coronavirus disease geneval world health organization estimating the mean and variance from the median, range, and the size of a sample a tool for early prediction of severe coronavirus disease (covid- ): a multicenter study using the risk nomogram in wuhan and guangdong, china clinical value of immuneinflammatory parameters to assess the severity of coronavirus disease abnormalities of peripheral blood system in patients with covid- in wenzhou, china covid- is distinct from sars-cov- -negative community-acquired pneumonia predictive values of blood urea nitrogen/creatinine ratio and other routine blood parameters on disease severity and survival of covid- patients multiple organ dysfunction in sars-cov- : mods-cov- platelet to lymphocyte ratio as a novel indicator of inflammation is correlated with the severity of metabolic syndrome: a single center large-scale study pathological inflammation in patients with covid- : a key role for monocytes and macrophages value of neutrophil to lymphocyte and platelet to lymphocyte ratios in pneumonia lymphopenia in severe coronavirus disease- (covid- ): systematic review and meta-analysis thrombocytopenia is associated with severe coronavirus disease (covid- ) infections: a meta-analysis hematologic, biochemical and immune biomarker abnormalities associated with severe illness and mortality in coronavirus disease (covid- ): a meta-analysis cell pyroptosis, a potential pathogenic mechanism of -ncov infection the heterogeneity statistic i( ) can be biased in small meta-analyses none. all data generated or analyzed during this study are included in the published article and its supplementary information files. the corresponding author (dms) can be contacted for more information. not applicable. the author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article. not applicable. the author(s) received no financial support for the research, authorship, and/or publication of this article. daniel martin simadibrata https://orcid.org/ - - - supplemental material for this article is available online. key: cord- -cco vh f authors: vultaggio, alessandra; agache, ioana; akdis, cezmi a.; akdis, mubeccel; bavbek, sevim; bossios, apostolos; bousquet, jean; boyman, onur; chaker, adam m.; chan, susan; chatzipetrou, alexia; feleszko, wojciech; firinu, davide; jutel, marek; kauppi, paula; klimek, ludger; kolios, antonios; kothari, akash; kowalski, marek l.; matucci, andrea; palomares, oscar; pfaar, oliver; rogala, barbara; untersmayr, eva; eiwegger, thomas title: considerations on biologicals for patients with allergic disease in times of the covid‐ pandemic: an eaaci statement date: - - journal: allergy doi: . /all. sha: doc_id: cord_uid: cco vh f the outbreak of the sars‐cov‐ ‐induced coronavirus disease (covid‐ ) pandemic re‐shaped doctor‐patient interaction and challenged capacities of healthcare systems. it created many issues around the optimal and safest way to treat complex patients with severe allergic disease. a significant numberof the patients are on treatment with biologicals and clinicians face the challenge to provide optimal care during the pandemic. uncertainty of the potential risks for these patients is related to the fact that the exact sequence of immunological events during sars‐cov‐ is not known. severe covid‐ patients may experience a “cytokine storm” and associated organ damage characterized by an exaggerated release of proinflammatory type and type cytokines. these inflammatory responses are potentially counteracted by anti‐inflammatory cytokines and type responses. this expert based eaaci statement aims to provide guidance on the application of biologicals targeting type inflammation in patients with allergic disease. currently, there is very little evidence for an enhanced risk of patients with allergic diseases to develop severe covid‐ with studies focusing on severe allergic phenotypes lacking. at present, non‐infected patients on biologicals for the treatment of asthma, atopic dermatitis, chronic rhinosinusitis with nasal polyps or chronic spontaneous urticaria should continue their biologicals targeting type inflammation via self‐application. in case of an active sars‐cov‐ infection, biological treatment needs to be stopped until clinical recovery and sars‐cov‐ negativity is established and treatment with biologicals should be re‐initiated. maintenance of add‐on therapy and a constant assessment of disease control, apart from acute management is demanded. the outbreak leading to the pandemic of sars-cov- -induced coronavirus disease has pushed health care systemsto the limits of their capacity across the globe. this infection can cause severe respiratory illness and multi-organ failure with clinical presentations greatly resembling sars-cov- and mers-cov, resulting in intensive care unit (icu) admission and high mortality. we discuss immunological and clinical considerations for patients on biologic agents (biologicals)targeting the type inflammatory response due to difficult-to-treat allergic diseases in the context of covid- . both innate and adaptive immune responses participate in anti-viral immunity. the interactions between sars-cov- and both arms of the immune system have been poorly clarified until now, particularly in the view of asymptomatic individuals, patients with mild disease, and those who fully recover. natural killer cells are involved in control of the acute phase of the viral infection, whereas cd + t cells are the key player in the following steps. antibody-secreting cells and t follicular helper cells are instrumental in the production of specific anti-viral iga, igm and igg antibodies early on. [ ] [ ] [ ] antibody-dependent macrophage activation as well as lymphocyte and macrophage pyroptosis (an excessive form of inflammatory cell apoptosis) might occur and contribute to more severe tissue damage, as described in sars-cov infection. [ ] [ ] [ ] [ ] among mediators, type i interferons (type i ifn) play a central role. in other coronavirus infections such as severe acute respiratory syndrome (sars), type i ifn are critical for the initiation of immune response and virus clearance. delayed production of type i ifn and aninsufficient cytotoxic responseis associated with a more severe clinical disease. observations from sars or middle east respiratory syndrome (mers) , and, more recently, covid- patients suggest an overshooting immune response in severe cases with wide-spread lung damage and disease aggravation around - days after onset. those severe covid- patients may also experiencea picture of a so-called "cytokine storm" and associated organ damage, particularly acute respiratory distress syndrome, acute kidney and liver failure, myocarditis and disseminated vascular coagulation. these manifestations are characterized byan exaggerated release of proinflammatory this article is protected by copyright. all rights reserved cytokines, such as il- , il- , il- , tnf- and many more ( figure ).consequently, these highly increased proinflammatory cytokines are believed to be potential targets for biological therapy.these type and type -driven inflammatory responses are counteracted by antiinflammatory cytokines,such as il- and tgf-, as well as potentially type responses. moreover, eosinophils have been reported to play a role in virus response. lower eosinophil counts were reported in association with severe cases, while an elevated eosinophil count was associated with a better prognosis although no functional relationship has been established so far and this finding may be an epiphenomenon. thus, probably all shades of cytokine responses (type and type , type , and regulatory cytokines) are required in the healing of sars-cov- infection. an appropriate induction and downregulation of individual response batteries is necessary to achieve an efficient viral clearance, an avoidance of excessive inflammatory reaction and irreversible tissue damage ( figure ). in line with a paucity of mechanistic data on covid- in the context of type inflammation, knowledge on the disease course in patients treated with biologicals targeting type inflammation due to severe asthma or other atopic diseases, such as csu, ad and crswnp, is scarce to absent. to our knowledge by april th , only studies presented disease characteristics of sars-cov- infection on patients with allergy or atopic diseases as a co-morbidity (table ) . while, in a study including patients infected with sars-cov- and admitted to icus of lombardy, italy, asthma was not referred to as a specific comorbidity and grouped under "others". allergic disease seemed to have no influence on presented symptoms and the course of the disease. [ ] [ ] [ ] [ ] none of these patients were on biologicals to treat their pre-existing allergic disease. in a recent report from the covid- -associated hospitalization surveillance network based on data from us states from march st - th % of hospitalized covid patients had asthma as a comorbidity. the highest percentage was in the - years old patient group with . % asthmatics. no information on severity of the disease and therapy has been provided. this supports the importance of a prospective assessment of atopic diseases in the context of covid . this article is protected by copyright. all rights reserved in the past years, new biological therapies for severe asthma, atopic dermatitis (ad), chronicrhinosinusitis with nasal polyps (crswnp) and chronic spontaneous urticaria (csu) have been developed targeting different aspects of the type immune response. [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] anti-il- monoclonal antibodies (mepolizumab and reslizumab) are approved for severe asthma with peripheral eosinophilia, uncontrolled under high intensity treatment. benralizumab, a monoclonal antibody that binds to the α subunit of il- receptor (il- rα) was also recently approved for uncontrolled eosinophilic severe asthma. dupilumab, a monoclonal antibody directed against the α subunit of the il- receptor (il- rα) acting as a dual antagonist of both il- and il- was approved for uncontrolled severe type asthma, moderate to severe ad and crswnp. omalizumab, a humanized monoclonal anti-ige antibody, has also been approved for igemediated persistent allergic asthma and csu. the low number of reports of patients on biologicals targeting type disease is encouraging since type diseases may predispose patients to viral infections due to compromised barriers. [ ] [ ] [ ] [ ] [ ] consequently, epidemiologic evidence closely links virus infections to both development and exacerbation of allergic diseases. [ ] [ ] [ ] the infection and persistence of respiratory viruses is attributed to impaired innate immune responses and a predisposition to mount strong type immune responses. in line with this argumentation some of these drugs provided evidence for a this article is protected by copyright. all rights reserved reduction of viral infections in asthmatics such as anti-ige treatment with omalizumab. itmay cause anti-inflammatory and immunomodulatory effects by restoring the capacity of human plasmacytoid dendritic cells (pdcs) to produce ifn-α, increasing antiviral activity and reducing viral-induced asthma exacerbations , .in severe asthma, clinical trials showed that rates of respiratory infections (upper respiratory tract infection, viral upper respiratory tract infection, influenza) were lower or similar in the anti-il- monoclonal antibody (mab)-and dupilumabtreated groups compared to placebo ( table ) . no data are available on the impact of anti-il- mab and dupilumabon virus-induced exacerbations and antiviral responses.for dupilumab, anincreased risk of herpes-virus reactivation has been reported in real-life uncontrolled studies and case reports. the pathogenesis of cytokine storm-related tissue injury has been repeatedly reported in covid- , dominated by proinflammatory type and type associated cytokines and linked inflammasome activationand neutrophilia. it has been reported that type response and treg response can antagonize these effects and may be beneficial. in this context, the inhibition of type response in severe and critical covid- cases may cause an aggravation of the disease. therefore, such biologicals should be discontinued in very severe disease. due to their long in vivo half-life in the range of a few weeks, it remains unclear to which extent such an action would impact the acute management and what the risk of losing disease control and co-morbidity later on could be. recent systematic reviews on approved biologicals in severe asthma showed that biologicals targeting il- -signalling pathway (mepolizumab, reslizumab and benralizumab) slightly increase drug-related adverse events (ae) in severe eosinophilic asthma. for anti-ige (omalizumab) and anti-il- rα (dupilumab) treatments rate ratios were rather small. benralizumab and omalizumab showed an increase in aes with low to moderate certainty in severe allergic asthma. there was an increased rate of dupilumab-related aes (low certainty) in severe asthma. data from clinical trials demonstrated good safety profiles of biologicals with regards to viral infections of the upper respiratory tract ( table ) . [ ] [ ] [ ] practical and clinical recommendations time restrictions did not allow for official guidelines to be published so far. however, several societies issued statements on the use of biologicals during the covid- pandemic (table ) . a consensus-based ad-hoc expert panel of allergy/immunology specialists from the us and canada this article is protected by copyright. all rights reserved recommends continuing administration of biologicals in patients with proven efficacy and converting the patient to a prefilled syringe for potential home administration if this is available or otherwise in-office application can occur with a plan to transition to home administration. initiation of biologic therapy for ad should be weighed very carefully, but it remains a viable option as this is administered at home. in a recent communication, the european task force on atopic dermatitis (etfad) suggested that targeted treatment selectively interfering with type inflammation, such as dupilumab, is not considered to increase the risk for viral infections and might thus be preferred compared to immunosuppressive treatments such as cyclosporine in a situation such as the covid- pandemic, although stressing that this theoretical advantage is not supported by robust clinical data. this article is protected by copyright. all rights reserved in case of hospital admission for moderate, severe or critical sars-cov- infection,management of the allergic disease should be in accordance with current guidelinesby involving the respective subspecialties. in particular for asthma inhalation therapy use preferably metered dose inhalations with chambers that are not to be shared and pulmonary function tests should be performed only if highly necessary (figure ). once resolution/recovery of the disease is established (e.g. via a negative sars-cov- test)but no shorter than weeks post onset of the disease/positive testing, the re-administration of the biological should be re-initiated ( figure ). in conclusion, current evidence does not suggest a higher risk for severe covid- in allergic individuals but data that allows estimating the risk of severe allergic phenotypes in case of sars-cov- infection is missing. treatment of patients on biologicals targeting type inflammation in allergic disease should be maintained in non-infectedindividuals. in case of an infection withholding the treatment is recommended until recovery. additional data on those patients with more severe phenotypes will provide more insight to define more precisely the risk profile of individuals with allergic disease who are of elevated risk.the collection of such data is imperative for future data-informed adaptations of these guidelines. this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved non-infectedpatients on biologicals for the treatment of asthma, ad, crswnp or csu should continue their biologicalstargeting type inflammation via self-application. in case of an active sars-cov- infection and moderate-to-severe covid- ,biological treatment needs to be stopped until clinical recovery and sars-cov- negativity is established. thereafter treatment with biologicals can be re-initiated. this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved according to who patients with chronic lung disease (e.g. such as asthma) may be prone to more severe disease. innate and adaptive immune responses to viral infection and vaccination dysregulated type i interferon and inflammatory monocyte-macrophage responses cause lethal pneumonia in sars-cov-infected mice breadth of concomitant immune responses prior to patient recovery: a case report of non-severe covid- profiling early humoral response to diagnose novel coronavirus disease (covid- ) pathogenic human coronavirus infections: causes and consequences of 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severe, uncontrolled asthma: results of the bora extension study position statement of expert panel of the polish allergology society on the management of patients with bronchial asthma and allergic diseases during sars-cov- pandemics this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved key: cord- - p g p authors: peiris, j s m; guan, y; yuen, k y title: severe acute respiratory syndrome date: - - journal: nat med doi: . /nm sha: doc_id: cord_uid: p g p severe acute respiratory syndrome (sars) was caused by a previously unrecognized animal coronavirus that exploited opportunities provided by 'wet markets' in southern china to adapt to become a virus readily transmissible between humans. hospitals and international travel proved to be 'amplifiers' that permitted a local outbreak to achieve global dimensions. in this review we will discuss the substantial scientific progress that has been made towards understanding the virus—sars coronavirus (sars-cov)—and the disease. we will also highlight the progress that has been made towards developing vaccines and therapies the concerted and coordinated response that contained sars is a triumph for global public health and provides a new paradigm for the detection and control of future emerging infectious disease threats. of these deletions, however, is not clear. similarly, sars-cov in individuals before february was genetically more diverse than the later isolates , , . the spike protein (the viral surface glycoprotein which mediates viral attachment and entry into the cell; fig. ) of early isolates contained higher rates of nonsynonymous mutations, probably reflecting the ongoing adaptation to the new host. the relative genetic homogeneity of sars-cov isolates from later in the outbreak - may reflect a virus better adapted to the new host. the fact that much of the global spread arose from one index case in hotel m in hong kong , may also contribute to this genetic homogeneity. a ban on the sale of wildlife in wet markets in guangdong imposed during the later period of the sars outbreak was lifted in september . between december and january , there were four new cases of sars, the first nonlaboratory-associated cases diagnosed in humans since the end of the sars outbreak in july . epidemiological linkage and phylogenetic data suggest that the associated viruses were new introductions from animals (y. guan, unpublished observations) , , . these human cases were relatively mild and did not lead to secondary transmission, reflecting that the animal precursor virus is probably not well adapted to efficient human-tohuman transmission. this is probably a recapitulation of events in late in the run-up to the sars outbreak in . this time, the findings led to the reintroduction of the ban on wild-game animal markets and there have been no further naturally acquired human cases since. it is likely that the precursor of sars-cov has repeatedly crossed the species barrier but only occasionally has it succeeded in adapting to human-human transmission. this adaptation clearly occurred in late and it may happen again in the future. but given the present understanding and awareness about sars, we expect that such reemergence is unlikely to lead to a global outbreak on the scale of . the major routes of transmission of sars are droplet infection, aerosolization and fomites (refs. , and world health organization, http://www.who.int/csr/sars/en/whoconsensus.pdf ) . deposition of infected droplets or aerosols on the respiratory mucosal epithelium probably initiates viral infection. whether infection can occur through the oral or conjunctival routes is unknown, but sars-cov has been detected in tears . although exposure to the animal precursor of figure the global spread of sars. the number of probable cases of sars and the date of onset of the first case in each country (or group of countries) is denoted. the countries denoted in red are those where substantial local transmission occurred. the data are based on world health organization, http://www.who.int/csr/sars/country/table _ _ /en_ /en/print.html and the figure is adapted from ref. . wet markets in guangdong: 'wet markets' selling live poultry, fish, reptiles and other mammals are commonplace across southeast asia and southern china to service the cultural demand for freshly killed meat and fish produce. in some regions (e.g., guangdong province, china), increasing affluence has led to the proliferation of markets housing a range of live 'wild' animal species, such as civet cats, pictured, linked to the restaurant trade servicing the demand for these exotic foods. , , once the virus had adapted to human-to-human transmission in the later part of the outbreak, asymptomatic infection seemed to be rare . other peculiarities about sars-cov transmission were also evident. transmission was infrequent during the first five days of illness and, unlike transmission of influenza, was relatively inefficient in the household setting . despite sars's fearsome reputation and global spread, the average number of secondary infectious cases generated by one case (r ) was low ( . - . ); in contrast, the r of influenza ranges from to (ref. ) . although not unique to sars, 'superspreading events' (in which a few affected individuals disproportionately contribute to transmission) were characteristic of the outbreak , . the factors underlying the superspreading phenomenon of sars are poorly understood but may include coinfection with other viruses and host factors, as well as behavioral and environmental factors. the clinical symptoms of sars-cov infection are those of lower respiratory tract disease [ ] [ ] [ ] [ ] . besides fever, malaise and lymphopenia, affected individuals have slightly decreased platelet counts, prolonged coagulation profiles and mildly elevated serum hepatic enzymes. chest radiography reveals infiltrates with subpleural consolidation or 'ground glass' changes compatible with viral pneumonitis. but although the main clinical symptoms are those of severe respiratory illness, sars-cov actually causes infection of other organs: some affected individuals have watery diarrhea, and virus can be cultured from the feces and urine, as well as the respiratory tract [ ] [ ] [ ] . in addition, rt-pcr has identified the virus in the serum, plasma and peripheral blood leucocytes , . individuals with sars also have a pronounced peripheral t-cell lymphocytopenia: numbers of cd + and cd + cells are both reduced, and more than one-third of individuals have a cd + t-cell count of less than cells/mm (refs. , ) , suggesting increased susceptibility to secondary infections. the mechanisms underlying the t-cell lymphopenia remain to be elucidated. around - % of individuals with sars require management in intensive care units and the overall fatality rate is ∼ % (world health organization, http://www.who.int/csr/sars/en/whoconsensus.pdf). the age dependence of disease severity and mortality is notable; during the outbreak, mortality rates of affected individuals in hong kong who were - , - , - and > -year old were %, %, % and %, respectively (world health organization, http://www.who.int/ csr/sars/en/whoconsensus.pdf). none of the - -year-olds infected with sars-cov in hong kong had disease severe enough to require intensive care or mechanical ventilation , . this progressive age dependence in mortality is not totally explained by comorbid factors and the underlying biological basis remains unclear. quantitative studies of viral load have provided insights into the pathogenesis of sars. viral load is higher in the lower respiratory tract than in the upper airways , . viral load in the upper respiratory tract and feces is low during the first days and peaks at around day of illness. in marked contrast, viral load in influenza peaks soon after onset of clinical symptoms . this unusual feature of sars-cov infection explains its low transmissibility early in the illness. it also explains the poor diagnostic sensitivity of the first-generation rt-pcr diagnostic tests on upper respiratory tract and fecal specimens collected early in the illness (reviewed in ref. ) . affected individuals with high serum viral loads have a poor prognosis . between days - of illness, high viral load in nasopharyngeal aspirates, feces and serum, as well as detection of virus in multiple anatomic sites, are independently predictive of adverse clinical outcome . serial studies of viral load throughout illness also reflect clinical outcome . taken together, these findings suggest that poor clinical outcome is associated with continued uncontrolled viral replication. sars-cov rna can be invariably detected in the lungs of individuals dying of sars, but viral load is higher in those dying earlier in the course of the illness (< days) . the respiratory tracts of affected individuals who die during the first ten days of illness show diffuse alveolar damage with a mixed alveolar infiltrate, lung edema and hyaline membrane formation. macrophages are a prominent component of the cellular exudates in the alveoli and lung interstitium , . multinucleate syncytia of macrophage or epithelial cell origin are sometimes seen later in the disease. immunohistochemistry, in situ hybridization and electron microscopy on autopsy or tissue biopsy have unequivocally demonstrated sars-cov replication in pneumocytes in the lung and enterocytes in the intestine [ ] [ ] [ ] [ ] . individual reports of virus detection by in situ hybridization or immunohistochemistry in other tissues await confirmation by electron microscopy . in the large and small intestines, the virus replicates in enterocytes . viral particles primarily are seen on the apical surface of enterocytes and rarely in the glandular epithelial cells. but there is no villous atrophy or cellular infiltrate in the intestinal epithelium and the pathogenic mechanisms responsible for watery diarrhea in individuals with sars is unclear. some human intestinal epithelial cell lines support productive replication of sars-cov and gene expression arrays have shown that virus replication is associated with the expression of an antiapoptotic host cellular response, perhaps explaining the lack of enterocyte destruction in vivo . studies using pseudotyped lentiviruses, carrying the spike, membrane and envelope surface glycoproteins of sars-cov (fig. ) separately and in combination demonstrated that the spike protein is both neces- sary and sufficient for virus attachment on susceptible cells [ ] [ ] [ ] [ ] . the sars-cov spike protein uses a mechanism similar to that of class fusion proteins in mediating membrane fusion , . there is no consensus as to whether the virus entry occurs through a ph-dependent receptor-mediated endocytosis or through direct membrane fusion at the cell surface , , . the receptor for sars-cov was identified as the metallopeptidase ace- (refs. , ) . the soluble ace- ectodomain blocks sars-cov infection , and amino acids - of the spike protein are required for interaction with ace- (ref. ) . other coronaviruses use different cell receptors and enter cells either by means of fusion at the plasma membrane or through receptor-mediated endocytosis . immunostaining techniques have identified ace- on the surface of type and pneumocytes, the enterocytes of all parts of the small intestine and the proximal tubular cells of the kidney. this localization explains the documented tissue tropism of sars-cov for the lung and gastrointestinal tract and its isolation from the urine. but it is notable that colonic enterocytes lack ace- protein expression although sars-cov replication does occur in colonic epithelium , . in contrast, whereas ace- is strongly expressed on the endothelial cells of small and large arteries and veins of all tissues studied and the smooth muscle cells of the intestinal tract, there is no evidence of virus infection at any of these sites. this lack of virus infection in tissues that express the putative receptor prompts the question of whether a coreceptor is required for successful virus infection . vasculitis is known to occur in individuals with sars but its relation to infection of endothelial cells is unknown. because only the basal layer of the nonkeratinized squamous epithelium of the upper respiratory tract expresses ace- (ref. ) , undamaged epithelium of the nasopharynx is unlikely to support sars-cov replication. other receptors for virus entry that are independent of ace- expression may exist. pseudotyped virus containing the spike protein has also been shown to bind to dendritic cell-specific intercellular adhesion molecule grabbing nonintegrin (dc-sign) . dc-sign is a type-ii transmembrane adhesion molecule found on dendritic cells consisting of a c-type lectin domain that recognizes carbohydrate residues on a variety of pathogens. unlike the ace- receptor on pneumocytes and enterocytes, dc-sign does not permit sars-cov infection of the dendritic cells. instead, binding of sars-cov to dc-sign allows dendritic cells to transfer infectious sars-cov to susceptible target cells . a similar mechanism has been described for dengue virus, human immune deficiency virus (hiv) and cytomegalovirus, and may be relevant in sars pathogenesis. many details of sars-cov pathogenesis remain to be elucidated, but the development of a full-length infectious cdna clone of sars-cov should permit precise manipulation of the virus genome and will help our understanding of the viral determinants of pathogenesis . several inflammatory cytokines (il- β, il- and il- ) and chemokines chemotactic for monocytes (mcp- ) and neutrophils (ip- ) are elevated in adults and children with sars [ ] [ ] [ ] [ ] . the increased levels of monocyte-tropic chemokines may contribute to the prominently monocytic macrophagic infiltrate observed in the lung . but increases of these same chemokines occur in other viral diseases (e.g., influenza) and are not a unique feature of sars. in addition, elispot assays of peripheral blood leukocytes have revealed prolonged immunological dysregulation in individuals with sars . it is difficult to evaluate the overall pathogenic significance of these findings because immunological markers in the peripheral blood do not always reflect the local microenvironment of the lung . genetic factors associated with susceptibility to, or severity of, sars are under investigation. hla-b* has been associated with severe sars disease in taiwan but not hong kong . hla-b* has also been associated with disease susceptibility and hla-drb * with resistance to sars. the coinheritance of b* and b was significantly higher in individuals with sars than in the general population . the mechanisms underlying these disease associations remain to be elucidated. key to the development of effective antiviral drugs and vaccines against sars-cov was the development of animal models of sars ( table ) . sars-cov seems to cause infection in cynomolgous macaques following intratracheal inoculation [ ] [ ] [ ] . but whereas some researchers find evidence of disease pathology reminiscent of that seen in individuals dying of sars and can show sars-cov antigen and viral particles in the pneumocytes of infected macaques , , others only find evidence of a mild upper-airway disease and low levels of virus by rt-pcr . these differences in outcome may reflect differences in the viral strain, pre-exposure history and age of the animals, route of inoculation, stage of infection at which necropsy was performed or other factors. other animal models include ferrets, cats, golden syrian hamsters, mice and african green monkeys ( table ) [ ] [ ] [ ] [ ] [ ] . these animal models support viral replication in the upper and lower respiratory tracts [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . ferrets and hamsters also develop notable lung pathology. infected cats and ferrets transmit sars-cov to noninfected animals held in the same cage . natural asymptomatic infection in cats was documented during the community outbreak at amoy gardens, hong kong (world health organization, http://www.who.int/csr/sars/en/ whoconsensus.pdf). these animal models of sars differ from natural human disease in that the period between infection and peak disease pathology or peak viral load is shorter than is found in human disease and because the disease pathology, when present, is self-limited and rarely progresses to a fatal outcome as occurs with sars. they also do not accurately reproduce the intestinal component of the human disease. but these models provide the only options presently available for addressing questions relevant to therapeutics and vaccine development. they can provide useful information providing their limitations are recognized. several potential antiviral agents have been evaluated in vitro, and a few have been tested in animal models. screening of currently available antiviral drugs and chemical libraries reveals that interferons, glycyrrhizin, baicalin, reserpine, niclosamide, luteolin, tetra-o-galloylβ-d-glucose and the protease inhibitors have in vitro activity against sars-cov - . differences in in vitro susceptibility of sars-cov to interferon (ifn)-β b, ifn-α and ribavirin , - probably relate to differences in the testing methods used. overall, ifn-αn /n , leukocytic ifn-α, ifn-β and the hiv protease inhibitors (especially nelfinavir) are consistently active in vitro and should be considered for animal studies and randomized placebocontrolled clinical trials. type interferons render uninfected cells refractory to sars-cov replication through a mxa-independent mechanism , whereas the hiv protease inhibitors may block the activity of the main sars-cov proteinase . so far, only interferons have been tested in animal models: in cynomolgous macaques, pegylated ifn-αn provided prophylaxis but was only marginally effective for early treatment . no randomized placebo-controlled trials have been performed for any of these antiviral drugs, although treatment studies using historical controls have suggested clinical benefit from ifn-α (infacon- ) and the combination of a protease inhibitor with ribavirin . the rapidity with which the sars-cov genome was sequenced, the determination of the structure of potential drug targets and the prediction of functional properties of sars-cov proteins based on prior knowledge of homologs from other coronaviruses have allowed identification of potential new drug targets. peptides derived from the heptad-repeat- region of the spike protein have been shown to block virus infection, albeit at much higher molar concentrations than similar inhibitors needed to prevent hiv entry , . short interfering rnas also seems to be effective in decreasing viral replication in cell lines [ ] [ ] [ ] , but this remains an experimental strategy rather than one immediately amenable to clinical application. screening of combinatorial chemical libraries has identified inhibitors of sars protease, helicase and spike-protein-mediated cell entry . for successful treatment of influenza, antiviral drugs must be administered within hours of disease onset to obtain substantial clinical effect. but because the sars-cov load increases until day of illness , and in light of the correlation of high viral load in the second week of illness with adverse outcome , the window of opportunity for antiviral therapy may be wider. much scientific effort has been focused on developing a vaccine to protect against future outbreaks of sars-cov. the commercial viability of developing a vaccine for sars-cov will ultimately depend on whether the virus re-emerges in the near future. as discussed above, it is unlikely that future outbreaks will reach global proportions, but nevertheless, vaccines or passive immunization would be relevant in the context of protecting high-risk individuals such as laboratory and health-care workers. a vaccine could also be considered in the setting of the farmed-game-animal trade, if farming of civets for human consumption continues. in the short time since the virus was identified, substantial progress has been made toward developing a vaccine. immunodominant b-and t-cell epitopes of sars-cov are being defined [ ] [ ] [ ] . natural human infection with sars-cov leads to a long-lived neutralizing antibody response and immune sera crossneutralize diverse human sars-cov , suggesting that active immu- (in the press) ( ) nization against sars may be a feasible proposition. but so far there has been no known instance of human re-exposure to sars-cov to confirm that the naturally acquired immune response confers protection from reinfection. when sars-cov spike, envelope, membrane and nucleocapsid proteins were individually expressed in an attenuated parainfluenza type vector, only the recombinants expressing the spike protein induced neutralizing antibody and protected from challenge in hamsters ( table ) . mucosal immunization of african green monkeys with this parainfluenza-spike protein chimeric virus led to neutralizing antibody and protection from viral replication in the upper and lower respiratory tracts after challenge with live sars-cov , and spike protein-encoding dna vaccines stimulated neutralizing antibody production and protection from live-virus challenge in mice . these studies confirm the assumption that the spike protein is the dominant protective antigen for sars. experiments using adoptive transfer and t-cell depletion showed that humoral immunity alone can confer protection . other vaccine strategies have included the use of naked dna - , adenoviral vectors or modified vaccinia (ankara) and inactivated whole virus , . many investigators have optimized the codon usage of the gene target to improve expression. in summary, all vaccines based on the spike protein seem to induce neutralizing antibody responses, and those carrying nucleoprotein can induce nucleoprotein-specific cellmediated immunity. but thus far only four studies have used live sars-cov to challenge immunized animals ( table ). an inactivated vaccine with alum adjuvant, which induces neutralizing antibody in mice, is entering phase human clinical trials in china . experience with coronavirus vaccines for animals is relevant for sars vaccine development . one problem facing animal coronavirus vaccines has been strain variation among field isolates, leading to variable vaccine efficacy. a further concern is the experience with feline infectious peritonitis coronavirus, in which prior immunization led to enhanced disease rather than protection . in the case of sars-cov, neither vaccination nor passive transfer of antibody has yet been reported to lead to disease enhancement, but challenge with live sars-cov has occurred soon after immunization. whether waning immunity or low titers of antibody lead to sars disease enhancement remains unclear; the recent suggestion that immunized ferrets became more ill after challenge clearly needs to be confirmed or refuted . passive transfer of immune serum protects naive mice from sars-cov infection , and hyperimmune globulin with sufficient neutralizing activity for use in humans could be prepared from pooled convalescent human plasma or from horses immunized with inactivated vaccine. alternatively, monoclonal antibodies with sufficient neutralizing antibody activity have been developed by screening phage-display antibody libraries and by immortalizing b-cell repertoires of convalescent sars individuals with epstein-barr virus ( table ) [ ] [ ] [ ] . one of these ( r) blocks the virus-ace- receptor interaction through binding to the spike protein s domain . passive immunization of ferrets and mice was effective in suppressing viral replication in lungs, but less so in the nasopharynx , . no randomized placebo control trial evaluated antibody therapy for pre-or post-exposure prophylaxis in at-risk groups during the sars outbreak. retrospective analysis of outcome in a limited human study using human sars convalescent plasma suggested that passive immunization had no obvious adverse effects . the antigenic diversity of sars-cov-like precursor viruses in the wild-animal reservoir is undefined. in the event of a new interspecies transmission event prompting another sars outbreak, the crossprotection afforded by current vaccine constructs based on the human sars-cov of is unknown and is likely to influence the efficacy of both passive and active immunization strategies. sars provided a painful reminder of the global impact of emerging infectious diseases. it illustrated how microbes, with their evolutionary drive to preserve and propagate their genes, exploit new opportunities and niches created by modern society . interspecies transmission of viruses to humans clearly has occurred throughout human history. but recent developments allowed sars-cov increased opportunity to adapt to human-to-human transmission and, subsequently, to spread globally. in particular, large centralized wet markets and hospitals proved to be venues for amplification of transmission to humans, and the burgeoning increase of international travel (currently ∼ million travelers annually) exploded the local outbreak of an emerging infection into a potential pandemic. because most recent emerging infectious disease threats have a zoonotic origin, we need to better understand the microbial ecology of livestock and wildlife. in the context of increased attention and research funding directed at preparedness to combat bioterrorism threats, it is relevant to note that nature remains the greatest 'bioterrorist.' although microbes that cause commercially important diseases in livestock are well studied, organisms that pose threats to human health are not necessarily ones known to cause disease in livestock, or for that matter, in wildlife. nipah virus, hendra virus and sars-cov all have a wildlife reservoir. furthermore, at present there is concern over the possible role played by wild birds and ducks in the maintenance and spread of avian influenza a (h n ) in asia . greater understanding of the viral ecology of apparently healthy domestic animals and wildlife is therefore important. for example, the attention on ecological studies arising from the nipah virus and sars outbreaks have already led to the identification of a number of new viruses, including tioman, menangle, australian bat lyssavirus and a novel group coronavirus , . some of these are now known to be associated with human or livestock disease. but prioritizing such research efforts and assessing the public health relevance-if any-of such findings, poses challenges. three incidents of laboratory-acquired sars have arisen from biohazard level and laboratories, with community transmission arising from one (world health organization, http://www.wpro.who.int/ sars/docs/update/update_ .asp). these incidents were associated with lapses in biohazard level and practices. sars-cov can be safely handled in biohazard level laboratories provided that biohazard level practices are rigorously complied with. but as hospital health-care workers learned to their cost, sars-cov is an unforgiving virus; one lapse may be one too many, and it is irrelevant whether the lapse occurs in a biohazard level or laboratory. despite the impressive speed of scientific understanding of the disease, the global success in containing sars owed much to traditional public health methods of clinical case identification, contact investigation, infection control at healthcare facilities, patient isolation and community containment (that is, quarantine) . but the application of such measures in modern society during the control of sars highlighted several ethical and medical dilemmas, many of which arose from the need to balance individual freedoms against the common good , . 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pathogenic and potentially pandemic h n influenza virus in eastern asia managing emerging diseases borne by fruit bats (flying foxes), with particular reference to henipaviruses and australian bat lyssavirus identification of a novel coronavirus in bats ethics and sars: lessons from toronto informed consent and public health world health organization. a multicentre collaboration to investigate the cause of severe acute respiratory syndrome germs, governance, and global public health in the wake of sars the authors declare that they have no competing financial interests. key: cord- -vwnpred authors: bermejo-martin, jesus f; ortiz de lejarazu, raul; pumarola, tomas; rello, jordi; almansa, raquel; ramírez, paula; martin-loeches, ignacio; varillas, david; gallegos, maria c; serón, carlos; micheloud, dariela; gomez, jose manuel; tenorio-abreu, alberto; ramos, maría j; molina, m lourdes; huidobro, samantha; sanchez, elia; gordón, mónica; fernández, victoria; del castillo, alberto; marcos, ma Ángeles; villanueva, beatriz; lópez, carlos javier; rodríguez-domínguez, mario; galan, juan-carlos; cantón, rafael; lietor, aurora; rojo, silvia; eiros, jose m; hinojosa, carmen; gonzalez, isabel; torner, nuria; banner, david; leon, alberto; cuesta, pablo; rowe, thomas; kelvin, david j title: th and th hypercytokinemia as early host response signature in severe pandemic influenza date: - - journal: crit care doi: . /cc sha: doc_id: cord_uid: vwnpred introduction: human host immune response following infection with the new variant of a/h n pandemic influenza virus (nvh n ) is poorly understood. we utilize here systemic cytokine and antibody levels in evaluating differences in early immune response in both mild and severe patients infected with nvh n . methods: we profiled cytokines and chemokines and evaluated the haemagglutination inhibition activity as quantitative and qualitative measurements of host immune responses in serum obtained during the first five days after symptoms onset, in two cohorts of nvh n infected patients. severe patients required hospitalization (n = ), due to respiratory insufficiency ( of them were admitted to the intensive care unit), while mild patients had exclusively flu-like symptoms (n = ). a group of healthy donors was included as control (n = ). differences in levels of mediators between groups were assessed by using the non parametric u-mann whitney test. association between variables was determined by calculating the spearman correlation coefficient. viral load was performed in serum by using real-time pcr targeting the neuraminidase gene. results: increased levels of innate-immunity mediators (ip- , mcp- , mip- β), and the absence of anti-nvh n antibodies, characterized the early response to nvh n infection in both hospitalized and mild patients. high systemic levels of type-ii interferon (ifn-γ) and also of a group of mediators involved in the development of t-helper (il- , il- , il- , il- ) and t-helper (tnf-α, il- , il- p ) responses were exclusively found in hospitalized patients. il- , il- p , il- constituted a hallmark of critical illness in our study. a significant inverse association was found between il- , il- and pao in critical patients. conclusions: while infection with the nvh n induces a typical innate response in both mild and severe patients, severe disease with respiratory involvement is characterized by early secretion of th and th cytokines usually associated with cell mediated immunity but also commonly linked to the pathogenesis of autoimmune/inflammatory diseases. the exact role of th and th mediators in the evolution of nvh n mild and severe disease merits further investigation as to the detrimental or beneficial role these cytokines play in severe illness. methods we profiled cytokines and chemokines and evaluated the haemagglutination inhibition activity as quantitative and qualitative measurements of host immune responses in serum obtained during the first five days after symptoms onset, in two cohorts of nvh n infected patients. severe patients required hospitalization (n = ), due to respiratory insufficiency ( of them were admitted to the intensive care unit), while mild patients had exclusively flu-like symptoms (n = ). a group of healthy donors was included as control (n = ). differences in levels of mediators between groups were assessed by using the non parametric u-mann whitney test. association between variables was determined by calculating the spearman correlation coefficient. viral load was performed in serum by using real-time pcr targeting the neuraminidase gene. results increased levels of innate-immunity mediators (ip- , mcp- , mip- β), and the absence of anti-nvh n antibodies, characterized the early response to nvh n infection in both hospitalized and mild patients. high systemic levels of type-ii interferon (ifn-γ) and also of a group of mediators involved in the development of t-helper (il- , il- , il- , il- ) and t-helper (tnf-α, il- , il- p ) responses were exclusively found in hospitalized patients. il- , il- p , il- constituted a hallmark of critical illness in our study. a significant inverse association was found between il- , il- and pao in critical patients. conclusions while infection with the nvh n induces a typical innate response in both mild and severe patients, severe disease with respiratory involvement is characterized by early secretion of th and th cytokines usually associated with cell mediated immunity but also commonly linked to the pathogenesis of autoimmune/inflammatory diseases. the exact role of th and th mediators in the evolution of nvh n mild and severe disease merits further investigation as to the detrimental or beneficial role these cytokines play in severe illness. the emergence of the new pandemic variant of influenza virus (nvh n ) has brought renewed attention to the strategies for prevention, treatment and minimization of the social and human costs of the influenza disease [ ] [ ] [ ] [ ] [ ] . the great majority of nvh n infections are mild and self-limiting in nature [ ] [ ] [ ] . nevertheless, a small percentage of the patients require hospitalization and specialized attention in intensive care units (icus) [ ] [ ] [ ] [ ] . many severe cases occur in healthy young adults, an age group rarely seriously affected by seasonal influenza [ ] [ ] [ ] [ ] [ ] [ ] . while pregnancy and metabolic conditions (including obesity and diabetes) have been identified as risk factors for severe nvh n disease, to % of fatal cases have no documented underlying medical condition [ , , ] . the new virus causes more severe pathological lesions in the lungs of infected mice, ferrets and non-human primates than seasonal human h n virus [ ] . the role of host immune responses in clearance of nvh n or the role, if any, of host immune responses in contributing to severe respiratory pathogenesis of nvh n infections is not known at this time. we have previously identified specific host immune response chemokine and cytokine signatures in severe and mild sars cov, h n and respiratory syncytial virus infections. in these studies, early host immune responses are characterized by the expression of systemic levels of chemokines, such as cxcl , indicative of innate anti viral responses [ ] [ ] [ ] [ ] . severe and mild sars and rsv illness could further be defined by chemokine and cytokine signatures involved in the development of adaptive immunity. interestingly, de jong et al. have demonstrated that hypercytokinemia of specific chemokines and cytokines is associated with severe and often fatal cases of human h n infections [ ] . to determine if host immune responses play a potential role in the evolution of mild or severe nvh n illness we performed an analysis of systemic chemokine and cytokine levels in serum from severe and mild nvh n patients shortly following the onset of symptoms. interestingly, we identified cytokine signatures unique to mild and severe patients. both hospitalized and outpatients were recruited during the first pandemic wave in the months of july and august in different hospitals within the national public health system of spain. inclusion criteria: critical patients with respiratory insufficiency, hospitalized non critical patients with respiratory insufficiency, and mild outpatients with no respiratory insufficiency attending to the participant centers with confirmed nvh n infection by molecular diagnostic methods (see below) were asked to donate a serum sample for the study in the first contact with the participant physicians. initially we enrolled hospitalized patients and outpatients. to determine systemic levels of chemokines and cytokines in sera from nvh n infected individuals, we analyzed sera from hospitalized, outpatients, and control subjects for levels of different mediators. the final number of patients used for analysis was based on exclusion and matching criteria listed in figure . exclusion criteria: patients with signs of bacterial infection defined by the presence of purulent respiratory secretions, and/or positive results in respiratory cultures, blood cultures, and/or positive urinary antigen test to legionella pneumophila or streptococcus pneumoniae were excluded from the analysis ( figure ). children under years old and one patient older than years old were also excluded in order to make groups comparable by age. pregnant women were also excluded to avoid confusion factors during the analysis of the immune response to the virus, since pregnancy induces physiological changes in the immune system ( figure ). informed consent was obtained directly from each patient or their legal representative and also from the healthy controls before enrollment. approval of the study protocol in both the scientific and the ethical aspects was obtained from the scientific committee for clinical research of the coordinating center (hospital clinico universitario de valladolid, spain). sample collection and transport blood samples were collected by experienced nurses. a single serum sample was obtained from each patient or control. serum samples were obtained after proper centrifugation and were sent refrigerated to the national influenza center of valladolid (spain), where they were stored at - °c until immune mediator profiling, haemagglutination inhibition activity (hi) and viral load evaluation. nasopharyngeal swabs preserved in virus transportation medium were sent to the world health organization (who) associated national influenza centers of valladolid, majadahonda and barcelona, spain for viral diagnosis purposes. viral rna from nasopharyngeal swabs was obtained by using automatic extractors (biomerieux ® (marcy l'etoile, france), roche ® (basel, switzerland) and viral presence was assessed by real time pcr based methods using reagents provided free of charge by the centers for disease control (cdc, atlanta, usa) or purchased from roche ® (basel, switzerland) (h n detection set) on -well plate termocyclers (roche ® lc (basel, switzerland) and applied biosystems ® (foster city, ca, usa) viral load measurement viral load was measured and compared between groups by real time reverse transcription pcr on rna extracted from serum. briefly, an external curve was obtained by using a serial dilution of human rna extracted from cultured monocytic leukemia (thp- ) cells, and human gene gapdh was employed as reporter gene. nvh n neuraminidase gene was amplified by qrt-pcr in each serum sample, and crossing points were extrapolated to the external curve. analysis of samples and standard curve was conducted by using the fast v . . software (applied biosystem™). results were given as relative comparisons in (pg rna/μl). '- ' sequences of primer pairs: (reverse); nvh n neuraminidase: '-tcagtcgaaatgaatgccctaa- ' (forward) and n r '-cacggtcgattcgagccatg- '(reverse). serum chemokine and cytokine levels were evaluated using the multiplex biorad © plex assay (hercules, ca, usa). this system allows for quantitative measurement of different chemokines, cytokines, growth-factors and immune mediators while consuming a small amount of biological material. furthermore, this system has good representation of analytes for inflammatory cytokines, anti-inflammatory cytokines, th cytokines, th cytokines, th cytokines and chemokines, allowing for the testing of differential levels of regulatory cytokines in the serum of severe and mild patients. additionally, interferon α, adiponectin and leptin were measured by using an enzyme-linked inmuno adsorbant assay (elisa) from r&d © systems (minneapolis, mn, usa). hi assays were performed on a μl aliquot of the samples at university health network (uhn), toronto, ontario, canada. the sera was treated with receptor-destroying enzyme (rde) of v. cholerae by diluting one part serum with three parts enzyme and were incubated overnight in a °c water bath. the enzyme was inactivated by a -minute incubation at °c followed by the addition of six parts . % physiological saline for a final dilution of / . hi assays were performed in v-bottom -well microtiter plates (corning costar co., cambridge, ma, usa) with . % turkey erythrocytes, as previously described [ ] , using inactivated pandemic influenza a/california/ / (nvh n ) antigens. data analysis was performed using spss . . comparisons between groups were performed using the non parametric u-mann whitney test. data are displayed as (mean, standard deviation) for clinical and laboratory parameters and as (median, interquartile rank) for data on sample collection timing and the immune mediators levels. association between variables was determined by calculating the spearman correlation coefficient (r) and data shown as (r, p value). significance was fixed at p value < . all the patients showed symptoms of acute respiratory viral infection at disease onset. the most frequent initial symptoms were (% of patients in each group: critical, hospitalized non critical, outpatients): fever ( , , ), cough ( , , ), headache ( , , ), tiredness ( , , ) and myalgia ( , , ) . hospitalized patients showed dyspnoea as the initial symptom in % of the cases and % developed respiratory insufficiency at the time of hospital admission (dyspnoea and/or hypoxemia defined as o saturation < % breathing at least two liters of oxygen). ten patients required admission to an intensive care unit (icu) due to their respiratory situation. the remaining were admitted to other different specialized hospital services. outpatients had no difficulties with respiratory function, showing respiratory rates under ×'. sex composition was the same for both critical and non critical hospitalized patients: % of the patients were male (n = ) and % female (n = ). fifty-three percent of the outpatients were male and % female (n = and respectively) ( table ). average age was as follows: hospitalized patients ( . ; . ), outpatients ( . ; . ) and healthy controls, ( . ; . ). critical patients were slightly older than the other hospitalized patients (table ) . seven patients with critical illness and four severe patients with non critical illness showed previous pathologies (table ). ten out of of the critical patients, and / of the severe non critical patients showed a pathological chest x-ray within hours of onset of the symptoms (table ) . outpatients had received just antipyretics (paracetamol) before sample collection (none of them had received oseltamivir). one hundred percent of the hospitalized patients (critical and non critical), had received oseltamivir at the time of sample collection (table ) . lymphopenia was a common finding in the critical patients (mean; sd) ( . ; . ). ldh levels were increased over normal levels in hospitalized patients, mostly in those critically ill (table ) . furthermore, critical patients also showed high levels of cpk, got, gpt and glucose in venous blood (table ) . critical patients stayed longer at the hospital than the other hospitalized patients (table ) . three critical patients ultimately died (five days after onset due to hypoxemia and septic shock; days after onset by refractory hypoxemia complicated by systemic candidiasis; and the third after days of supportive therapy by multiorganic failure). hi activity (a/california/ / ) was present in serum from only two critically ill patients of and years old (titres / and / respectively) and in one -year-old outpatient (titre / ). serum from those three patients showing hi showed also the ability to block viral replication, as assessed by microneutralization assay against a/california/ / (data not shown). this data supports the notion that at the time of sampling the vast majority of the patients had yet to produce table clinical and laboratory characteristics of the patients hospitalized, non critical illness (n = ) antibodies against nvh n and was in the early stages of disease. the virus induced in both mild and severe patients a systemic elevation of three chemokines that have been shown to be expressed early during viral infections, cxcl- (ip- ), ccl- (mcp- ) and ccl- (mip- β), with no differences in the levels of these mediators between them (data on immune mediators profiling are shown in figure and additional file ). il- , ifn-γ, il- , il- levels were higher in the hospitalized patients than in outpatients and controls (p < . ). il- behaved in a similar way. while both critical and non-critical hospitalized patients showed higher levels of il- and tnf-α than controls, only severe non critical patients showed significant higher levels of il- and tnf-α than mild. on the other hand, il- and il- p increased exclusively in critical patients, who in addition showed the highest levels of il- of the compared groups. to determine if systemic viral load plays a role in chemokine or cytokine expression levels we evaluated serum for nvh n levels. fifty-seven percent of critical patients, % of hospitalized non critical patients, and % of mild patients showed positive virus in serum. for those with positive virus in serum, we found no differences in viral load between critical patients, hospitalized non critically ill, and mild outpatients (figure ). we found significantly higher levels of il- and il- in those hospitalized patients with negative virus in serum compared to those with virus in serum (data not shown). similarly, inverse correlations were found between viral load and il- , il- in patients requiring hospital admission (figure ). when mediator levels were correlated with the clinical parameters, a significant inverse association was found between il- and pao in hospitalized patients (figure ). exclusively in the critical patients group, il- inversely correlated with pao [- . ; . ]. in the non critically ill hospitalized patients group, a negative association was observed between il- and pao [- . ; . ]. in a first attempt to understand the role host immune responses play in the evolution of severe and mild nvh n disease, we assessed systemic levels of chemokines and cytokines in the sera from hospitalized and outpatients. consistent with our previous studies on early elevated expression of cxcl , ccl and ccl in sars cov and rsv infected patients [ ] [ ] [ ] [ ] , we found in the present study elevated expression of these chemokines in severe patients (critical and non critical) and mild patients. the early expression of these chemokines in all patients likely is indicative of innate antiviral host responses. one of the most intriguing observations in our present study is the dramatic increase of mediators which stimulate th- responses (ifn-γ, tnf-α, il- , il- p ) and th- ones (il- , il- , il- , il- ) in the severe patients ( figure ). th- adaptive immunity is an important response against intracellular microbes such as viruses [ ] . th- immunity participates in clearing pathogens during host defense reactions but is involved also in tissue inflammation in several autoimmune diseases, allergic diseases, and asthma [ ] [ ] [ ] [ ] [ ] . increase in ifn-γ il- , il- , il- and il- in both critical and non critical hospitalized patients compared to mild ones indicates that they constitute hallmarks of severe disease. ifn-γ and il- promote antiviral immunity but also respiratory tract inflammation by recruiting neutrophils and mononuclear cells to the site of the infection [ ] [ ] [ ] . il- is a th cytokine that induces differentiation of th- cells [ ] . il- and il- show immunomodulatory properties. il- attenuates th- cytokine production [ ] . il- is known to be an anti-inflammatory cytokine. in a murine model, mckinstry et al.revealed that il- inhibits development of th- responses during influenza infection, correlating with compromised protection [ ] . increase of il- and tnf-α in hospitalized patients over control indicated that they also parallel severe disease, but the significantly higher levels of il- and tnf-α in severe non critical patients compared to mild (difference not found for critical ones), could reflect a beneficial role of these cytokines in this particular subset of patients. the patient who died five days after disease onset showed high viral load and undetectable il- levels in serum. this could reflect a protective role of il- in severe patients. il- , il- p , il- constituted a hallmark of critical illness in our study. these three cytokines also mediate both antiviral and pro-inflammatory responses. il- is a potent regulator switching immune responses from the induction of foxp + regulatory t cells to pathogenic th cells in vivo [ ] . il- promotes cd t cells homeostatic proliferation [ ] in response to infection. il- plays a key role in the switch from innate to adaptive immunity [ ] . levels of immune mediators in the four groups levels of immune mediators in the four groups. *significant differences with control at the level p < . . high levels of th- and th- related mediators could support the hypothesis of a th- +th- inflammatory response in the origin of the severe respiratory disease caused by nvh n infection. alternatively, an increase in th- and th- cytokines may reflect a vigorous antiviral host response necessary for clearance of virus during severe lower respiratory infections. while the ability of influenza a virus to induce the production of chemotactic (rantes, mip- α, mcp- , mcp- , and ip- ) and pro-inflammatory (il- β, il- , il- , and tnf-α) th related mediators is well know from previous reports on seasonal influenza [ , ] , this is the first report evidencing th response as a signature of severe influenza disease in humans [ , ] . since there are immunomodulatory drugs which have shown to down-modulate the activity of both th and th [ ] , the results obtained here supports the development of further studies on animal models aimed to clarify the role of these mediators in the pathogenesis of the acute respiratory disease showed by severe nvh n infected patients. analysis of the immune mediators involved in host responses to the virus in mild and severe cases revealed th and th cytokine responses as early distinctive hallmarks of severe respiratory compromise following infection with nvh n . the exact role of th and th mediators in the evolution of nvh n mild and severe disease merits further investigation as to the detrimental or beneficial role these cytokines play in severe illness. the influence of th -dominant conditions (autoimmune diseases) or th deficient ones (hiv infection) on disease outcome should also be explored. furthermore, the impact of other regulatory cytokines elevated in severe disease (il- , il- ) on the evolution of host immune responses to nvh n infections may represent alternative therapeutics for controlling severe illness. correlation studies correlation studies. from left to right: correlation between il- level and viral load in serum; correlation between il- level and viral load in serum; correlation between il- serum levels and pao . • the great majority of infections caused by the new influenza pandemic virus are mild and self-limiting in nature. nevertheless, a small percentage of the patients develop severe respiratory disease. analysis of the immune mediators involved in host responses to the virus along with the evaluation of the humoral responses in mild and severe cases may help understand the pathogenic events leading to poor outcomes. • early response to the virus in both hospitalized and outpatients was characterized by expression of chemokines (cxcl , ccl and ccl ), also observed in the response to sars cov, h n and rsv, which previous literature describes to correspond to innate antiviral responses. • patients who develop respiratory compromise in the first days following infection with nvh n typically showed th and th hyper-cytokinemia, compared to mild patients and healthy controls. these cytokine profiles have been previously reported to participate in both antiviral and pro-inflammatory responses. • increased systemic levels of il- , il- p , il- constituted a hallmark of critical illness. these mediators are known to promote the development of adaptive responses and also pro-inflammatory ones in other viral infections. • our findings constitute a major avenue to guide the design of further works studying the beneficial or detrimental role of th and th responses in this disease. the following additional files are available online: predominant cytokine profiles paralleling early nvh n disease by clinical severity predominant cytokine profiles paralleling early nvh n disease by clinical severity. emergence of a novel swine-origin influenza a (h n ) virus in humans lurie n: h n influenza, public health preparedness, and health care reform swine flu. after delays, who agrees: the pandemic has begun a race against time to vaccinate against novel h n virus the transmissibility and control of pandemic influenza a (h n ) virus europe's initial experience with pandemic (h n ) -mitigation and delaying policies and practices national public health service for wales, hpa northern ireland swine influenza investigation teams: epidemiology of new influenza a (h n ) virus infection influenza a(h n )v in germany: the first , cases critical care services and h n influenza in australia and new zealand hospitalized patients with h n influenza in the united states critically ill patients with influenza a(h n ) infection in canada león-gil c: intensive care adult patients with severe respiratory failure caused by influenza a (h n )v in spain severe respiratory disease concurrent with the circulation of h n influenza epidemiology of fatal cases associated with pandemic h n influenza in vitro and in vivo characterization of new swine-origin h n influenza viruses gene expression analysis of host innate immune responses during lethal h n infection in ferrets human immunopathogenesis of severe acute respiratory syndrome (sars) dj: interferon-mediated immunopathological events are associated with atypical innate and adaptive immune responses in patients with severe acute respiratory syndrome arranz e: persistence of proinflammatory response after severe respiratory syncytial virus disease in children fatal outcome of human influenza a (h n ) is associated with high viral load and hypercytokinemia concepts and procedures for laboratory-based influenza surveillance. us department of health and human services the effector t helper cell triade interleukin- and systemic lupus erythematosus: current concepts il- and th cells development and function of th cells in health and disease il- induces differentiation of th cells and enhances function of foxp + natural regulatory t cells molecular mechanisms of cytokine and chemokine release from eosinophils activated by il- a, il- f, and il- : implication for th lymphocytesmediated allergic inflammation both conventional and interferon killer dendritic cells have antigenpresenting capacity during influenza virus infection molecular pathogenesis of influenza a virus infection and virus-induced regulation of cytokine gene expression il- mrna in sputum of asthmatic patients: linking t cell driven inflammation and granulocytic influx? a functional il- receptor is expressed on polarized murine cd + th cells and il- signaling attenuates th cytokine production il- deficiency unleashes an influenza-specific th response and enhances survival against high-dose challenge oukka m: il- controls th immunity in vivo by inhibiting the conversion of conventional t cells into foxp + regulatory t cells loss of il- r and il- r expression is associated with disappearance of memory t cells in respiratory tract following influenza infection protective immunity to influenza: lessons from the virus for successful vaccine design tc , a unique subset of cd t cells that can protect against lethal influenza challenge critical role of il- ra in immunopathology of influenza infection confronting an influenza pandemic with inexpensive generic agents: can it be done? this work has been made by an international team pertaining to the spanish-canadian consortium for the study of influenza immunopathogenesis. the authors would like to thank lucia rico and verónica iglesias for their assistance in the technical development of the multiplex cytokine assays, to begoña nogueira for her technical support, and to nikki kelvin for language revision of this article. this work was possible thanks to the financial support obtained from the ministry of science of spain and consejería de sanidad junta de castilla y león, programa de investigación comisionada en gripe, gr / , programa para favorecer la incorporación de grupos de investigación en las instituciones del sistema nacional de salud, emer / , and proyectos en investigación sanitaria, pi . cihr, nih and lksf-canada support djk. this sponsorship made possible reagent acquisition and sample transportation between participant groups. the authors declare that they have no competing interests. tp, jr and iml assisted in the design of the study, coordinated patient recruitment, analysed and interpreted the data, and assisted in writing the paper. pr, mcg, cs, dm, jmg, sh, es, mg, ac, bv, cjl, jad, ch, ig and pc supervised clinical aspects, participated in patient recruitment, assisted in the analysis, interpretation of data, and writing the report. at, mjr, mlm, vf, mam, mrd, jcg, rc, sr and jme performed viral diagnosis, assisted in the analysis, interpretation of data, and writing the report. ra performed cytokine profiling, and assisted in supervision of laboratory work and writing the report. nt collected clinical data, and assisted in writing the report. tr, db performed the hai assays and assisted in writing the report. dv and al designed and performed the quantitative pcr method for viral load measurement. jfbm, djk and rol were the primary investigators, designed the study, coordinated patient recruitment, supervised laboratory works, and wrote the article. key: cord- -jh qmoso authors: ortiz, justin r.; jacob, shevin t.; eoin west, t. title: clinical care for severe influenza and other severe illness in resource‐limited settings: the need for evidence and guidelines date: - - journal: influenza and other respiratory viruses doi: . /irv. sha: doc_id: cord_uid: jh qmoso the influenza a (h n ) pandemic highlighted the importance of quality hospital care of the severely ill, yet there is evidence that the impact of the pandemic was highest in low‐ and middle‐income countries with fewer resources. recent data indicate that death and suffering from seasonal influenza and severe illness in general are increased in resource‐limited settings. however, there are limited clinical data and guidelines for the management of influenza and other severe illness in these settings. life‐saving supportive care through syndromic case management is used successfully in high‐resource intensive care units and in global programs such as the integrated management of childhood illness (imci). while there are a variety of challenges to the management of the severely ill in resource‐limited settings, several new international initiatives have begun to develop syndromic management strategies for these environments, including the world health organization's integrated management of adult and adolescent illness program. these standardized clinical guidelines emphasize syndromic case management and do not require high‐resource intensive care units. these efforts must be enhanced by quality clinical research to provide missing evidence and to refine recommendations, which must be carefully integrated into existing healthcare systems. realizing a sustainable, global impact on death and suffering due to severe influenza and other severe illness necessitates an ongoing and concerted international effort to iteratively generate, implement, and evaluate best‐practice management guidelines for use in resource‐limited settings. the influenza a (h n ) pandemic highlighted the essential role of hospital care of the severely ill in the response to public health emergencies. early reports from mexico and canada of intensive care units (icus) filled to capacity with patients with severe respiratory infections helped to calibrate the early, aggressive global public health response. [ ] [ ] [ ] the critical care medical community and public health reacted with unprecedented coordination to describe severe disease, to disseminate data on the epidemiology and care of pandemic patients, and to convey the impact of the outbreak on health systems. [ ] [ ] [ ] [ ] [ ] while the worst-case scenario of a -scale pandemic was avoided, there were reports of patients with severe disease stressing critical care services in communities throughout the world. , [ ] [ ] [ ] these reports show that severely ill patients can divert resources and impact the balance of care delivery even in hospitals where the overall capacity is not exceeded. this problem is particularly acute in resource-limited settings where there is decreased capacity to manage severe illness. while building resource-intensive and highly technological icus is not feasible for many parts of the world, much can be done to improve care for severely ill patients in more austere environments. in this review, we discuss the inter-related issues of pandemic influenza, severe seasonal influenza, and severe illness more generally. given the burden of severe illness in resource-limited settings, it is vital to improve capacity to care for severely ill individuals in these environments. estimates of h n pandemic influenza morbidity and mortality differ considerably among countries. , [ ] [ ] [ ] in highresource countries, large epidemiologic studies have shown that the overall incidence of pandemic influenza requiring hospitalization was comparable to interpandemic seasons, [ ] [ ] [ ] [ ] but there were important demographic groups that experienced substantial increases in severe influenza disease. these groups included young adults, pregnant women, obese persons, and indigenous persons. in low-and middleincome countries, where laboratory testing for clinical care or for public health surveillance was limited, the full impact of the pandemic has been harder to measure. [ ] [ ] [ ] however, modeling studies estimate that there were disproportionately increased pandemic deaths in resource-limited regions. compared with the americas, the risk of pandemic respiratory and circulatory mortality was Á times higher in southeast asia and Á times higher in africa. these data underscore the importance of improving clinical management of influenza disease in resource-limited settings. a who consultation on the clinical care of pandemic influenza noted that there were limited clinical data and guidelines for the management of severe manifestations of viral infection in resource-limited settings. in light of the apparent paucity of data about supportive management of severe influenza-specific illness, the who public health research agenda for influenza commissioned the international respiratory and severe illness center (intersect) at the university of washington to perform a systematic review on non-antiviral, supportive management of persons with severe pandemic influenza a (h n ). a update of this review, limited to randomized controlled trials, controlled prospective cohort studies, and systematic reviews/meta-analyses found only seven pertinent studies, mostly of adults. one study found benefit of convalescent plasma infusion for severe illness, three studies found no benefit of corticosteroids for severe respiratory disease, [ ] [ ] [ ] and three studies had mixed results on the benefit of extracorporeal lung support for severe respiratory disease. [ ] [ ] [ ] no study identifying a therapeutic benefit from an intervention was applicable to healthcare delivery in resourcelimited settings. while pandemic influenza gained attention worldwide, the burden of seasonal influenza is perhaps less widely recognized. yet, the contribution of seasonal influenza to severe illness is substantial, especially in resource-limited settings. a variety of data support this assertion. among children younger than years of age in sub-saharan africa and south asia, influenzainfection and co-infections are commonly identified during hospitalizations for acute lower respiratory infections (alri). [ ] [ ] [ ] [ ] the only systematic review and metaanalysis that estimates the global burden of severe influenza disease was recently published. the study, which analyzes only early childhood disease, reports that % of influenza-related cases of severe alri among children younger than years occur in low-and middle-income countries, and that % of all alri in this age group are associated with influenza virus infection. the study dispels the myth that influenza is only a problem in temperate regions. the authors calculated that the incidence rate of severe influenza-associated alri in early childhood is similar in developing countries as compared with industrialized, temperate countries (both are around - episodes per child-years). this pediatric study underestimates the overall burden of severe influenza, as research has demonstrated that influenza mortality among adults is considerably higher than in children. in the united states, approximately children die annually from influenza virus infection, as compared with greater than influenza-attributable deaths among persons older than years. the burden of influenza among adults may be even higher in developing settings. recent data from south africa suggest that the risk of influenza mortality among the elderly in that country is greater than in the united states. further, countries with a high prevalence of hiv/aids and minimal availability of highly active antiretroviral therapy may also experience substantial risk of influenza mortality among non-elderly adults. thus, beyond pandemic planning and response, it is critical to optimize care of severe seasonal influenza in resource-limited settings. more generally, the global burden of severe illness is poorly understood. in the united states, acute respiratory distress syndrome (ards) and sepsis are two of the most common severe illnesses requiring critical care. , among adults worldwide, extrapolated data suggest that - million cases of sepsis and Á - Á million cases of ards occur annually. however, considering that about % of the global burden of respiratory mortality is in children < years of age, these estimates are a substantial underestimate of the total global burden of severe illness. the vast majority of severe illness occurs in low-and middle-income countries, yet there are currently little clinical data or evidence-based management guidelines to improve hospital care for patients in these settings. in intensive care medicine, severe illness treatment is syndromic in approach. clinical management of two common syndromes encountered in intensive care units -ards and sepsisfollow standardized guidelines. , these syndrome-focused guidelines facilitate the rapid recognition and treatment for life-threatening conditions, even before specific etiologies are identified. this approach also promotes the widespread adoption of research-proven interventions such as low tidal volume lung protective ventilation for ards. , similarly, several studies suggest that sepsis outcomes may be improved by the use of protocolized sepsis care pathways. , syndromic management is therefore an important tool in improving care of severely ill patients. in resource-limited settings, where advanced diagnostic equipment may not be available, syndromic management of severely ill patients using readily available tools offers a practical and feasible strategy for care. evidence of the benefit of syndromic management of hospitalized patients in resource-limited settings comes from the who integrated management for childhood illness (imci) program. imci guidelines were developed using existing clinical evidence and expert opinion to standardize healthcare provider training and care delivery in resourcelimited settings. the imci guidelines are designed to identify children in need of care by presenting signs and symptoms, and they do not require diagnostic tests that are likely unavailable in most austere settings. studies evaluating training and implementation of imci guidelines have shown a substantial impact on improved management and survival related to childhood pneumonia and other common illnesses. [ ] [ ] [ ] [ ] [ ] for example, a cluster randomized trial in kenya evaluating the efficacy of a multi-faceted quality improvement intervention for the management of severely ill children resulted in improved quality of clinical care when compared with a less comprehensive approach. similarly, in a pediatric outpatient and emergency unit in malawi, directed trainings based on imci to improve triage and emergency care resulted in streamlined healthcare delivery and a % decrease in pediatric in-hospital mortality. modifications of imci have also been shown to improve pneumonia outcomes when implemented in the community setting. [ ] [ ] [ ] moreover, economic analyses have found imci protocolized care to be cost effective and comparable to preventive interventions such as routine childhood pneumococcal conjugate immunization. it is important to highlight that clinical management guidelines may have only modest effects on important outcomes in resource-limited settings for several reasons, including an incomplete evidence base and numerous challenges to implementation. further, some studies have shown limited or no effects of knowledge translation activities, such as clinical guideline implementation. , nevertheless, even modest treatment effects on a high burden disease can have a massive impact when widely implemented. the imci experience demonstrates the potential benefits of syndromic management for persons with severe respiratory infections in resource-limited settings. global disparities in access to care for severe illness are substantial. , , who reported in that poor clinical outcomes of pandemic influenza were associated with delays in seeking health care, limited access to supportive care, and "rapidly progressive overwhelming lung disease which is very difficult to treat." disparities in access to critical care may partly explain some of the pandemic influenza mortality differences reported among countries. , for example, during the early phase of the h n pandemic, reported icu mortality in middleincome mexico was twice that of high-income canada. , , access to typical health technologies used to manage critically ill patients such as pulse oximetry, invasive hemodynamic monitoring, blood gas analyzers, and mechanical ventilation may be limited or absent in resource-limited settings. , hospital care is often delivered by nurses and non-specialist doctors who may have limited time, resources, training, and access to information to manage severely ill patients, , particularly during a public health emergency like the influenza pandemic. from the few surveys of icu resources in resource-limited settings, data suggest that many hospitals are ill equipped to dedicate sufficient personnel and supplies required by patients with severe illness. [ ] [ ] [ ] [ ] alarming shortages of reliable electricity, clean water, and supplemental oxygen have been reported from hospitals in sub-saharan africa and southeast asia. , developing country health systems may be weak and hospital support from government ministries is often lacking. thus, strategies to improve care delivery in low-and middle-income countries must address challenges of improving access to care, training and retention of healthcare providers, supply chain management, and strengthening healthcare systems. any new healthcare intervention designed to improve management of severe influenza disease must be integrated into the current health system structure and strengthen healthcare delivery overall if it is to be successfully adopted and remain sustainable. despite the absence of sophisticated equipment and abundant resources, including icus, it is likely that many lives in resource-limited settings can be saved by promoting the basic tenets of severe illness management. , , examples include simple triage systems to rapidly identify ill patients, protocolized supplemental oxygen therapy managed by nonphysician staff, infection source control, and early antimicrobial therapy for sepsis based on local antimicrobial clinical care for severe influenza ª blackwell publishing ltd susceptibility testing, prompt fluid resuscitation for septic shock, and standardized infection control measures such as hand cleaning. several recent global initiatives have developed guidelines for the syndromic management of severe influenza and other severe illness in resource-limited settings. during the h n influenza pandemic, the who assembled a group of experts to generate a document addressing management of severe respiratory distress and shock in resource-limited settings. more recently, similar advice was produced by the who for the clinical management of novel coronavirus from outbreaks in in the middle east (publication pending). the who integrated management of adult and adolescent illness (imai) program, a sister initiative to imci, created a comprehensive manual for the care of hospitalized patients by clinicians in district hospitals that includes sections on severe illness management. the european society of intensive care medicine also recently produced sepsis management guidelines targeted to resource-limited settings. these largely expert opinion-based documents fill major gaps in management guidelines but require quality clinical research to provide missing evidence and refine best-practice recommendations. a reminder of the importance of evidence generated by studies in at-risk populations was provided by the unexpectedly harmful effect of fluid boluses for kenyan children with severe infection in the feast (fluid expansion as supportive therapy) trial. severe illness, influenza-related and otherwise, causes a profound burden of disease in resource-limited settings. quality clinical management is an essential element in mitigating this burden. in support of this, a recent call to action by unicef, and several governmental and nongovernmental organizations, focuses global attention on treatment of preventable childhood deaths, including respiratory infections. syndromic management approaches to the care of severe illness seem practical and feasible. however, there are few data to guide the optimal management of severely ill patients in these resource-limited settings. while several recent initiatives now provide long awaited guidance for clinicians in resource-limited settings, a sustainable, global impact on outcomes due to severe influenza and other severe illness will require an ongoing and concerted international effort to implement, evaluate, and refine these guidelines. epidemiological characteristics and underlying risk factors for mortality during the autumn pandemic wave in mexico critically ill patients with influenza a(h n ) in mexico critically ill patients with influenza a(h n ) infection in canada a systematic review to inform institutional decisions about the use of extracorporeal membrane oxygenation during the h n influenza pandemic critical care services and h n influenza in australia and new zealand surveillance of illness associated with pandemic (h n ) virus infection among adults using a global clinical site network approach: the insight flu and flu studies critically ill children during the - 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t gf c authors: agbuduwe, charles; basu, supratik title: hematolological manifestations of covid‐ : from cytopenia to coagulopathy date: - - journal: eur j haematol doi: . /ejh. sha: doc_id: cord_uid: t gf c s emerging data from the management of patients with coronavirus disease (covid‐ ) suggests multisystemic involvement, including the hemopoietic system. the hematological manifestations of covid‐ include blood count anomalies notably lymphopenia and neutrophilia which are of prognostic significance. hyperferritinemia and elevated lactate dehydrogenase have also been associated with increased mortality. furthermore, there is considerable evidence of a distinct coagulopathy associated with covid‐ characterised by elevated d‐dimers and an increased risk of thrombotic events. this comprehensive review summarises the latest evidence from published studies and discusses the implications of the various hematological manifestations of covid‐ with a view to guiding clinical management and risk stratification in this rapidly evolving pandemic. severe acute respiratory syndrome (sars) and middle east respiratory syndrome (mers), have been caused by betacoronaviruses, sars-cov and mers-cov respectively in humans. [ ] sars-cov- shares % sequence identity with sars-cov, the virus responsible for the severe acute respiratory syndrome (sars) outbreak - . [ , ] while much of the pathogenesis of covid- remains to be unravelled, it is known that sars-cov- , like sars-cov, binds to host cells via its receptor, angiotensin converting enzyme (ace ), which is expressed across a wide range of human cell types including lung type ii pneumocytes and the endothelium of blood vessels. [ , ] the clinical spectrum of covid- ranges from asymptomatic to severe pneumonia or acute respiratory distress syndrome (ards) resulting in respiratory failure and death. [ , ] while covid- is considered to be primarily a respiratory infection, there is increasing evidence of multi-systemic complications of the disease. with more case series being published, a number of hematological manifestations have been now come to light. the key hematological abnormalities of covid- are summarised in table . the most commonly reported blood count abnormality is lymphopenia which occurs in %- % of patients. [ ] [ ] [ ] lymphopenia is also more frequent and the absolute lymphocyte count much lower in severe cases of covid- . [ ] [ ] [ ] in one study of hematological parameters in hospitalised covid- patients in singapore, the median nadir of the absolute lymphocyte count (alc) was significantly lower in patients requiring admission to intensive care unit (icu) ( . vs . x /l) as was neutrophilia ( . vs . x /l). [ ] another study from wuhan revealed a similar pattern with more severe cases having higher neutrophil ( . vs . × ⁹/l ; p < . ) counts, lower lymphocytes counts ( . vs . × ⁹/l ; p < . ), higher neutrophil-to-lymphocyte ratio (nlr) ( . vs . ; p < . ) as well as lower percentages of monocytes, eosinophils and basophils. [ ] in addition to a significant reduction in both cd + and cd + t lymphocyte subsets in covid- patients, severe cases had much lower cd + lymphocytes and a subsequent increase positively correlated with improved clinical outcomes. [ ] accepted article this article is protected by copyright. all rights reserved mild thrombocytopenia ( - x /l) has been reported in up to - % of covid- cases [ ] [ ] [ ] however, severe thrombocytopenia (< x /l) is unusual. in a case series of patients admitted to an intensive care unit in wuhan, a platelet count of < x /l was observed in only % of patients. [ ] similar findings were reported in a french cohort study with mild thrombocytopenia identified in about a quarter of covid- patients on admission to hospital and this was independently predictive of the risk of admission to icu, mechanical ventilation or death. [ ] furthermore, data from multiple studies suggest that anemia is not a prominent feature of covid- even in severe cases. [ , , ] a case series of autoimmune haemolytic anaemia occurring in covid- patients was recently published but about half of the patients in this study had an underlying haematological disorder known to be associated with autoimmune haemolysis. [ ] peripheral blood film and morphological features examination of the peripheral blood smear remains a crucial part of the hematological assessment and can often aid the diagnosis of many conditions. notable features reported in covid- cases include an increased frequency of reactive and plasmacytoid lymphocytes, [ ] [ ] significant left-shifted granulopoiesis with hypergranular, occasionally vacuolated neutrophils [ ] and leucoerythroblastic features [ ] . the presence of schistocytes or red cell fragments has not been reported. furthermore, from the experience of the authors, bone marrow hemophagocytosis can be a feature of severe covid- and this has been observed in a small number of patients managed at new cross hospital, wolverhampton, united kingdom. the bone marrow aspirate from one of the patients is shown in figure . this pathologic finding corroborates recent reports of cases of covid- meeting the clinical criteria for secondary hemophagocytic lymphohistiocytosis (shlh). [ , ] alongside other acute phase markers such as c-reactive protein (crp), procalcitonin and erythrocyte sedimentation rate (esr), an elevated ferritin has been associated with increased mortality in covid- . [ , , ] . significant elevation of lactate dehydrogenase (ldh) has also been observed in patients with severe disease. [ ] observations from the clinical, biochemical and serological manifestations of covid- strongly support an immunological basis for the severe manifestations of the disease. [ ] the marked elevation of proinflammatory markers such as il- β, il- , il- , il- , il- , tnf-α and ifnγ frequently seen in severe this article is protected by copyright. all rights reserved covid- , [ ] [ ] [ ] point to a state of disordered and exaggerated immune response to sars-cov- infection, often referred to as the "cytokine storm". several complex pathways have been implicated in the pathogenesis of covid- cytokine storm including the renin-angiotensin-aldosterone system (raas), jak/stat and complement activation pathways. [ , , ] furthermore, parallels can be drawn with the cytokine release syndrome (crs) associated with chimeric antigen receptor (car) t cell therapy and hemophagocytic lymphohistiocytosis (hlh), which are both states of immune dysregulation and hyperinflammation frequently encountered by haematologists. crs, thought to be due to t cell activation, is characterised by marked elevation of inflammatory markers and cytokines notably il- , fever, hypotension and respiratory insufficiency following the infusion of car t cells or other immune therapies. [ , ] similarly, hlh is characterised by uncontrolled activation of cytotoxic t lymphocytes, natural killer cells and macrophages resulting in hypercytokinemia and immune-mediated organ damage. [ ] the aetiology of hlh may be primary (as a result of inherited genetic mutations) or secondary to other conditions including viral infections and often presents with features of crs in addition to histological evidence of hemophagocytosis, cytopenias, hyperferritinemia, organomegaly, coagulopathy and multi-organ failure. [ ] secondary hlh (shlh) is probably an under-recognised feature of severe covid- . a recent case series of autopsies of patients who died from covid- published as a preprint, [ ] reported histologic evidence of haemophagocytosis within pulmonary and hilar lymph nodes in the majority of cases. however, in a study of critically-ill covid- patients admitted to an intensive care unit, only ( . %) met the cut off for shlh using the hscore. [ ] this observation was most likely due to the absence of certain cardinal features such as organomegaly and hypofibrinogenemia in the majority of covid- patients, possibly indicating a distinct mechanism of sars-cov- -associated shlh. understanding the link between crs, hlh and covid- cytokine storm is crucial because this would expedite the repurposing of therapies for severe covid- . indeed, early clinical studies have indicated that certain immunomodulatory therapies used in crs and shlh such as dexamethasone [ ] , tocilizumab [ ] and anakinra [ ] may be effective in severe covid- . given the pivotal role of il- signalling in crs and its prognostic significance in covid- , [ ] there has been considerable interest in the therapeutic potential of the il- receptor antagonist, tocilizumab. two separate case reports of the use of tocilizumab in acute chest syndrome of sickle cell disease precipitated by covid- reported rapid responses in an adult [ ] and a teenager [ ] . tocilizumab administration also resulted in clinical improvement in a patient with covid- and multiple myeloma. [ ] a recently published retrospective cohort study of patients treated with tocilizumab, the largest to date, observed a significant reduction in the risk of mechanical ventilation or death in tocilizumab-treated patients accepted article compared with controls. [ ] several randomised clinical trials of tocilizumab are ongoing and the results of these studies are eagerly awaited. recent data emerging from the management of patients with covid- suggests an increased thrombotic tendency. approximately one-third of patients with covid- had ct scan evidence of pulmonary embolism (pe) in a french study. [ ] notably, two thirds of the patients without pe in this cohort also had elevated d-dimers with a higher cut off value of µg/l being more predictive of pe in this cohort. a dutch study of covid- patients admitted to icu similarly identified a % incidence of thrombotic events including pe, deep vein thrombosis (dvt) and ischaemic strokes despite all patients having received prophylactic anticoagulation. [ ] a retrospective study of covid- patients admitted to icu identified dvt in % with advanced age, lower lymphocyte counts and elevated d-dimers being significant risk factors. [ ] the prognostic importance of d-dimer testing was further demonstrated in a prospective study of hospitalised covid- patients which revealed that significantly higher d-dimer and prolonged prothrombin time (pt) were associated with a higher probability of mortality. [ ] in a study investigating prolonged activated partial thromoplastin time (aptt) in patients with covid- , the lupus anticoagulant was detected in %. [ ] in addition, markedly elevated von willebrand factor (vwf) levels (vwf:antigen- %, vwf:activity- %) and factor viii (clotting activity of %) in addition to antiphospholipid antibodies were observed in a patient with severe covid- . [ ] taken together, these findings strongly support the existence of a syndrome of covid- -associated coagulopathy characterised by derangements in clotting tests (pt and aptt), elevated d-dimer and an increased thrombotic tendency. in a study comparing coagulation parameters in hospitalised covid- patients, ( . %) of nonsurvivors met the international society on thrombosis and haemostasis (isth) criteria for overt disseminated intravascular coagulation (dic) compared with ( . %) of survivors. [ ] therefore, the isth dic scoring system which includes platelet count, fibrinogen, d-dimer and prothrombin time ( table ) is likely to be a useful prognostic tool for covid- -associated coagulopathy. the coagulopathy of covid- appears to be distinct from dic due to other causes in elevated fibrinogen, [ ] modest prolongation of the aptt and the absence of schistocytes on the blood film. [ ] interestingly, despite the derangements in coagulation tests, abnormal bleeding is unusual. [ , ] furthermore, much of the data for coagulopathy in covid- has been from hospitalised patients with the severe form of the disease. it is yet unclear but unlikely that patients with mild covid- are at increased risk of thrombosis. while still not fully understood, the immune response to sars-cov- is thought to be two-phased; an initial t-cell mediated adaptive response during the pre-symptomatic or non-severe stage which in a accepted article proportion of individuals is ineffective, leading to uncontrolled viral replication, triggering an exaggerated innate immune response which results in multi-organ failure in the most severe cases. [ , ] the dysregulated secretion of pro-inflammatory cytokines perpetuates hyperinflammation and is thought to be the basis of the cytokine storm in covid- . this two-phased model of the immune response would probably account for the lack of efficacy of some immunosuppressive therapies in non-severe cases of the disease. [ ] furthermore, aging is generally associated with diminished adaptive immune function and an exaggerated innate immune response to pathogens, notably a shift towards myelopoiesis and neutrophil accumulation in animal models. [ , ] these age-related changes in the immune system may, at least partly, explain the increased mortality of covid- among the elderly. a lot is still not known about the marked difference between immune responses to sars-cov- between individuals and it is likely that genetic and environmental factors also play a role. of particular interest is the association between certain abo blood group genotypes and the likelihood of severe covid- . in a recently published genomewide association study, blood group a and a few other single nucleotide polymorphisms (snps) were associated with an increased risk of covid- -induced respiratory failure (blood group o was apparently protective). [ ] interestingly, blood group a has also been shown to be associated with increased odds of thromoboembolic and cardiovascular events. [ , ] these findings are hardly surprising because blood group a individuals (as well as other non-o blood groups) are known to have higher plasma von willebrand levels [ , ] and abo blood group antigens have innate immune functions. [ ] currently, the evidence base for the clinical management of covid- is mostly limited to case series and other relatively small observational studies of hospitalised patients. despite the limitations, these studies provide useful insight into the manifestations of the disease. similar to findings in sars patients, [ ] lymphopenia is the most commonly reported hematological abnormality in covid- and recent data shows that it can be predictive of disease severity. however, as an isolated finding, lymphopenia is not specific for covid- and is a common finding in the elderly. the predictive utility of lymphopenia is likely to be improved when the trend is considered alongside other parameters particularly the presence of neutrophilia a higher neutrophil-lymphocyte ratio. furthermore, assessments of lymphocyte subsets in covid- patients have revealed a significant reduction in t and b lymphocytes along with natural killer (nk) cells, more marked among patients with severe disease. [ , , ] of particular interest was the finding of reduced regulatory t cells (treg) in severe cases. [ ] the precise mechanism of the development of lymphopenia in covid- is not known but lymphocytes have been shown to express ace [ ] and lymphoid cell apoptosis may be a consequence of sars-cov- infection. [ ] [ ] trafficking accepted article of lymphocytes away from the peripheral blood to the lungs or other sites of infection may also play a role. [ ] the neutrophilia observed in severe cases of covid- is most likely a response to the cytokine storm which has been implicated in the most severe manifestations of the disease. furthermore, significant elevations of acute-phase markers such as ferritin, crp and procalcitonin have been associated with mortality from covid- and these biomarkers are positively correlated with increased proinflammatory cytokines such as interlukin- and tnf-α. [ , ] therefore, serial monitoring of these markers may facilitate early therapeutic intervention with experimental immunomodulatory agents. with recent reports of thrombotic complications in patients with covid- , there is increasing recognition of a distinct coagulopathy associated with covid- . the underlying mechanism is likely to be multifactorial including direct endothelial damage from sars-cov- or immune cells, [ ] inflammatory cytokine-induced activation of the coagulation cascade [ ] , the development of antiphospholipid antibodies and an increase in acute phase pro-coagulants such as factor viii and fibrinogen. derangements of coagulation reported among covid- cases include prolongation of the pt and to a lesser extent, aptt. the relative shortening of the aptt is possibly due to significantly increased levels of factor viii. furthermore, hyperviscosity has been postulated as a possible link between hyperinflammation and coagulopathy following the association of an increased risk of arterial and venous thromboses with plasma viscosity levels greater than . centipoise (twice the upper limit of normal) in a study of icu patients with covid- . [ ] however, the hyperviscosity in covid- is most likely due to the marked elevation of acute phase proteins and does not completely explain the severe thrombophilic state which is out of proportion to other conditions associated with comparable degrees of plasma viscosity. plasma exchange may nevertheless be beneficial as this could theoretically replace most of the elevated circulating cytokines and pro-coagulant factors. the use of convalescent plasma may, in addition, provide neutralising antibodies against sars-cov- and a small-scale clinical trial has reported modest but encouraging results in severely-ill but not in critical covid- patients. [ ] the lack of efficacy in the latter group probably reflects the irreversibility of end-organ damage from hyperinflammation and the relatively minimal contribution from sars-cov- at this stage of the disease. this article is protected by copyright. all rights reserved the mainstay of the management of cac is prevention as well as early recognition and treatment of thrombotic events. regarding the prevention of thrombotic events in hospitalised covid- patients, evidence from china suggests that prophylactic heparin reduces mortality in high-risk patients. [ ] consequently, the recently published isth interim guidance recommends prophylactic anticoagulation with low molecular weight heparin (lmwh) for all hospitalised patients with covid- . [ ] lmwh is the preferred anticoagulant as it does not interfere with the pt or aptt and the anti-inflammatory properties of heparins could indeed provide additional benefits in this setting. [ ] unfractionated heparin with anti-xa monitoring may be a useful alternative given its widespread use in the intensive care setting but close monitoring for the development of heparin-induced thrombocytopenia (hit) is recommended. furthermore, since cac is not commonly associated with a bleeding phenotype, anticoagulation is recommended regardless of derangements in the pt or aptt, provided other contraindications are excluded. moreover, transfusion of blood products such as fresh frozen plasma (ffp) or cryoprecipitate to correct abnormal clotting parameters in the absence of bleeding is not recommended as this could be detrimental. cautious transfusion of blood products is however indicated in bleeding patients. given the histological finding of widespread fibrin deposition consistent with microvascular thrombosis reported in covid- cases, [ ] clinical trials are needed to evaluate the efficacy of experimental fibrinolytic therapies such as tissue plasminogen activator (tpa). [ ] in view of the reported high incidence of thrombotic events despite prophylactic anticoagulation in critically ill patients with covid- , [ , , ] there is a strong argument for intensification of anticoagulation in this setting but the benefit of this strategy remains to be determined due to the lack of clinical trial data. while there is currently insufficient evidence to recommend therapeutic anticoagulation in all patients with covid- in the absence of venous thromboembolism (vte), an individualised riskbased approach to intensification of anticoagulation may be considered. in summary, severe covid- represents a state of immune dysregulation and hyperinflammation which account for the multisystemic manifestations, including the hematological anomalies associated with the disease. therefore, monitoring of hematological parameters such as the absolute lymphocyte count, neutrophil-to-lymphocyte ratio and d-dimers can offer prognostic insight in the management of covid- and will help with early identification of the high risk group of patients requiring more intensive care. in view of the increased thrombotic risk associated with covid- , prophylactic anticoagulation with low accepted article molecular weight heparin is recommended for all hospitalised patients with the disease and clinical trials are needed to investigate the role of more intensive anticoagulation and other experimental therapies. in addition, further translational research is needed to fully unravel the pathogenesis of covid- , particularly the host immune response, with a view to developing effective therapies. a novel coronavirus from patients with pneumonia in china modes of transmission of virus causing covid- :implications for ipc precaution recommendations the proximal origin of sars-cov- genomic characterisation and epidemiology of novel coronavirus: implications for virus origins and receptor binding a pneumonia outbreak associated with a new coronavirus of probable bat origin 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and thrombophilia? effect of convalescent plasma therapy on time to clinical improvement in patients with severe and life-threatening covid- accepted article this article is protected by copyright. all rights reserved isth interim guidance on recognition and management of coagulopathy in covid- using heparin molecules to manage covid- complement associated microvascular injury and thrombosis in the pathogenesis of severe covid- infection: a report of five cases tissue plasminogen activator (tpa) treatment for covid- associated acute respiratory distress syndrome (ards): a case series high incidence of venous thromboembolic events in anticoagulated severe covid- patients pulmonary embolism in covid- patients: awareness of an increased prevalence running title: poissy et al.; covid- and pulmonary embolism n the scoring system of the scientific and standardisation committee on disseminated intravascular coagulation of the international society on thrombosis and haemostasis: a -year overview the authors wish to thank dr richard whitmill for the bone marrow aspirate image included in this publication.funding statement: ca is a recipient of a clinical research fellowship funded by the cruk-city of london centre clinical academic training programme award [c /a ] the authors declare no conflict of interests in relation to this article. this article is protected by copyright. all rights reserved key: cord- - dn a authors: peng, liang; gao, zhi-liang; wang, yu-ming; he, deng-ming; zhao, jing-ming; bai, xue-fan; wang, xiao-jing title: clinical manifestations and laboratory tests of aechb and severe hepatitis (liver failure) date: - - journal: acute exacerbation of chronic hepatitis b doi: . / - - - - _ sha: doc_id: cord_uid: dn a this chapter describes the clinical symptoms and signs of aechb and hbv aclf, classification, grading of hbv aclf and their features, diagnostic principles and standards in liver pathology, biochemistry, and virology of hbv aclf. . liver failure is defined as serious damage to the liver cause by a variety of etiologies, leading to liver function disorder or even decompensation, and clinical syndromes with coagulopathy, jaundice, hepatic encephalopathy, and ascites. . severe hepatitis b can be indicated pathologically by apparent hepatocellular necrosis, including extensive multifocal, confluent, bridging, sub-massive or massive necrosis. . laboratory tests during the course of severe exacerbation of chronic hepatitis b can reflect pathological changes and liver function in a timely manner, providing objective and informative reference data for evaluation of disease severity and treatment efficacy. among the most important laboratory tests are those for prothrombin activity, international normalized ratio, and increases in total bilirubin concentration. . severe hepatitis b is associated with interactions between the virus and host factors. detection of hbv dna, hbv genotype, quasispecies and hbv mutation can provide important theoretical bases for the prevention, control or mitigation of the progress of severe hepatitis b. . noninvasive imaging modalities can be used to visualize the entire liver and parts of it. measuring liver volume to evaluate liver size and liver reserve capacity is regarded as important in diagnosis, surgical approach and prognostic evaluation of patients with severe exacerbation of chronic hepatitis b and liver failure. . model for end-stage liver disease (meld) is the first quantitative method developed to assess whether a patient with liver failure requires a liver transplant. the predictive value of the meld model has been improved by the meld-na, imeld, and meso models. several other valuable prognostic models have been developed. for example, for patients with hbv-aclf, the established tppm scoring system was found to be more predictive than meld score. can reflect pathological changes and liver function in a timely manner, providing objective and informative reference data for evaluation of disease severity and treatment efficacy. among the most important laboratory tests are those for prothrombin activity, international normalized ratio, and increases in total bilirubin concentration. . severe hepatitis b is associated with interactions between the virus and host factors. detection of hbv dna, hbv genotype, quasispecies and hbv mutation can provide important theoretical bases for the prevention, control or mitigation of the progress of severe hepatitis b. . noninvasive imaging modalities can be used to visualize the entire liver and parts of it. measuring liver volume to evaluate liver size and liver reserve capacity is regarded as important in diagnosis, surgical approach and prognostic evaluation of patients with severe exacerbation of chronic hepatitis b and liver failure. . model for end-stage liver disease (meld) is the first quantitative method developed to assess whether a patient with liver failure requires a liver transplant. the predictive value of the meld model has been improved by the meld-na, imeld, and meso models. several other valuable prognostic models have been developed. for example, for patients with hbv-aclf, the established tppm scoring system was found to be more predictive than meld score. liang peng, zl huang, yy mei and zhi-liang gao currently, both clinical and pathophysiological diagnoses are made of severe hepatitis (liver failure) in china. according to the guideline for the prevention and treatment of viral hepatitis ( ), severe hepatitis is classified as acute severe hepatitis, subacute severe hepatitis, and chronic severe hepatitis. acute severe hepatitis is initially diagnosed due to acute jaundice that rapidly progresses to liver failure within weeks. subacute severe hepatitis can be identified in patients with acute jaundice hepatitis that progresses to liver failure anywhere from days to weeks. chronic severe hepatitis often develops with pre-existing chronic liver diseases. the clinical manifestations of chronic severe hepatitis are similar to those of subacute severe hepatitis in some patients, or, in some patients, appear similar to decompensated cirrhosis at disease onset. the diagnostic criteria for severe hepatitis in china remain to be fully developed and hence have not been introduced internationally. to meet the clinical requirements and standardize the diagnosis and therapy of liver failure, the branch of infectious diseases and the branch of hepatology of the chinese medical association invited experts in china to develop the first guidelines for the diagnosis and therapy of liver failure in . in those guidelines, liver failure refers to severe liver damage caused by multiple factors. that damage to the liver results in either the severe impairment or decompensation of synthesis, detoxication, excretion, and biotransformation in the liver and subsequent clinical manifestations characterized by coagulation disorder, jaundice, hepatic encephalopathy, and ascites. on the basis of pathological features and disease progression, liver failure is classified as acute liver failure (alf), subacute liver failure (salf), acute-on-chronic liver failure (aclf), and chronic liver failure (clf). alf is characterized by the rapid appearance of clinical manifestations. patients with alf usually develop a clinical syndrome of liver failure characterized by highgrade hepatic encephalopathy (he, >grade ) within weeks. patients with salf typically present with a clinical syndrome of liver failure anywhere from days to weeks. finally, aclf refers to the acute decompensation of the liver function in the presence of pre-existing chronic liver diseases, and clf refers to chronic decompensation of the liver function characterized by ascites or portal hypertension, coagulation disorder, and he due to progressive liver dysfunction in the presence of hepatic cirrhosis. the published guidelines systemically and extensively reflect the current status of the diagnosis and therapy of liver failure. in addition, the guidelines, for the first time, focus on liver failure rather than severe hepatitis, which broadens our horizons and highlights practicability. in china, acute severe hepatitis, subacute severe hepatitis, and chronic severe hepatitis correspond closely to alf, slf, and aclf, respectively, as illustrated in table . . in some patients, chronic severe hepatitis is similar to clf in other countries. on the basis of available guidelines for liver failure, we define severe hepatitis b as liver failure due to hepatitis b virus infection. clf is the most common, and alf and slf are rare. acute exacerbation of chronic hepatitis b (aechb) is a dynamic process, including mild, moderate, severe chronic hepatitis b and chronic aclf defined in above guidelines. the reference index of abnormality in laboratory examination is shown in table . . in addition to viral replication, other factors also contribute to the pathogenesis of hepatitis b-induced liver failure, such as concomitant infection of other hepatitis viruses (especially the hepatitis e virus), immune status, pregnancy, drug and/or alcohol consumption, concomitant bacterial infection, mental stress, and concomitant disease processes (e.g., hyperthyroidism). the liver is the largest solid organ in humans and has complex functions. hepatic parenchymal cells are responsible for metabolism, secretion, synthesis and bioconversion. factors that can cause severe damage to hepatocytes (i.e., parenchymal cells, kupffer cells) may result in disorders of metabolism, secretion, synthesis, detoxication and immunity. in turn, that damage can lead to jaundice, liver shrinkage, coagulation dysfunction, hemorrhage, secondary infection, hepatorenal syndrome, he, and other clinical entities described in detail here. the physical condition of patients deteriorates, and affected individuals usually develop weakness, extreme fatigue, and a severely diminished quality of life. they frequently require assistance to perform basic personal needs, such washing their face, brushing their teeth, and using the toilet. in the early stage of jaundice, in addition to developing extreme fatigue, gastrointestinal symptoms become evident, including extremely poor appetite, anorexia, intolerance of oily foods, nausea, vomiting, abdominal discomfort, and hiccups. in the jaundice stage, the gastrointestinal symptoms deteriorate further. patients can develop refractory vomiting, hiccups, evident abdominal distension, and reduced/ lack of borborygmus. clinically, patients initially note their urine color darkens, becoming a strong-tea like color. next, a yellowish pigmentation of the ski and conjunctival membranes develops. that jaundice progressively becomes deeper, characterized by hepatocellular jaundice. in this stage, serum bilirubin increases rapidly. in fact, the daily increment in serum bilirubin may be > μmol/l (> mg/dl). the sulfur-containing amino acids in the intestine are degraded into mercaptans that have the odor of rotting fruit. mercaptans cannot be metabolized in the liver and are therefore excreted from the respiratory tract. this distinctive odor is specifically noted in patients with he. the severity of hepatic foetor may, in some cases, reflect the severity of liver injury. the occurrence of coagulation dysfunction is primarily ascribed to the reduced synthesis of coagulation factors by the liver. a majority of the both coagulation and anticoagulant factors are synthesized in the liver. in addition, some coagulationrelated factors and their inhibitors are also metabolized in the liver. the outcome of coagulation dysfunction is dependent on the severity of damage to the hepatocytes. thus, even patients in an early stage of liver failure may present with coagulation dysfunction. prothrombin (pt) activity is often abnormal in the early stages of liver failure and may therefore serve as a sensitive indicator for the prognosis of liver failure. common clinical manifestations of coagulation dysfunction are mucocutaneous bleeding (i.e., spontaneous bruising, gingival bleeding, subconjunctival hemorrhage), ecchymosis at the site of injection/puncture, and purpura in more severe cases. gastrointestinal bleeding is also common in affected individuals, whereas bleeding into/from the genitourinary tract, lung, kidney, and retroperitoneum is rare but occasionally observed in some patients. if intracranial hemorrhage develops, it is frequently life threatening. in ahf, the incidence of bleeding and severe bleeding is as high as and > %, respectively. another cause of coagulation dysfunction is thrombocytopenia and platelet dysfunction. of the two, thrombocytopenia is more common. because platelets are derived from megakaryocytes in bone marrow, bone marrow fibrosis and either reduced bone marrow regeneration or invasion of lymphoma cells in the bone marrow can reduce the number of platelets. platelets perform multiple activities, including adhesion, aggregation, release, and shrinking blood clots. additionally, they play an important role in coagulation. platelet dysfunction may also increase capillary permeability and fragility, which may cause either spontaneous bleeding of the skin and mucous membranes or difficult hemostasis following vascular injury. in patients with slf, thrombocytopenia is mainly diagnosed in the latter stage of disease in which massive hepatocyte necrosis leads to posthepatic cirrhosis, portal hypertension, and hypersplenism. in clf patients, thrombocytopenia might be present, and hepatocyte necrosis may aggravate portal hypertension and hypersplenism, resulting in worsening thrombocytopenia. splenomegaly and splenic sinus hyperplasia increase the phagocytosis and destruction of platelets. further, splenomegaly can cause enlargement of the platelet pool within the spleen. as a result, the platelets in the spleen may account for > % of platelets in the body. the above pathological changes may finally cause a reduction in the circulating platelets. the reason for thrombocytopenia in liver disease patients without hypersplenism is still poorly understood and might be ascribed to following factors ( ) the hepatitis b virus may significantly inhibit the megakaryocyte system of the bone marrow, resulting in reduced production of platelets; ( ) the thrombopoietin (tpo) level is reduced. the division of megakaryocytes into platelets in the bone marrow is controlled by both megakaryocyte colony stimulating factor (meg-csf) and tpo. meg-csf primarily regulates the proliferation of megakaryocyte progenitor cells, whereas tpo stimulates the maturation of megakaryocytes and production of platelets. tpo is almost exclusively produced by hepatocytes, and only a minority of tpo is produced in the kidney and other organs. tpo is a key factor affecting the production of platelets, and the synthesis of tpo is reduced significantly in patients with either severe hepatitis or hepatic cirrhosis, which affects the production of platelets. in patients with parenchymal liver diseases, abnormalities of platelets are present in both quality and quantity. for example, when the platelet membrane glycoprotein gpi -ix is reduced, the aggregation of platelets following ristocetin treatment and the shrinkage of blood clots are markedly compromised; and ( ) patients with liver diseases usually develop immune dysfunction and are therefore susceptible to infection. bacterial toxins and systemic inflammatory response syndrome may also cause thrombocytopenia. one published study of icu patients found that infection was an independent risk factor of thrombocytopenia. he he is both a neuropsychiatric syndrome, a type of central nervous system dysfunction, and metabolic disturbance due to hepatocellular dysfunction and portosystemic shunting. he is clinically characterized by mental and neurological abnormalities, such as abnormal personality and behaviors, irritability, sleep perversion, drowsiness, and complete loss of consciousness or coma. he is one of the major causes of severe complications and death in patients with liver failure and is typically classified into one of the four following stages: stage : the prodromal stage. this stage usually manifests with mild abnormal personality changes and behaviors, such as euphoric excitement, indifference, taciturnity, being sloppily dressed, and inappropriate defecation/urination. the affected individual can usually provide correct responses to questions but they are inarticulate and have slow speech. flapping tremor/hepatic tremor might also be present. to test for flapping tremor, patients are asked to close their eyes with their arms stretching straight, elbows flexed, palms in dorsal extension, with separated fingers. a positive response is determined when the metacarpophalangeal joint, wrist, elbow, and shoulder show irregular movements (jitter) when held in that position within s. physicians may also ask the patients to hold the their hand for min. if the physician feels the hand tremor, the test suggests a positive diagnosis of flapping tremor. the condition is caused by afferent dysfunction of joint-reticular formation of the brainstem and a characteristic neurological manifestation. that said, flapping tremor has no specificity fro he and can also be found in patients with either uremia or hypoxemia due to chronic respiratory disease/heart failure. the presence of flapping tremor in a patient with severe liver disease, however, is helpful for early diagnosis of he. patients with he usually have a normal electroencephalogram. stage of he lasts anywhere from several days to several weeks. several patients with he in the prodromal stage may have no evidence of clinical symptoms; therefore, misdiagnosis is possible. stage : the precoma stage. patients with he in this stage usually presents with confusion, sleep and behavioral disorders, and symptoms as described in the prodromal stage further deteriorate. patients suffer from disorientation and understanding disorders as well as conceptual confusion over time, place, and person. patients are unable to perform simple intellectual composition (e.g., building blocks, arranging matchstick into pentagon), and have decreased computing capacity (e.g., - and continuing). slurred speech, writing disorders, and abnormal behaviors are also common. sleep perversion and daytime sleep and night awaking may be present. further, hallucinations, fear, and mania are also observed, and some patients can be misdiagnosed with mental diseases. patients with liver failure in this stage usually have evident neurological signs such as tendon hyperreflexia, increased muscle tone, ankle clonus, and presence of the babinski sign. flapping tremor and an abnormal electroencephalogram can also be observed. patients may also suffer from uncontrolled muscular activities and ataxia. stage : the lethargic stage. patients with he in the lethargic stage mainly manifest lethargy and insanity, and neurological signs continue and deteriorate. in the majority of time, patients are in a lethargic state, but can be waken up. patients respond to questioning, but may present confusion and hallucination. flapping tremor is also present. muscular tension increases, and there is resistance in the passive limb movements. pyramidal signs and abnormal waves in eeg can also be noted. stage : the coma stage. patients have complete loss of consciousness and are unable to be awakened. in a light coma, patients are responsive to painful stimuli and uncomfortable postures, have tendon hyperreflexia, and increased muscular tension. patients in this stage are usually unable to co-operate during an examination, and a flapping tremor may not be inducible. in a deep coma, various reflexes disappear; muscular tension reduces; pupils become dilated; and there are paroxysmal convulsions, ankle clonus, hyperventilation, and abnormalities on an electroencephalogram. stage of he is an important indicator of severity of disease. it may reflect not only the severity of brain damage but also the severity of liver disease. it is important to recognize that there is no clear boundary between two neighboring stages and that there might be some overlap between two neighboring stages (therefore missing the middle stage of he). when the disease condition either deteriorates or improves after therapy, the severity of he may be reduced by one or two stages. as mentioned above, the initial symptoms of he are personality changes. patients with extrovert personalities (i.e., lively, cheerful) may become depressed, whereas patients with introverted personalities (i.e., withdrawn, reticent) may become euphoric and garrulous. the second most common symptom is a change in behaviors. initially, patients have sloppy behaviors, such as meaningless behaviors like scattering garbage all over the place and defecating/urinating anywhere, looking at clothes, and touching the bed. those changes are usually only identified by close observation and careful experience. there are also changes in sleep habits. patients are often drowsy during the daytime but have difficult sleeping at night or show sleep perversion, which predicts imminent he. hepatic foetor is also an important feature of he. he patients usually have brain edema and present with nausea, vomiting, dizziness, headache, and either irregular breathing or even apnea. as blood pressure increases there might be a paroxysmal or sustained increase in systolic blood pressure. bradycardia may be also observed. muscular tension can increase or the patient can develop a decerebrate posture or even opisthotonus with severe he. the pupillary light reflex can become blunt/absent, the pupils can become mydriatic, and anisocoria can occur. achilles and knee tendon hyperreflexia may be observed. it is important to note that some signs might not be obvious in a patient with late-stage he. in clinical practice, clinicians may indirectly evaluate the severity of brain edema according to chemosis. accurate evaluation of brain edema is dependent on the subdural, epidural, or cerebral parenchymal measurement of intracranial pressure. the normal intracranial pressure is < . kpa ( mmhg), and brain edema is diagnosed once intracranial pressure is > mmhg. the most important sign of he is flapping tremor, which means the presence of he in stage ii. in addition, thinking and intelligence tests (such as number connection test, signature test, mapping test, and computing capability test) are abnormal in he patients. in some he patients (especially those with hyperammonemia due to he), slow waves with high amplitude may be observed on electroencephalogram, and positive-evoked potential is also a characteristic change. brain edema is a complication of alf. typical clinical manifestations of brain edema are sustained increase in blood pressure, abnormal pupils, irregular respiration, and papilledema. more than % of patients with he in stage or are likely to develop brain edema, and severe brain edema may result in cerebral hernia. brain edema has the clinical presentations of increased intracranial pressure and cerebral dysfunction, which can sometimes overlap with the manifestations of he. it is therefore sometimes difficult to differentiate the two, potentially resulting in misdiagnosis. he patients with brain edema may present dysphoria, irascibility, and increased muscular tension, which are more common than in patients with he without brain edema. if there are concomitant changes in pupils and respiration together with convulsions and/or seizures, cerebral hernia is suspected. in the late stages of liver failure, patients may develop intracranial hemorrhage, causing respiratory and circulatory arrest and even sudden death. thus, once cardiopulmonary arrest of unknown cause is present, intracranial hemorrhage should be considered. concomitant gastrointestinal bleeding in patients with severe hepatitis can be caused by multiple factors, including ( ) decreased coagulation factor synthesis by hepatocytes and/or significant inactivation of active coagulation factors in the liver; ( ) endotoxemia and disseminated intravascular coagulation consuming a large amount of coagulation factors; ( ) hypersplenism causing abnormalities in the quality and quantity of the platelets; ( ) portal hypertension causing the rupture of esophageal and gastric varices; and ( ) stress response in severe hepatitis leading to diffuse gastric corrosive erosion. of the possible complications occurring in liver failure patients, bleeding is the most common and severe. in clinical practice, gastrointestinal bleeding with severe hepatitis seems to make the primary disease worse. it may worsen liver ischemia and hypoxia and aggravate liver dysfunction and ascites. blood in the gastrointestinal tract can be degraded into ammonia and increase the production of sulfur-like substance, resulting in he. in addition, bleeding may reduce immune function, which make infections difficult to control. the reduction in effective circulating blood volume may also induce hepatorenal syndrome. taken together, bleeding may cause multiple organ dysfunction, thereby complicating treatment and reducing the success rate of therapy. the causes of upper gastrointestinal bleeding are different among patients with different types of liver failure. in alf and slf, bleeding is related to reduced synthesis of coagulation-related factors and stress-induced gastric mucosal lesions. in clf, however, rupture of esophageal and gastric varices and gastric mucosal lesions secondary to portal hypertension are the main causes of gastrointestinal bleeding. in some cases, there is more than one cause of bleeding. in liver failure, the ability of the monocyte-macrophage system to clear intestinerelated endotoxins is reduced significantly, which may lead to intestine-related endotoxemia and deterioration of liver function. this clearly forms a vicious cycle and may cause multiple organ failure if it is severe enough. in addition, patients usually have compromised immune function and are susceptible to infection. invasive manipulations and use of broad-spectrum antibiotics and immunosuppressants further increase the possibility of secondary infection. concomitant infection in liver failure patients has the following characteristics: ( ) a high incidence; ( ) infection may occur at different sites either simultaneously or sequentially, and abdominal and biliary tract infection is the most common. once pulmonary infection is present, the disease condition will likely deteriorate, directly causing death; ( ) a majority of infections are nosocomial infection, and pathogens are usually resistant to common antibiotics, making therapy challenging; ( ) the pathogens causing infection are diverse but mainly gram-negative bacteria, although the incidence of gram-positive and fungal infections is increasing; ( ) infection is closely related to the prognosis for liver failure patients. in sum, the more severe the disease, the higher the incidence of infection is and secondary infection may worsen the condition or cause death. the early diagnosis of secondary infections is based on clinical findings such as signs of infections (i.e., fever, increase in peripheral white blood cells, deterioration of primary disease, specific symptoms of infection of a particular organ). some patients may not present with an obvious fever and instead only show focal signs of infection. for example, in spontaneous bacterial peritonitis, examination could reveal abdominal tenderness and rebound tenderness and a slight increase in peripheral white blood cells and polymorphonuclear proportion (although they are in normal ranges). in contrast, pulmonary infections can present only with fever while the respiratory symptoms are not obvious/absent and thoracic radiographs fail to show abnormalities in affected patients. in such cases, computed tomography is required to identify the pulmonary lesions. in addition, liver failure patients are vulnerable to fungal infection, especially for those receiving long-term therapy with broad-spectrum antibiotics. gastrointestinal candidiasis is the most common fungal infection. oral candida albicans infection is characterized by thickening and a bean residue-like coating on the tongue, gastrointestinal fungal infection is characterized by increased stool frequency and stool with mucus, and pulmonary fungal infection (especially aspergillus infection) is a severe complication of liver failure that can progress rapidly and has a high mortality rate. once a pulmonary fungal infection is suspected, computed tomography of the thorax should be performed to confirm the diagnosis, and effective antifungal therapy should be initiated as early as possible. hepatorenal syndrome (hrs) refers to progressive functional renal failure in the absence of primary kidney disease in patients with severe liver diseases. hrs is most often diagnosed in the late stages of severe hepatitis and hepatic cirrhosis. the main clinical manifestations of hrs include: . late stages of liver failure; . renal failure after a reduction in effective circulating blood volume (e.g., water and electrolyte disorder, following paracentesis for ascites, excessive urination due to diuresis, gastrointestinal bleeding, secondary infection, vomiting, and diarrhea). however, hrs may present abruptly with no evident/discoverable causes; . hrs is often found in patients with moderate to severe ascites; . hrs has no significant relationship with jaundice and he; and . blood pressure reduces during hrs. thus, when patients are treated with propranolol for portal hypertension, physicians should pay attention to the baseline blood pressure because reduction in blood pressure after pharmacotherapy may reduce the blood supply to the kidney and decrease glomerular filtration rate, inducing hrs; . the abrupt decrease in urine output suggests the presence of hrs. diuretics usually fail to increase the urine output. patients often have a reduction in urine sodium and concomitant hyponatremia; . urinalysis shows similarities to prerenal azotemia but displays opposite features to acute tubular necrosis; . the symptoms of uremia may overlap with those of liver failure and cause the deterioration of original symptoms. in patients with progressive liver diseases, secondary renal dysfunction is closely related to the deterioration of their general condition, suggesting the aggravation of liver failure. in addition, the presence of uremia may contribute to metabolic complications. coagulation dysfunction in liver disease patients may be deteriorated due to compromised aggregation of platelets during uremia. uremia may also aggravate immune dysfunction. on the basis of clinical characteristics, hrs can be classified into two types. type i hrs is rare, has an acute onset, and is characterized by progressive renal dysfunction. serum creatinine may be either × that at baseline (i.e., > μmol/l or . mg/dl) within weeks or creatinine clearance decreases by % within h (i.e., creatinine clearance of < ml/min). patients with type i hrs have a poor prognosis, with % of patients dying within weeks of diagnosis and only % of patients can survive for > months. the course of disease is short, and symptoms of uremia are not obvious. in type ii hrs, which is often found in clf patients with pre-existing hepatic cirrhosis, has a chronic onset. ascites patients with type ii hrs are usually nonresponsive to diuretics. in type ii hrs, renal failure shows a slow progression (lasting for several weeks to months), but the survival rate of patients is lower than that of hepatic cirrhosis patients with ascites. the main clinical consequence is refractory ascites nonresponsive to diuretics in patients with type ii hrs. a follow-up study of hepatic cirrhosis patients with ascites showed the accumulative incidence of hrs was % within year and % within years. a retrospective study showed about % of patients with ascites on admission had hrs and hrs patients accounted for % of hepatic cirrhosis patients died. however, for hepatic cirrhosis patients, the -year and -year incidence of hrs is % and % after development of ascites. a majority of patients ( - %) die within weeks after development of azotemia. hps refers to a series of pathophysiological changes and clinical manifestations (including hypoxemia) due to abnormal pulmonary vascular dilation, gas exchange disorder, and abnormal arterial oxygenation. abnormal arterial oxygenation due to a gas exchange disorder may increase the alveolar-arterial oxygen pressure difference. hypoxemia is an important pathophysiological basis of hps, and hps is a severe pulmonary complication of end-stage liver disease that is clinically characterized by dyspnea and cyanosis. hps was first reported by rydell hoffbauer in , but it wasn't until that kenned and knudson proposed the full concept of hps. hps per se refers to pulmonary vascular dilation and the shunting of venous blood with low oxygenation to arteries in the presence of severe liver disease. hps is mainly identified in patients with clf (child c hepatic cirrhosis). in addition, patients with either acute or chronic liver disease may present with a pulmonary vascular abnormality and arterial hypoxemia. hps occurs most commonly in patients with hepatic cirrhosis secondary to chronic liver disease, including hepatitis-induced cirrhosis, cryptogenic cirrhosis, alcoholic cirrhosis, and primary biliary cirrhosis, all of which have similar pathophysiological processes as hps. in hps, severe ascites, portal hypertension, and arterial hypoxemia (pao < kpa) may be related to the intrapulmonary vascular shunt, excessive production of nitric oxide, lung ventilation-perfusion imbalance, and interstitial fibrosis. the incidence of hps varies among studies. the incidence of hps is about - % in chronic liver disease patients but higher in patients with hepatic cirrhosis. the most common clinical manifestations of hps are dyspnea, hypoxemia, and cyanosis caused by intrapulmonary vascular dilation and poor arterial oxygenation in the presence of primary liver disease: patients usually progressively develop respiratory manifestations (e.g., cyanosis, dyspnea, clubbed-fingers/toes, orthostatic hypoxia, supine breathing). progressive dyspnea is the most common pulmonary symptom of hps, and cyanosis is a unique and reliable clinical sign. supine breathing and orthostatic hypoxia are characteristic manifestations of hps. pulmonary examination often fails to identify clinically important signs, and hps is not associated with the either the cause or severity of liver disease. in a fraction of patients with stable liver disease, there is progressive lung dysfunction. research shows that hps is associated with esophageal varices and spider angiomas. intrapulmonary vascular dilation (i.e., pulmonary spider angiomas) is frequently found in liver disease patients with subcutaneous spider angiomas susceptible to hypoxemia. spider angiomas have been regarded as a marker of extrahepatic involvement. if patients have no primary heart and lung disease, concomitant lung disease (such as chronic bronchitis, emphysema, pneumonia, and pleural effusion) may coexist with hps. affected patients usually have obvious respiratory symptoms; therefore, physicians should differentiate between the conditions. hps is an independent risk factor for prognosis. specifically, studies have reported that the median survival time is . months after the diagnosis of hps. to date, no effective strategies have been developed for the therapy of hps. orthotopic liver transplantation should be performed as early as possible for hps patients. in liver failure, there is massive hepatocyte necrosis that may cause a reduction in glycogenolysis and abnormal gluconeogenesis. thus, patients are vulnerable to hypoglycemia, shock, coma, and impaired glucose tolerance. the synthetic function of the liver is impaired in such patients, and the serum level of cholesterol and triglycerides decreases. serum cholesterol has been used as an indicator for the prediction of prognosis of liver failure patients. the frequent use of diuretics can cause water and electrolyte imbalance, of which hypokalemia and hyponatremia are the most common. such imbalances may also induce he and brain edema. ap is a rare, but severe, complication of liver failure. the inciting cause(s) and pathogenesis of ap in patients with viral hepatitis remain unclear but might be associated with viral infection, biliary tract lesions, drugs (steroids and diuretics), and other factors. the reported incidence is . - %, but one autopsy study shows that the incidence of ap is as high as % in patients with severe hepatitis and hepatic cirrhosis. evidence shows that the incidence of ap is relatively high in patients with advanced liver failure. further, high serum bilirubin, low albumin, and a significant reduction in prothrombin activity may predict a poor prognosis for ap patients with severe hepatitis and a high mortality. two of the following three criteria are required for the diagnosis of ap: ( ) patients have abdominal pain characteristic of ap; ( ) serum amylase and/or lipase is ≥ × the upper limit of normal; and ( ) there are characteristics of ap on medical imaging. that said, in cases of severe hepatitis with concomitant ap, the symptoms of ap are usually atypical, diverse, and easy to be masked by symptoms of severe hepatitis. thus, severe hepatitis is often considered the cause of abdominal distension, nausea, and vomiting, even in the presence of ap. in some patients, ap may be misdiagnosed as spontaneous bacterial peritonitis, cholecystitis, or gastritis, which may delay treatment and therefore worsen the patient's condition. the clinical manifestations of ap are usually atypical in patients with preexisting liver failure, therefore, physicians should highlight the diagnosis of ap in affected patients. when the following findings are observed, ap should be suspected and laboratory and imaging examinations should be performed as soon as possible for the confirmed diagnosis: . patients with severe hepatitis develop abrupt and persistent upper abdominal pain/peritoneal irritation that is nonresponsive to general antispasmodics; . patients present with severe vomiting, severe sialorrhea of unknown cause, and refractory hiccups; . patients manifest repeated and transient episodes of conscious disturbance, which are refractory and not caused by hepatic coma and hypoglycemia-like reaction; . patients have prior chronic cholecystitis or gallstones, receive treatment with diuretics or steroids, and have symptoms and signs described in ( ) after exclusion of spontaneous peritonitis. for patients with severe hepatitis, routine blood testing and urine amylase detection should be performed dynamically. imaging examinations can be performed simultaneously. abdominal ultrasonography may be performed within - h after the onset of abdominal pain, which is helpful for the morphological change in the pancreas and the exclusion of biliary tract disease. however, gas in the gastrointestinal tract during ap may affect the performance of ultrasonography and make accurate diagnosis of ap impossible. thus, computed tomography is recommended as a standard imaging examination for the diagnosis of ap. computed tomography is helpful for the early diagnosis and subsequently timely therapy, which may improve the prognosis. to facilitate the determination of therapeutic efficacy and the evaluation of prognosis, the branch of infectious and parasitic diseases and branch of hepatology of chinese medical association published the guideline for the prevention and therapy of viral hepatitis in (xi'an conference). on the basis of those guidelines, severe hepatitis can be classified as early, intermediate, and advanced severe hepatitis. specifically, early severe hepatitis meets the diagnostic criteria for severe hepatitis (i.e., severe fatigue, gastrointestinal symptoms, deepening jaundice, serum bilirubin > × the upper limit of normal, prothrombin activation of ≤ - %, or pathological characteristics), but patients have no evidence of he and no ascites. intermediate severe hepatitis patients have grade he or obvious ascites, bleeding tendency (i.e., bleeding point, ecchymosis, and a prothrombin activation of ≤ - %). advanced severe hepatitis patients develop refractory complications and hrs, gastrointestinal bleeding, severe bleeding tendency (i.e., ecchymosis at the injection site), severe infection, refractory electrolyte imbalance, he >grade brain edema, or a prothrombin activation of ≤ %. currently, some investigators classify the natural history of liver failure into the following: prejaundice stage, bilirubin increase stage, bilirubin plateau stage, and bilirubin reduction stage. those stages are based on disease progression, serum bilirubin level, and recovery of liver failure patients. in the prejaundice stage, patients have fatigue, anorexia, and an intolerance of oil. they deteriorate gradually, the urine becomes yellow, liver function detection usually shows a significant increase in aspartate aminotransferase and alanine aminotransferase (higher than several thousand), and the prothrombin activity increases. serum bilirubin increases progressively (i.e., a daily increment of > . μmol/l), and symptoms (fatigue, anorexia) deteriorate after the appearance of jaundice (which is different than manifestations of acute jaundice hepatitis). when the serum bilirubin peaks and remains relatively stable, the disease may be in the bilirubin plateau stage in some patients with no severe complications but present improved mental status and appetite. with the regeneration of hepatocytes, the disease progresses into the bilirubin reduction stage, in which the coagulation, mental status, and appetite improve. when the disease recommences its deterioration, it may progress from the so-called bilirubin increase stage directly to the end stage. in patients with alf, the bilirubin plateau stage is not obvious, and patients might die shortly after disease onset. if patients survive alf, the disease may be pathologically classified as a hepatocyte edema type, and liver function will improve in a short period. not all types of liver failure (including severe hepatitis b) have clear stages based on their natural history and characteristics, and the respective features are discussed in detail as described in the following sections. there is still no consensus on the definition of alf. in , the us acute liver failure study group published guidelines for the management of acute liver failure. in those guidelines, they emphasized that liver failure within weeks after onset can be diagnosed with alf in mother to child transmission of hepatitis b infection (or autoimmune hepatitis), although it has the possibility of progressing into hepatic hepatitis. in addition, some physicians propose that liver failure with an abrupt attack either secondary to chronic hepatitis b or in the presence of other hepatitis virus infection can also be classified as alf. the pathological basis of alf may be classified as necrosis-and degeneration-dominant (acute edema) type. in alf of the necrosis-dominant type, hepatocytes become diffuse and massive necrosis occurs soon after disease onset. in alf of the degeneration-dominant type, hepatocytes show diffuse and severe swelling. alf secondary to acute hepatitis b virus (hbv) infection is rare in clinical practice. patients with alf secondary to acute hbv infection usually have no history of hbv infection, are relatively young, and often have predisposing factors (e.g., stress, absence of rest after disease onset, malnutrition, alcoholism, use of liver damaging drugs, pregnancy, concomitant infection). moreover, it usually progresses rapidly, and patients may develop coagulation dysfunction before the jaundice becomes evident. such patients present with symptoms of liver failure characterized by he >grade within weeks, a prothrombin activation ≤ %, an obvious bleeding tendency (i.e., massive petechiae at an injection site), patients have no ascites, disease progresses rapidly and has a poor prognosis, and patients frequently die of complications such as brain edema or cerebral hernia within weeks. some patients may recover rapidly after appropriate therapy and are usually diagnosed with liver failure of extensive hepatocyte swelling. after recovery, the risk for hepatic cirrhosis is relatively low. another situation is the presence of a history of hbv infection in which patients have a good liver condition and no evidence of/mild liver lesions. for hbv patients with alf, the liver condition is good (as in alf patients without prior hbv infection) and both alf patients with and without prior hbv infection share pathological basis, pattern of disease onset, and clinical course. alf usually progresses rapidly, and the four stages of alf (i.e., prejaundice stage, bilirubin increase stage, bilirubin plateau stage, and bilirubin reduction stage) are difficult to identify. alf may result in high mortality, and a majority of patients directly develop alf of the bilirubin increase stage or even terminal stage. pathologically, slf not only has extensive hepatocyte necrosis but also an obvious inflammatory reaction and formation of regenerative nodules in residual hepatocytes. slf usually has an origin of alf. when slf occurs in patients with or without mild liver lesions, it often shows an abrupt onset. in the early stages, slf is similar to acute icteric hepatitis and patients progressively deteriorate. affected individuals may also develop clinical symptoms of liver failure from days to weeks, including severe fatigue, loss of appetite, frequent vomiting, and deepening jaundice (i.e., a daily increment of > . μmol/l or > mg/dl and an increase in serum bilirubin of > μmol/l or mg/dl). patients usually have hepatic foetor, refractory abdominal distension, ascites (susceptible to concomitant peritonitis), evident bleeding tendencies, and mental and neurological symptoms. in the late stages, hepatorenal syndrome may be present and patients often develop complications (such as gastrointestinal bleeding and hepatic coma) before death. the liver either shrinks or remains normal in size. the course of slf lasts for several weeks to several months. patients surviving slf following therapy usually develop postnecrotic hepatic cirrhosis. clinically, slf can be divided into two types. first, the ascites type results in profound jaundice (serum bilirubin of ≥ μmol/l or > × the upper limit of normal), ascites, and evident bleeding tendencies (i.e., a pta ≤ %). he might be absent or present in the late stages. patients often die of hrs, upper gastrointestinal bleeding, severe secondary infection, and intracranial hemorrhage. slf of the ascites type accounts for a majority of slf. second is the encephalopathy type. such patients have he as an initial symptom and present manifestations as in ash except for course of disease lasting for > days. patients usually die from either brain edema or cerebral hernia. slf of the encephalopathy type is also not rare. slf often has an abrupt onset, and the four stages (i.e., the prejaundice, bilirubin increase, bilirubin plateau, and bilirubin reduction stage) of liver failure are difficult to identify. it is usually associated with a high mortality rate. the pathological basis of aclf is similar to that of slf; therefore, they both share clinical characteristics. a majority of patients with aclf have ascites, spontaneous peritonitis, and biliary tract infection. in the late stages, patients may develop portal hypertension and other complications, repetitive he and hrs, and most die of gastrointestinal bleeding and hrs. according to the guideline for the prevention and therapy of viral hepatitis ( ), a fraction of patients with csh meeting the diagnostic criteria can be grouped with aclf. that is, patients have either chronic hepatitis or compensated hepatitis cirrhosis that remain stable, but some predisposing factors cause the deterioration of liver function, which, thereafter, progresses to liver failure. aclf refers to acute decompensated liver function in the presence of chronic liver disease. the previously mentioned guidelines emphasize pre-existing chronic liver disease and liver failure due to acute liver dysfunction. it is important to note that controversy regarding the basis of chronic liver disease persists. in , an english physician proposed that aclf was diagnosed in chronic liver disease patients with compensated liver function presenting with acute aggravation of liver function within - weeks due to accidents characterized by jaundice, he, and/or hrs. german physicians subsequently proposed that the diagnostic criteria for aclf included ( ) the liver has the histological, laboratory, or ultrasound evidence of hepatic cirrhosis; and ( ) patients develop jaundice, ascites, coagulation dysfunction, and/or grade - he, meeting the definition of decompensated liver function. the guideline for the diagnosis and therapy of liver failure ( ) does not detail chronic liver diseases as a basis of liver failure. however, in general, hbv carrier status may not serve as a baseline liver disease for patients with either chronic hepatitis or hepatic cirrhosis. the term aclf also highlights that acute or subacute deterioration of liver function occurs, which rapidly progresses to liver failure. patients often display an abrupt onset and develop severe fatigue and evident gastrointestinal symptoms. in the early stages, there is acute liver damage; therefore, patients usually present with a significant increase in transaminase levels. thereafter, the disease condition becomes aggravated, and patients may manifest symptoms of liver failure. aclf can also be divided into the brain type and ascites type, of which aclf of the brain type has a higher incidence. further, the four stages (prejaundice, bilirubin increase, bilirubin plateau, and bilirubin reduction) of liver failure are very clear in patients with aclf. one goal for physicians and researchers is to determine individualized therapy for aclf patients according to the specific stage of aclf. patients with clf usually have decompensated hepatic cirrhosis that progressively evolves into chronic liver failure, resulting in clinical manifestations of chronic decompensated liver dysfunction characterized by ascites, portal hypertension, coagulation dysfunction, and he. the pathological basis of chronic liver failure is hepatic cirrhosis, chronic and progressive aggravation of hepatocyte injury, and reduction in hepatocytes that are unable to maintain normal liver function. physical examination usually shows signs of chronic liver diseases (such as liver palms and spider angiomas), imaging examination shows characteristics of chronic liver diseases (such as spleen thickening), and laboratory examination also supports the diagnosis of chronic liver diseases (increased gamma-globulin and reduced/inverted albumin/globulin ratio). of note, a majority of patients have no clear history of liver disease and may initially be misdiagnosed with alf. further examinations may provide evidence of hepatic cirrhosis. when patients with hepatic cirrhosis become decompensated, the liver dysfunction usually presents with acute deterioration due to complications or gradually aggravates in a small fraction of patients. on the basis of the above findings, liver failure secondary to decompensated hepatic cirrhosis can be divided into slowly progressive liver failure and acutely deteriorating liver failure. the former shows a chronic status of liver failure and is characterized by repetitive ascites and he. the latter shows an acute deterioration of liver function in the presence of chronic liver dysfunction, which is similar to aclf in the disease onset and clinical course. hepatopulmonary syndrome is often noted in acutely deteriorated liver failure, and patients usually die of heavy gastrointestinal bleeding, hrs, and severe infection. clf is generally characterized by slow progression of liver failure, and the course of clf is relatively long. the four stages (prejaundice, bilirubin increase, bilirubin plateau, and bilirubin reduction) of liver failure are difficult to identify. as such, finding ways to best preserve residual hepatic function reserve is one of the important therapeutic goals in affected individuals. yu-ming wang, deng-ming he liver failure is a clinical syndrome with high mortality by severe liver damage. it is caused by a variety of causes, results in serious obstacles or decompensation of liver synthesis, detoxication, excretion and biotransformation and appears with coagulation disorders, jaundice, hepatic encephalopathy and ascites as main manifestation. hepatic failure can be divided into acute liver failure (alf), subacute liver failure (salf), acute-on-chronic liver failure (aclf), and chronic liver failure (clf). although the incidence of liver failure is not high in western countries, the relevant papers, reviews, conferences and other exchanges have increased markedly in recent years. aasld, easl and apasl had established a thematic seminar and the definition diagnosis and classification of liver failure has been consistent. at the same time, there are different understandings. therefore, it is necessary to discuss the main differences of liver failure diagnosis and classification, so as to develop a more rational diagnosis and classification scheme. the classification of liver failure involves classification of hepatic injury. a variety of factors (drugs, virus, alcohol, etc.) can cause liver cell damage. although course and prognosis of liver cell damage are different, the most common mechanism is inflammation. wieland et al. found that there were two mechanisms of liver cell injury in the immune clearance of hbv; non-soluble cell damage occurring early and soluble cell damage, early mainly non-soluble cell injury, by the study of gorillas with hbv infected. in , bonino et al. proposed the theory of nonsoluble liver cell damage in the study of fibrosing cholestatic hepatitis (fch) study. however, this theory was ignored because many scholars believed that it ignored the background of immunosuppression. at that time, fch was still considered as the injury of endoplasmic reticulum and golgi apparatus by the excess replication of hbv as well as overexpression of hbv antigen during immune inhibition. however, in , masayoshi et al. reported that most effective antibodies had been detected in children after living donor liver transplantation who received chickenpox vaccine, attenuated vaccines in children, such as measles, rubella, and mumps. according to this, immune suppression cannot stop antibody production and in fch, non-cellular immune injury may be present. recently, we found that there were two types of hbv reactivation in immune-suppressed; high alt type (> × uln) and low alt type (< × uln) and both with bad prognosis. we proposed that there may be different injury mechanisms, both immune and non-immune damage. in , rolando et al. found that . % of acute liver failure patients with systemic inflammatory response syndrome (sirs), and there was significant correlation between the progress of hepatic encephalopathy and infection and between the degree of hepatic encephalopathy and the occurred rate of infection. infection aggravates degree of illness and fatality rate in liver failure patients. accordingly, we speculate that serious primary liver injury can cause injury to other organs by cytokines, while injury to other organs can aggravate liver injury. corresponding with this, we summarized relevant literature and referred that alf can occur secondary in the development process of multiple organ failure induced by non-primary liver damage. this suggests that this type of liver failure is a special type of liver failure, which is a result of rapid changes in internal environment and inflammatory factors induce liver damage. therefore, liver failure can be divided into one with primary injury and with secondary injury. liver failure mainly caused by non-soluble cell injury is extremely rare; as "paralyzed type", "stunned type" or "edematous type", and with the good prognosis. in , small-for-size syndrome after partial liver transplantation was defined by dahm et al., which is a hepatic failure due to less of liver tissue. actually, this syndrome can also be seen as a special type of liver failure caused by non-soluble liver cell damage. soluble cell injury of liver is relatively common, such as the early hepatic failure in hepatocellular carcinoma after interventional therapy, some drug-induced alf, etc. liver failure caused by immune injury is more common in liver failure caused by autoimmune liver disease. liver failure caused by non-immune injury is more common in liver failure patients with severe cellular immunity damaged (hiv/ aids patients, chemotherapy patients) or immunity inhibited (in patients after organ transplantation). primary type is common in fulminant hepatic failure (fhf), the secondary type is seen in severe systemic diseases, such as severe sepsis and acute hemorrhagic. in fact, clinical liver failure is the result of combination of different proportions of various types of damage factors. based on the role of various factors in liver failure, it can be categorized. according to acute and sustained, the clinical course and the image changes, liver failure can be divided into different stages with some or certain factors dominated. we have divided it into two categories, necrosis type and the decompensation type. although this classification is based on practice and clinical management, large-sample analysis is needed in the future. no matter what the cause, liver failure may be divided into two major categories on the pathophysiology. one type is necrosis induced by hepatic inflammation; the other type is decompensation of liver cells. in particular, alf and salf are types of necrosis, aclf and clf are decompensated type. mixed type, both necrosis type and decompensation type, is possible, which treatment should be considered. the relevance of these two type of liver failure mainly reflects in treatment. necrosis type mainly focuses on treatment of the cause (as antiviral treatment for hbv infection and corticosteroid treatment for autoimmune hepatitis) and symptomatic support treatment (as anti-inflammatory treatment and integrated symptomatic support treatment). decompensation type mainly focuses on intensive treatment (as control infection for sirs and control bleeding for gastrointestinal hemorrhage). some pathophysiological processes such as hepatic encephalopathy (he) are in both type, but are different in different liver failure type. for example, cerebral edema is more prominent and progressive and less to do with high protein diet in he patient of necrosis type of liver failure, while is just the opposite in decompensation of hepatic failure. based on pathologic features and speed of progression, liver failure can be divided into four categories: alf, salf, aclf, and clf (tables . and . ). alf occurs liver failure syndrome characterized with varying degrees of hepatic encephalopathy ( ): - (article in chinese) [ ] in weeks except for liver cirrhosis. salf occurs liver failure syndrome in days to weeks. aclf is acute hepatic decompensation on the basis of chronic liver disease. clf refers to chronic hepatic decompensation characterized by ascites, portal hypertension, coagulation disorders or hepatic encephalopathy based on cirrhosis. diagnosis and classification of liver failure are the most controversial part, but has tended to unify in recent years. in , professor roger william from the university of london proposed the same type criteria as chinese. the only difference is limited to weeks for alf and no aslf. due to aslf belonging to alf, this part is not contradictory for two criteria. in recent years, a discussion of aclf proposed many times by sarin from india made aclf more valued and accepted. clf is also getting more recognition. recently, to clarify meaning and avoid misunderstandings, it was recommended that clf be converted into end-stage liver failure (eslf). although issues related to classification of liver failure have largely agreed, there are some differences in practical application. dispute over whether to set up subtypes (namely aclf/saclf) of aclf occurred during drafting the new version of guide. two suggestions of modification are: ( ) with existence of cld, clinical manifestation of acute (within weeks) and sub-acute ( ~ weeks) liver function decompensation occur; ( ) with existence of cld, clinical manifestation of acute (within weeks) liver function decompensation occur (this actually is the original version). the reasons are: ( ) the classification of guide has only been published for years, and it was not easy for it to be widely recognized home and abroad. it requires more time to accumulate experience in this field, so frequent revision are inappropriate; ( ) as is generally accepted home and abroad, aclf mostly occurs within month ( weeks) after onset, while those as late as weeks after onset are rare (the document of our department indicates the same result); ( ) it still requires medical evidence and extensive clinical summary and proof to have the new diagnostic term "saclf" in english and chinese established and accepted; ( ) clinical significance and importance of salf classification are neither prominent nor urgent. after multiple discussions, diagnostic classification in the guide applied the latter one (table . ). despite the differences, academia has become unified about the classification of liver failure in the world. the differences towards the convergence of: ( ) in terms of naming and classification. naming has simply become to acute liver failure (including acute and subacute) and clf (include acute-on-chronic and chronic decompensation), and tend to be more simplified. aasld guidelines clearly stated that nouns used to differentiate the length of the course (such as hyper acute, acute, and subacute) had claimed not to use. ( ) in terms of clinical diagnosis. because of many cause of liver failure, it is very difficult to achieve unity. the only way is to combine the clinical diagnosis (such as acute hepatitis) and the pathophysiologic diagnosis (such as alf). ( ) if hepatic encephalopathy as a prerequisite for liver failure. currently, it is a prerequisite for alf, and not necessary for clf because hepatic decompensation is the main clinical manifestations. according to the severity of the clinical manifestations, liver failure can be divided into early, middle and late stage. . extreme weakness, severe gastrointestinal symptoms such as significant loss of appetite, vomiting and abdominal distension; . progressive jaundice (serum total bilirubin ≥ μ mol/l or increased by ≥ . μ mol/l daily); . bleeding tendency, % < prothrombin activity (pta) ≤ % (or . < inr ≤ . ); . no hepatic encephalopathy or other complications. based on the early stage of liver failure, further develop to one of the following two: . below grade ii hepatic encephalopathy and/or obvious ascites and infection. . obvious bleeding tendency (bleeding point or ecchymoses), and % < pta ≤ % (or . < inr ≤ . ). based on the middle stage of liver failure, further aggravating, severe bleeding tendency (such as ecchymoses on injection site), pta ≤ % (or inr ≥ . ), achieve one of the following four: hepatorenal syndrome, upper gastrointestinal bleeding, severe infection and above ii° hepatic encephalopathy. considering the notorious difficulty to treat hepatic failure and its high mortality rate, special attention has to be paid to and active treatment has to be performed on patients showing the following early-stage clinical features of hepatic failure. . extreme weakness, severe gastrointestinal symptoms such as significant loss of appetite, vomiting and abdominal distension. . progressive jaundice ( μ mol/l ≤ t.bil ≤ μ mol/l), and increased by ≥ . μ mol/l daily; . bleeding tendency, % < prothrombin activity (pta) ≤ % (or . < inr ≤ . ). clinical practice has shown that, with or without history of chronic liver disease, there are patients with grade ii hepatic encephalopathy in a short period, with rapid development, poor prognosis. these patients should be regarded as fulminant type. meanwhile, in asia, including china, there are some patients with severe jaundice, ascites and bleeding as the main presentation, with relatively slow development and very poor prognosis, but without hepatic encephalopathy. these patients should be classified as subacute type. fulminant type must have a hepatic encephalopathy. however, it is not necessary for subacute type, which mainly characterized by severe jaundice and ascites. compared with severe hepatitis in china, fulminant type amounts to acute severe hepatitis and chronic severe hepatitis with acute onset, subacute type amounts to subacute severe hepatitis and chronic severe hepatitis with subacute onset. currently, a large divergence of these two types is about time, from days to weeks. according to clinical features of alf, alf can be further divided into fulminant type and subacute type interval for weeks. however, according to more researches, fulminant hepatitis, characterized by massive necrosis of liver, brain edema, and hepatic encephalopathy, concentrated in weeks, most of them in days or less. taking into account the subacute type belongs to acute category, subacute are not be established in international classification. alf are defined as liver failure in weeks. on the difference between acute and chronic liver failure, most chinese scholars depend on the past history, which be ignored internationally. the difference lies in this onset. acute inflammation, necrosis and chronic decompensated were classified as acute and chronic processes, respectively. most typical example is that patients with acute heavy syndrome of onset in hbv carrier were divided into alf by the scholars from hong kong, macao and taiwan. similarly, liver failure caused by hepatitis flares in chronic hbv carriers, reactivation of chronic hepatitis b, super infection with hdv and hbeag seroconversion are included in alf. it is inconsistent with classification methods in china. the reason lies in greater emphasis on the continuous development processes of hepatitis chronicity and severity in china, and focused on the acute effects this time abroad. some scholars suggest that considering the significant difference between clf and the other three types in clinical manifestation, it is worth discussing whether to list clf as a type of hepatic failure. we believe that significance and importance of clf classification are: ( ) clf are similar to crf (uremia) in nephrology and chronic cardiac failure (congestive heart-failure) in cardiology. although their clinical manifestation differ significantly, the "coexistence of acute and chronic failures" is shared by failures of all those organs; ( ) clf classification has been generally recognized at home and abroad, and the necessity of classification are further proved by the difference between clf and the other three types; ( ) clf cases are relatively large in proportion (nearly %), which is still increasing (since the proportion of alf/salf are lowering); ( ) complications of clf are common and are found in various forms, with bad prognosis; ( ) in clf patients with correlation to hbv, virus replication are commonly found, which is closely related to decompensation. the efficacy of nucs are satisfying, which, if taken for a long term, can reverse decompensation, avoiding liver transplantation; it also increases support means in a fast rate, creating more chances for treatment. if the strict definition of acute and subacute liver failure as "no past history of liver disease" is executed, how to name the patients who had a history of chronic liver disease (caused by hbv from mother to child transmission in china)? as for alf and salf, rigorous definition for the past history of liver disease (including hbv carrying history) is necessary in china, and more interested is in this attack instead of the latent infection in the past, even a dominant attack in europe and america. in clinical practice, past history should not be ignored, because patterns of chronic hepatitis b reactivation vary. we summarize them into four types: ( ) burst type: suddenly attack based on immune tolerance state, eventual liver failure; ( ) recurrent type: repeated unequal flares, finally developing into liver failure; ( ) occult type: no obvious attack, presenting with symptoms of decompensation; ( ) document type: compensated cirrhosis, acute decompensation in certain situations (mainly due to sepsis and other infections). in burst type absence of history, or only carrier state, the past history can be ignored. in recurrent type, history of recurrent injury is important, which the significance lies in the extent of the occurrence, duration and consequences. because these factors determine the basis of injury to the patient's liver, the patients with mild liver disease have mild or even have no hepatic fibrosis and liver cirrhosis, otherwise, the symptoms will be serious and obvious. the former attack often leads to liver necrosis, the latter often lead to decompensation. the difference between occult type and document type is in the speed of decompensation; the former is slow and the latter is fast. in summary, although history has the certain reference value, pathophysiological changes in attack is main of necrosis or decompensation, or a combination of both. whether hepatic encephalopathy should be considered as a complication of liver failure is controversial, because many scholars have listed it as a prerequisite for liver failure, but in recent years, some patients do not necessarily have encephalopathy. from the complete course and early prevention and treatment of liver failure, it is necessary to incorporate non-encephalopathy type, but the effect and prognosis of the rescue treatment should be divided into the encephalopathy type and nonencephalopathy type, because they are different. hepatic encephalopathy is divided into a, b, c type in international guidelines. type a is acute hepatic encephalopathy (alfa-he), which does not include acute hepatic encephalopathy associated with chronic liver disease. some patients with a long-term hbv carrier were diagnosed with aclf or clf on the first time severe, especially in china. in fact, this type of patients with acute or subacute liver necrosis caused by alf. the mechanism of hepatic encephalopathy in this type liver failure is different from alfa-he, as well as treatment. liver failure classification has the greatest impact on treatment of hepatic encephalopathy, based on the following facts: ( ) in acute phase of alf, fasting protein diet on the first day, unnecessary in the short term ( days); however, chronic hepatic encephalopathy of clf don't have to fast; ( ) the metabolism of branched chain amino acids (bcaa) in alf was reduced and increased in clf. this suggests that the former should not be added bcaa, and the latter can supplement the bcaa. ( ) high blood ammonia in clf is more than in alf. the effect of clf was better than that of alf patients, but the effect was not good for deamination drugs. ( ) because cerebral edema in alf is more than in clf, it is better to reduce the intracranial pressure in alf treatment, and in clf with poor efficacy; ( ) as for type a and c according to the international consensus of hepatic encephalopathy are equivalent to the current alf and clf. severe cerebral edema has been found in alf and its mechanism is not clear. study on cerebral edema treatment have been found that hypothermia therapy can reduce the cerebral blood flow and brain edema. in th easl, larsen from affiliated hospital of university of copenhagen in denmark reported that therapeutic hypothermia did not support application in the treatment of patients with alf in a prospective, multicenter randomized controlled experimental study (the study was carried out in - ). the study results are different from previous studies. we believe that the reason is likely from the bias of group selection. in addition, there is a great difference of the mechanism of hepatic encephalopathy in patients with chronic hepatic failure and alf, so the response to hypothermia therapy will vary greatly. based on the existing research, hypothermia therapy has better effect on hepatic encephalopathy in alf patients caused by cerebral edema, and has bad effect on hepatic encephalopathy in the chronic decompensated liver failure caused by metabolic abnormalities. as for hepatic encephalopathy in aclf patients, the effect depends on the roles of cerebral edema in pathogenesis. therefore, it is recommended that the patients should be carefully screened to obtain a comparable result in the study of the hypothermia therapy in hepatic encephalopathy. we believe that, with the gradual elucidation of the pathogenesis of liver failure, the treatment measures will also be more targeted, the efficiency may be further improved. glucocorticoids (gcs) therapy in chronic active hepatitis b began in the s to s. however, compared with the control group, gcs did not show better effect. there have been reports that, after stopping the treatment of immunosuppressive therapy, early re-given long-term high-dose gcs can prevent severe hepatitis in hbv reactivation patients. however, this result had not been affirmed in the future clinical practice. although more application of gcs before s, each effect is different. there is a negative trend in s. we have analyzed, the effect may involve two major factors: one is the choice of indications, dose and duration of treatment; another is the prevention of adverse reactions and complications of gcs. according to the guidelines for diagnosis and treatment of liver failure ( ) in china, liver failure without viral infections, such as autoimmune hepatitis and acute alcoholism (severe alcoholic liver disease), etc. are gcs indications. at the early stage of liver failure caused by other reasons, in the patients with developed rapidly and no serious infection, bleeding and other complications, gcs may be appropriate to use. gcs can improve the survival rate of patients with autoimmune hepatitis and severe alcoholic hepatitis, and has been recognized by most scholars. for hepatitis flares in chb patients, if on the basis of combined application of nucs, gcs will inhibit excessive hyperactivity of host cellular immunity and excessive release of cytokines, and help preventing liver cell death. at the same time, the role of gcs in the treatment of chb and the specific usage, as well as the effect of combined treatment is still controversial. the reason is that the advantages and disadvantages of gcs are very prominent. the key of success or failure lies in the clinical skills. in recent years, the reports of gcs for the treatment of hbv related liver failure increased. there are three main reasons: ( ) nucs can effectively resist hbv replication due to gcs; ( ) application of proton pump inhibitors can effectively prevent gastrointestinal bleeding due to gcs; ( ) increasing of infection prevention and treatment can effectively fight infection due to gcs. even so, the above three aspects of the problem have not been satisfactorily resolved. therefore, we put forward several viewpoints on the current application of gcs: first, to fully analyze the advantages and disadvantages, to consider the main function and purpose after the gcs application and the risk of adverse events before expanding the indications of gcs. secondly, both short course treatment ( - days) and long course treatment has drawbacks. the former may not be sufficient to adequately inhibit a strong immune response, and induce a stronger immune response after a sudden stop; the latter can induce bleeding, infection or viral resistance. finally, in a common clinical liver failure induced by hbv reactivation under immunosuppression, the mechanism is often unrelated with immune activation. the typical representative is fch, and the prognosis is extremely bad. it should be vigilant, focus on prevention. our department has treated a chronic hepatitis c (chc) patients after kidney transplant and taking large doses of gcs. severe fch was induced, resulting in liver failure and eventually died. in recent years, the hepatic stem cell therapy of liver failure was concerned, human stem cell transplantation in the treatment of clinical study on severe hepatitis/liver failure were carried out and the results were satisfactory. however, due to the difference of the patient's condition or stage, and most of which are case report, it is difficult to draw an objective conclusion. we have repeatedly reported that the human umbilical cord blood stem cells and bone marrow stem cells in vitro and in vivo can be successfully transformed into liver cells, but the biggest obstacle to its application to clinical practice is the limited number. it is difficult to repopulate the whole liver or at least part of liver compensatory function. in addition, considering the part liver failure patients with cirrhosis background, which clinical manifestations as aclf and clf in the practical application, there are the following issues: ( ) it is difficult to provide effective growth and functional support for implanted cells in a diseased liver; ( ) the original portal hypertension can be aggravated by portal vein implanted cells; ( ) input cells by vein can obstruct the pulmonary vein, thus affect the pulmonary circulation; ( ) if the implanted cells for allogeneic or xenogeneic tissue, may have compatibility problems; ( ) when using gcs to prevent rejection, severe hepatitis patients are easy to infection and other related adverse reactions. finally, it is pointed out that there is a tendency of the academic circles to the understanding of liver failure for decades, that is, the existence of the chronic process is often neglected in the process of analysis, and vice versa. a typical example, until nearly a few years ago, many scholars in the world still do not recognize the existence of aclf and clf. chinese research papers on liver failure published very little in the international, the main reason is that the aclf and clf is not clearly defined. the papers are often rejected because the diagnosis of type does not conform to the international. in contrast, as previously mentioned in the definition of aki and hrs, the existence of alf (including salf) is ignored. another example: in the report of antiviral treatment of hbv related liver failure, some foreign scholars put forward that the liver failure should be changed to the decompensated liver cirrhosis, the reason is that acute hbv infection is self-limited. however, at present, there has been a clinical recognition that the hbv related alf is actually an acute episode of chronic carrying process and should be referred to as aclf. in chinese guidelines for the diagnosis and treatment of liver failure ( ), liver failure was divided into four types, mainly because of their different mechanisms and treatment (sometimes even the opposite). due to limitation of the space and data, we have not discussed infection, bleeding and other complications in relation to liver failure. these pathological processes are closely related to the classification, and should be studied further in relation to liver failure. in future, we should develop classification of liver failure according to the mechanism of liver injury with different causes, and provide the basis for clinical types of hepatic failure. in order to improve the level of diagnosis and treatment of liver failure, the mechanism based classification should be carefully assessed and evaluated. standard' of definite diagnosis, severity evaluation and treatment effect assessment, and it is irreplaceable compared with other examinations. histopathological evaluation of aechb and liver failure not only contribute to the definite diagnosis of severity of aechb and provide pathological evidence for effective clinical treatment of hepatitis b, but also is helpful to the early detection of histopathological proof of aechb by pathological examinations and of pre-warning function for clinical treatment of aechb. this section mainly focuses on pathological features of aechb and other types of liver failure. aechb, clinicopathological manifestation of aggravation of liver necroinflammation and disease deterioration of chronic hepatitis b, tends to be of poor prognosis without positive and effective intervention. its pathological characteristics mainly include: ballooning degeneration of diffuse hepatocytes, significantly increased focal necrosis, confluent necrosis, bridging necrosis, extensive and intensive interface hepatitis, massive or submassive necrosis, many neutrophil infiltrations in hepatic lobules and portal areas and moderate intrahepatic cholestasis. prominent hepatocyte necrosis is the pathological foundation of aechb and manifests extensive multifocal necrosis, confluent necrosis, bridging necrosis and other forms of necrocytosis. severe ones can even occur submassive and massive necrosis leading to the extreme form of aechb, severe hepatitis (liver failure). ballooning degeneration of diffuse hepatocytes is one of pathological characteristics of aechb ( fig. . ). the hepatocytes manifest sparse and granular cytoplasm, sometimes can be micro-bubble like. the degenerated hepatocyte is - times bigger than the normal hepatocyte. sometimes the ballooning degenerated hepatocytes can fuse and transform into multinucleated cells, and this lesion is similar to that of neonatal giant cell hepatitis when it is relatively extensive. the general performance of the liver is increased volume, tense capsular and cutting edge eversion due to tension. ballooning degeneration of hepatocyte is not specific histological manifestation of hepatitis band also can occur in liver tissues of hepatitis caused by factors such as alcohol or drugs. extensive and diffuse ballooning degeneration of hepatocyte can make the hepatocytic plate wider, and hepatic sinusoid is pressed to be narrower, causing microcirculation disorder of liver tissue and exacerbation of disease. liver lobular inflammation activity is enhanced, and apoptotic bodies and focal necrosis increased significantly when aechb occurs. hepatocyte apoptosis is the programmed necrosis and one of the major forms of hepatocyte death in hbv infection. histopathology manifests cell membrane shrinkage, deepened cytoplasmic staining, eosinophilic degeneration. free apoptotic bodies in liver sinus are large or the apoptotic cell fragments, sometimes containing nuclear fragments. focal necrosis is another form of liver cell necrosis and manifests as an interruption of the hepatocytic cords or replacement by focal lymphocytes and macrophages, with hepatic regeneration that often causes irregular arrangement of hepatocytic cords. this necrosis often inferred from the disappearance of the hepatocytes and the infiltration of inflammatory cells rather than what is actually seen under a microscope. swelling hepatocytes presented increased size and loosing cytoplasm, further develop to ballooning degeneration showed almost spherical in size and transparent cytoplasm, predominantly in the lower right confluent necrosis and bridging necrosis are the common histopathological changes, play important roles in the progression of aechb and are closely related to the adverse prognosis of hepatitis b. due to the larger necrosis range of confluent necrosis and bridging necrosis, even during repair stage after going through the active stage, the liver tissue often undergoes fibrous repair, resulting in liver fibrosis and hepatic lobule reconstruction, thereby causing liver cirrhosis. statistics show that about % of the patients with viral hepatitis who had bridging necrosis progress to cirrhosis. confluent necrosis is regional lytic necrosis of hepatocytes on a larger scale and is common in active stage or aggravation of viral hepatitis, or drug-induced liver injury, which often occurs around the central veins and inflammatory cell infiltration is not obvious. specific confluent necrosis can also be seen in other parts. take ferrous sulfate poisoning for example, confluent necrosis is more common in zone of liver acinus, and when confluent necrosis expands to connecting vascular or portal area, bridging necrosis occurs ( fig. . ). bridging necrosis is large area hepatic lytic necrosis that connect the portal area to central area (p-c), portal area to portal area (p-p), and central area to central area (c-c). it can be caused by the expansion and confluence of interface hepatitis, or a one-time large-scale translocular necrosis. p-c necrosis: it starts on the periphery of the lobules, affects the central hepatic lobules when it expands, and forms bridging necrosis phenomenon. the currently acknowledged mechanism is as follows: the initial pathological change is serious periphery necrosis of hepatic lobules. with the aggravation of the disease, microcirculatory disturbance occurs in the lobules and causes the hypoxia, degeneration and necrosis of central area liver cells. p-p bridging necrosis: most scholars think it is caused by the expansion of interface inflammation, especially based on the fibrous septum, hbv load increasing significantly, immune response enhancing, activating the signal pathway of liver cell death. with the enhancement of the lesion activity, fresh and severe necrosis occurs. c-c bridging necrosis: it is usually seen in serious disease with bridging necrosis of hepatocytes, but mononuclear cells infiltration is rarely seen in the necrosis area and serum transaminase increases significantly (up to iu/l above). histological manifestations of bridging necrosis can vary due to the different stages of the disease. in the early stage, the liver parenchymal cells necrotize and then disappears, reticular framework residue accompanied by the infiltration of lymphocytes and macrophages. with the time extending, reticular framework collapses and forms the sparse interval crossing the liver tissues. when bridging necrosis is accompanied with reticular framework collapse, with hepatic necrosis and regeneration, disorder of hepatic lobules occurs. at this time, it is difficult to distinguish the fibrous septa between bridging necrosis and chronic hepatitis, and elastic fiber staining can help to solve this problem. the elastic fiber staining of bridging necrosis is negative, because elastic fiber formation often takes several months. interface hepatitis, formerly known as piecemeal necrosis, is one of the symbolic histological manifestations in the chronic activity of chronic hepatitis b. it mainly refers to single or small clusters of liver cells around the portal areas necrose and shedding, leading to worm-eaten defect of limiting plates. significant lymphocytes infiltration is commonly seen in and around the portal area. mononuclear cells extend to the hepatic lobules along the destructive limiting plates and encase the necrotic liver cells, resulting in the enlargement of portal areas ( fig. . ) . interface hepatitis increases significantly and extensive interface hepatitis occurs in aechb. the interface hepatitis area can exceed % of portal areas periphery and be more than a third of the depth of hepatic lobules, even causing bridging necrosis (p-p and p-c). because the limiting plate of the lobules is an important structure to maintain a whole hepatic lobules, interface hepatitis destroys the integrity of hepatic lobule structure. extensive and intensive interface hepatitis can often cause bridging necrosis and bridging fibrosis, and is an essential part of poor prognosis of aechb. massive necrosis and sub-massive necrosis are considered to be the basic pathological changes of severe hepatitis (liver failure) and major substratum for histologic diagnosis of liver failure. massive necrosis is the diffuse lytic necrosis of the liver parenchyma involving more than / of hepatic lobules. thorough and rapid liver tissue necrosis is shown with invisible necrosis process, and only reticular framework remained and is filled with red blood cells ( fig. . ) . sub-massive necrosis is diffuse liquefaction necrosis of the liver parenchyma involving / - / of hepatic lobules. reticular framework collapses and forms reticular fiber bundles, residual liver cells and bile ducts proliferate. massive necrosis and sub-massive necrosis will seriously affect the prognosis of patients with high fatality rate once they occur. the cause of massive necrosis and sub-massive necrosis remains unclear, and the possible causes include excessive virus replication, virus mutation, overlapping with other virus infection and microcirculation, etc. when extensive confluent necrosis, massive necrosis and sub-massive necrosis involve the entire hepatic lobule and even several adjacent hepatic lobule, causing a lobular or adjacent several lobular hepatocytes lytic necrosis, then the panacinar or multiacinar necrosis occur, which is the most severe form of necrosis of aechb. in panacinar and multi-acinar necrosis, with large range of liver cell necrosis, only a small amount of liver cells remain. residues of clump, rosetting, island or glands-like arrayed liver cells are commonly seen around the collapsed reticular framework after necrosis or loose fibrous connective tissue. due to distortion, normal structure cannot be recognized and sometimes can only be identified by portal area range around the necrotic area. cells proliferation can be observed in periportal area, arraying like bile duct structure and these cells can express hallmarks of hepatocyte and bile duct epithelial cells at the same time, which is considered to be the histological manifestation of liver stem cells (hepatic stem cell) activation and proliferation. infiltrating inflammatory cell types tend to be multiple, and the quantity varies. when there is less infiltrating inflammatory cells, the main cell type is macrophage, often containing brown pigment particles. notably, the liver biopsy might be error in diagnosing the necrosis of wide range of diffuse dissolved necrosis of liver parenchyma (above / lobule), with only the mesh stents remained the whole lobules and multi-lobules, and that is due to limited amount of liver biopsy specimens. for example, multi-acini necrosis occurs only in the area under the liver capsule, and liver biopsy pathological examination may overestimate the severity of illness. in aechb, types of infiltration inflammatory cells in the liver tissue are various, and most of them contain many neutrophils. it is different from obvious lymphocyte accumulation within liver parenchyma and portal area common seen in hepatitis b. usually, cd + t cells/cytotoxic t lymphocytes (ctls) are the major effector cells of the inflammatory response in hepatitis b. but when exacerbation occurs, the neutrophils in inflammatory cells infiltrating liver acinus and portal area increase significantly. neutrophils play an important role in innate immunity, and are the first inflammatory cells migrating to the lesion during inflammatory response. these neutrophils kill the invading pathogenic microorganisms through releasing protease and anti-microbial proteins, and producing reactive oxygen. meanwhile, neutrophils have important functions in activation and regulation of innate immunity reaction and adaptive immunity reaction and could release cytokines such as il- to participate in regulating adaptive immunity reaction. although cytokines produced by neutrophils are less than mononuclear macrophages, since neutrophils are the first inflammatory cell moving to inflammatory lesion, the immune regulator function of neutrophils is more important during the early or acute stage of immunity reaction. evidently increased neutrophils in liver tissues of aechb might be beneficial for eliminating infected cells, but it is also a "double-edged sword". accumulation of neutrophils may cause extreme immune response and excessive inflammatory reaction might lead to deterioration. therefore, when increasing neutrophils appear in liver tissues of hepatitis b patients, more attention should be paid to identify whether virus mutation, overlap infection, drug-induced liver injury occur and cause aechb. intrahepatic cholestasis (cholestasis), especially moderate or severe cholestasis, is one of the common histologic manifestations of aechb. intrahepatic cholestasis takes shape from the bile thrombus within the cholangioli around the central vein which is hard to be identified, forming cholestasis in expansive interlobular bile duct and large "bile lake" in hepatic tissue ( fig. . ) . microscopically, bile can be characterized by dark brown, green or yellow color, occasionally also can present the gray which is difficult to recognize. bilirubin is revealed as green in van gieson staining, which is helpful to pathological diagnosis of intrahepatic cholestasis. moderate and severe intrahepatic cholestasis often cause feather-like degeneration of hepatocytes, even intrahepatic cholestasis infarction. with the extension of duration of intrahepatic cholestasis, the inter-hepatocyte structure relation of - normal liver cells surrounding the capillary bile duct also changes. the number of liver cells around the bile duct increases, and the bile canaliculi expands, causing the cholestasis related rosette structure forms and the fusing multi-nucleus giant hepatocyte appears. kupffer cells with brown bile pigment can be seen in the hepatic sinusoid. remarkably, although intrahepatic cholestasis is often associated with clinical symptoms such as jaundice and risen serum bilirubin, the severity of intrahepatic cholestasis is not consistent with clinical symptoms and serum bilirubin level. severe hepatitis b (liver failure) is the most severe liver syndrome complex, which is characterized by poor clinical course and high mortality rate. over the years, scholars have continued to explore the definition, classification, diagnosis and treatment of liver failure, but so far no consensus has been reached. according to histopathological features and progression of the disease, china released guidelines on diagnosis and treatment of liver failure in , which divided liver failure into four categories: acute liver failure (alf), sub-acute liver failure (salf), acute-onchronic liver failure (aclf) and chronic liver failure (clf). hbv infection is the most common cause of liver failure in china. acute liver failure is characterized by acute onset, and hepatic encephalopathy (above stage ii) often develops within weeks of the onset, which results in high mortality rate. former knowledge about acute severe hepatitis mostly comes from the autopsy. nowadays, with the development and universal application of biopsy technique, and the further study on acute liver failure, we have better understanding of the development and process of necrotic lesions, and can predict prognosis and outcome according to the necrotic area and type. in alf, liver atrophy is significantly present in gross pathology inspection, especially for the left lobe. coverings shrinkage, thin edge, soft liver texture, section may be yellow or reddish-brown, and some area is red alternating with yellow, and the weight of the liver drops sharply to - g. histopathology emphasizes extensive and consistent liver cell necrosis caused by one-time strike, and most patients die in the short term. the morphology of liver tissues is relatively simple, manifested as massive or sub-massive necrosis of liver cells, dissociation of liver cords and hepatolysis. the regeneration of liver cells is not obvious, and surviving liver cells show clear ballooning degeneration. hepatic sinus expand and, congest with blood and occur hemorrhage. kupffer cell proliferates and sinusoidal mesh stent does not collapse or completely collapse. quantity of liver cell necrosis is closely correlated with prognosis. if the amount of necrosis is over %, mostly the patient will not survive. if the amount is less than %, the patient is expected to resume with rapid regeneration of hepatocytes. if there is diffuse small steatosis, the prognosis is often poor. concerning the hepatic regeneration of acute liver failure, former pathology emphasizes on liver cell and bile duct cell proliferation of sub-acute liver failure, and relatively neglects cell regeneration of acute liver failure. based on the authors' knowledge, in some acute liver failure cases, liver tissue demonstrates obvious bile duct-like or acinar-like regeneration within days after onset. the regenerated liver cells were co-expressing albumin and ck , ck , indicating these cells have double markers of hepatocytes and biliary epithelial cells, which presumably come from liver pluripotent stem cells. the regenerated liver cells in acute liver failure have their unique characteristics that degenerated and regenerated cells co-exist in the liver. as the time of liver biopsy is different, the morphological change is also varied because of the rapid restoration. the usual dual-core, large nuclear or nuclear fission are rarely seen in regenerated liver cells; liver cell body swelling, transparency of cytoplasmic periphery and slightly basophilic center are commonly seen in regenerated liver cells. because of cell enlargement and transparent cytoplasm, it is often difficult to distinguish from serious ballooning degeneration; in some cases, it shows bile granules within the cytoplasm, bile thrombus within bile capillary with cell swelling and transparency symptoms, which is also similar to the feather-like degeneration caused by bile salt siltation. however, unlike feather-like degeneration which was scattered by small groups or disorderly arranged severe ballooning degeneration, cell enlargement often shows the pole adenoid arrangement, which is known as a sign of liver cell proliferation. the outcomes of continuous liver biopsy also prove its rapid regeneration. nayak et al. also proposed that hepatocyte swelling during the acute liver failure recovery stage indicates good prognosis. the continuous liver biopsy of liver transplant centers in europe also confirmed that the appearance of liver cells enlargement after days of partial liver transplant of acute liver failure with massive necrosis, cells linked to sheets, and lobular structure are basically recovered in to months. the vacuolation, cholestasis and duct-like structure presented by these regenerated enlarged liver cells will be gradually disappeared. the onset of salf is acute, and liver failure syndromes appear within days to weeks, mostly caused by delay of acute liver failure. in salf, liver atrophy is mainly presented as in gross inspection, and variable sizes of regenerative nodules, the yellow-green cutting surface due to cholestasis. histopathology manifests the new and old sub-massive necrosis of liver tissues, or bridging necrosis. in older necrosis area, reticular fibers collapse, or collagen deposit. survived liver cells may have varied degrees of regeneration, and are arranged in nodular. fine, small bile duct proliferation and cholestasis are commonly seen in the periphery lobe. the sinusoids congest in the early stage, collapse in mid-stage, and occlude in late stage. the histopathological distinction of salf and alf is based on the consistency of necrotic lesions. alf emphasizes consistency of necrotic lesions, that is, 'onetime strike', while the necrotic lesions of salf are mixed, caused by 'multiple attack'. in addition, differences also exist in aspects such as cell regeneration and extracellular matrix (ecm) expression between alf and salf. liver stem cells play an important role in the liver cell regeneration process of alf. the regenerated liver cells express dual markers of liver cell and bile duct cell, and are often orderly proliferated along mesh stents. whereas salf presents unipolar regeneration of liver cell and bile duct cell, and the regenerated liver cells is disorderly arranged. due to the different length of disease course, there is no obvious ecm deposit in alf, while salf presents in iii collagen-based ecm deposit. this type of liver failure often develops into post-necrotic cirrhosis. acute-on-chronic liver failure (aclf) refers to acute liver function decompensation occurring on the basis of chronic liver disease. liver gross manifestation of acute-on-chronic liver failure differs with the different stages of chronic liver disease. for instance, aclf occurring in the stage of cirrhosis is accompanied by hepatic cirrhosis nodules besides liver atrophy. the main histology of aclf is new and varied degrees of liver cells necrosis, hepatocyte focal and spotty necrosis, bridging necrosis, confluent necrosis, massive necrosis and sub-massive necrosis on the basis of chronic liver injury. the common chronic changes are as follows: fibrosis in collapsed reticular framework or periportal area with obvious extracellular matrix deposit, forming of large number of fibrous septa, sparse scars, or bridging fibrotic septa when distortion of lobule structures associated with disproportionate numbers of central veins and portal area, pseudolobule formed; twin-cell or multiple-cell of liver plate is commonly seen and the liver plate lose the radiated array, activated regeneration of liver cell caused the occurrence of tumor-like cell. chronic liver failure is chronic liver function decompensation caused by progressive deterioration of liver function on the basis of liver cirrhosis, with ascites, portal hypertension, blood coagulation dysfunction and hepatic encephalopathy as main symptoms. liver gross appearance of clf is significant liver atrophy and nodular liver cirrhosis. histopathological changes are mainly those of liver cirrhosis, including diffuse liver fibrosis, nodular liver cirrhosis with unevenly distributed liver cells necrosis. progression of hepatitis b is an interaction between hbv infection and body response. development of aechb is mainly caused by obvious increased viral load and/or decreased immune clearance. large number of hbv replication can activate hepatocyte death pathways, leading to serious liver inflammation, necrosis and aggregation of disease. additionally, hbv infection overlapping with hcv/hiv, or with etiological factors like drugs and ethanol, could also affect disease progression. fibrosing cholestatic hepatitis (fch), a new clinicopathological type, develops in stages of severe immunosuppression caused by various reasons, especially in hepatitis virus-infected patients lots of immunosuppressant after organ transplant. due to immunosuppressor, hbv replicates rapidly in the patients, leading to quick progression of hepatitis and progressive failure of liver function. the histopathological features of fch are as follows: fibrosis straps starching from the portal area to hepatic sinusoid and circumvoluting basal plates of biliary epithelial; obvious intrahepatic and hepatocytes cholestasis, bile embolism forms in small bile duct; hepatocytes ballooning degeneration with disappearance of cells; mass ground-glass hepatocyte; mild to moderate mixed inflammatory reaction ( fig. . ) . fch could quickly proceed to liver failure with blood coagulation dysfunction and hepatic encephalopathy, and mostly die in several weeks to months. due to common transmission, coinfection with hbv and hcv or hiv is not rare clinically. - % of the chronic hbv infected patients carry hcv antibody and there are - million coinfectious patients all over the world. studies showed that hbv and notable intrahepatic cholestasis (black arrow) and ballooning degeneration and/or feathery degeneration (red arrow), a large number of ground glass hepatocytes (blue arrow), associated with inflammatory response (green arrow) hcv coinfection could promote the synthesis of collagen and promote disease progression to liver fibrosis. compared to the single hbv infection, hbv and hcv coinfection presents more severe liver fibrosis and inflammatory necrosis. studies demonstrated that hbv and hcv coinfection could promote chb progression, cause severe damage of the liver function and then exacerbation, increasing the probability of liver fibrosis, liver cancer and liver failure in chb patients. in hbv and hiv coinfection, hiv infection can affect the natural history of hbv and accelerate the development to end-stage liver disease and liver cirrhosis. immune deficiency induced by hiv infection fosters hbv replication, and even fibrotic cholestasis hepatitis in severe cases. histologically, chb caused by hbv and hiv coinfection had a severer fibrosis than that by simple hbv infection. cases of hepatitis b overlapping drug or alcohol induced liver damage are not rare. even antiviral drugs can cause aechb, and there is previous case report on hepatitis b patient died of acute liver failure induced by anti-hepatitis b virus medication lamivudine. abuse or nonstandard drug use and alcoholism have become the common causes of aechb. pathology manifests features of overlapping drug or alcohol induced liver injury on the basis of hepatitis b changes. for instance, in aechb caused by overlapping drug-induced liver injury, liver tissues present histological characteristics of hepatitis b accompanied with drug-induced liver injury, such as evident increased percentages of infiltrated eosinophils and neutrophils in liver tissues, confluent necrosis with less inflammatory cell infiltration in acinus three area, cholestasis of bile canaliculi and so on. in summary, aechb has its relative histopathological features. understanding of these pathological characteristics can not only help with clinical diagnosis and effective treatment, but also aid to prevent aechb. it is important to note that despite the value of histopathological examination in diagnosis, classification and prognosis assessment, considering the significantly decreased coagulation function of liver failure patients and liver biopsy examination has certain risk, hence more attention should be paid to indications of liver biopsy in clinic. laboratory tests for liver diseases is the important basis to help and ascertain the clinical diagnosis, and the important reference to evaluate disease severity, make classification, predict outcome and guide therapy. the laboratory tests may reflect the pathological change and the functional status of liver in time, and may provide the objective and detailed data as reference for clinical classification and evaluating therapeutic effects, so that clinical intervention and effective treatment can be performed successfully. the liver is a complicated organ and the laboratory test items of relevance to severe hepatitis b are many, there are various biochemical items reflecting liver function, including coagulation function, immune and inflammatory cells and genetic markers. in this section only those laboratory tests that are relevant to acute exacerbation of chronic hepatitis b and severe hepatitis b will be described. for nonspecific laboratory tests, the reader is referred to other more general pathology books and literature. serum bilirubin is not a sensitive parameter of hepatocellular injury, but a significant increase (commonly ≥ ten times of upper limit of normal value) is usually a specific manifestation of acute exacerbation of chronic hepatitis or liver failure, which is also necessary condition to diagnose severe hepatitis or liver failure. in the course of acute exacerbation of hepatitis both direct and indirect bilirubin rise markedly due to the disturbance of bilirubin metabolism and secretion because the injury and hypofunction of hepatocytes, paracholia, and the rupture of bile capillary and biliary duct. the main enzymes reflecting liver function are alanine aminotransferase (alt), aspartate aminotransferase (ast), γ-glutamyltransferase (γ-gt, ggt) and cholinesterase (che). enzyme protein content in the liver account for . % of total liver protein. because the aminopherase content of the liver is times that of blood, in a pathological condition, as long as % of the enzyme in the liver is released into blood and keep active, this will be enough to keep the activity of enzyme in the serum increasing at rate of double. alt is an enzyme with the highest increasing amplitude and highest positive incidence when acute liver damage is occurring, with activity in the liver times that of serum. there is a large range of activity in daytime, commonly higher in the afternoon than the morning. although the activity of alt is almost coincident with the degree of liver damage, the activity of the enzyme decreases rapidly when liver failure or hepatocyte necrosis becomes widespread, with significantly increasing levels of serum bilirubin, manifesting the disassociation of enzyme and bilirubin. alt mainly resides in the cytoplasm of the hepatocyte, whilst ast is found more in the mitochondria. when liver cellular necrosis and change of cell membrane permeability appear, there is more release of alt than ast, but in very severe damage of liver, mitochondria damage is witnessed with elevation of ast and significant elevation of ast/alt. che in the serum is mainly produced by liver, with its activity and synthesis decreasing when liver damage occurs. although the change of che is less compared to the aminopherase when liver cell damage occurs, it will drop dramatically when there is severe liver necrosis and liver decompensation, especially in liver encephalopathy. the half life of human serum albumin is - days. because the occurrence of low levels of serum albumin is usually a late marker, the albumin level cannot accurately reflect acute liver cell damage, especially during the acute exacerbation of chronic hepatitis b. serum globulin, especially the γ-globulin, is only elevated in particular chronic hepatitis situations such as decompensated cirrhosis and autoimmune liver disease. the half life of serum prealbumin is only . days, and its reaction is more rapid and sensitive, which can reflect liver cell damage earlier. serum prealbumin has special diagnostic value with acute exacerbation of liver especially acute and subacute severe hepatitis. total cholesterol is composed by cholesterol ester and free cholesterol in healthy people. when hepatocellular damage appears, its cholesterol esterification noticeably decreases. it has been reported that α lipoprotein decreased significantly in patients with liver failure and this indicates a poor prognosis, while the changes of other lipoprotein and serum triglycerides were not specific for the severe type of patients. it has been reported that patients with severe liver dysfunction usually have bile acid metabolism abnormalities, indicating that serum total bile acid is a sensitive indicator reflecting functional recovery of liver cell and improvement of pathogenetic condition, just like alt. detecting serum total bile acid plays an important role in predicting disease prognosis of severe liver failure and evaluation of therapeutic effect. plasma coagulation factor xii (hageman factor) can produce activating factor xii by surface activation, which can further activate kallikreinogen (also called prekallikrein, pk) to produce kallikrein, kallikrein then makes prokinin release bradykinin, which cause vasodilatation, increasing capillary permeability with decreasing blood pressure. because of the short plasma half-life of pk, its plasma content decreases rapidly during liver failure, so pk has important diagnostic value with acute severe type hepatitis and liver failure. it was reported that its content was ± % in healthy controls, ± % in survivors with decompensatory liver cirrhosis, while only ± % in non-survivors. a content level below % predicts poor prognosis, with patients usually dying of liver dysfunction within - days. gst is a protein rich in the liver, the renal tubules and intestine cells in mammals. its major function is detoxication by combining with multiple metabolistic organics such as bilirubin, bromphenol, cholecystographic agent and epoxide. because of the high content of gst in the hepatocyte, its small molecular weight and short half-life (only min), it can be released into the blood after hepatocyte necrosis with high concentration. thus, gst has become a good predictor of observing hepatocyte necrosis especially in patients with acute liver failure and fulminant hepatitis, not only for early diagnosis of hepatocyte necrosis, but also for predicting prognosis. other factors such as multiple circular lysosomal enzyme and serum hyaluronate are all significantly elevated during a course of liver failure. these markers can also be used for monitoring and diagnosis of the disease. severe hepatitis and liver failure caused by hbv infection and other pathogenic factors are not obviously different under laboratory examination, although they have different etiological factors, pathogenesis and clinical manifestation. the major liver function, including protein (especial various kinds of coagulation factors) synthesis, metabolism and detoxication, are the first to lose function, and so result in severe hemorrhage and hepatic encephalopathy. because of the difference on the production of coagulation factors and the steps they involve in, not all coagulation factor detections are suitable for detection of severe type hepatitis and acute exacerbation of hepatitis b. vitamin k dependent factors mainly includes prothrombin (factor ii), pre-convert in (factor vii), christmas factor (factor ix) and fibrin stabilizing factor (factor x). patients with severe type hepatitis can manifest vitamin k deficiency caused by bile accumulation exterior and interior of liver, ingestion reduction or diarrhea. factor vii with the shortest half-life ( - h) is influenced firstly, which cause prolonged prothrombin time (pt). factor ii is not sensitive on vitamin k deficiency, while factor ix and factor x are moderately sensitive. factor vii can be considered as a reliable indicator, and it has important clinical value on prediction of prognosis of acute liver failure because of its short half-life and less influence by other factors, such as inflammation, dic, fibrinolysis, etc. it was reported that when the level of factor vii was below % of normal controls, the probability of death increased significantly, with predicting value of % sensitivity and % specificity. human fibrinogen-like protein /fibroleukin prothrombinase is a mediator of inflammation produced by activated macrophages, which belongs to fibrinogen superfamily and can catalyze and convert prothrombin to activate thrombin directly, thereby starting the clotting process and promoting thrombogenesis. some studies have demonstrated that the level of human fibrinogen-like protein /fibroleukin prothrombinase, if expressed highly and specifically on pbmcs and liver tissue in patients with acute-on-chronic liver failure, can be correlated with disease severity. other clotting factors such as factor vii, xi, xii, i, c protein, plasminogen and platelet count are all decreased in various types of liver diseases, with no special changes on the course of severe-type hepatitis or acute exacerbation. so they are not suitable for evaluation of acute exacerbation of chb. some scholars considered that combination detection of antithrombin iii (at iii), hepaplastin test (hp) and thrombin time (tt) has important value on early predicting fulminant hepatitis. pt is mainly used to detect activity of factor vii, x, ii, v and i. it has three measurable methods: one is the prolonged pt, normal pt is to s, with abnormality above s of normal control value; another is prothrombin activity (pta), which can be calculated by mathematical formula. normal pta is - %, usually below % in liver failure. the third is international normalized ratio (inr), which can be calculated by certain correction factor: pt in patients/pt in healthy. inr is above . in abnormality, and it is generally over . in liver failure. pta and inr detection values have been included in diagnostic criteria of international and domestic liver failure. ptt is a screening test of intrinsic coagulation system. ptt can prolong when it is faced with factor vii, ix, xi and xii deficiency or factor i, ii, v and x reduction and increment of anticoagulant substances. because ptt can be prolonged in a variety of liver diseases, demonstrating that ptt is not necessarily specific for the diagnosis of liver failure. tt tests for the activity of plasma fibrinogen. tt can be prolonged when fibrin degradation product (fdp) is increasing, fibrinogenolysis activity increasing, or fibrinogen (fib) decreasing, or heparin-like anticoagulant substances occurrence. apart from tissue-type plasminogen activator (tpa) and plasminogen activation inhibitor- (pai- ), other proteins and molecules involved in the course of fibrinolysis are all synthesized in liver. so plasminogen, α antiplasmin and thrombin activation fibrinolysis inhibitor (tafi) all decreased significantly in severe liver disease such as decompensated cirrhosis. as a result of dysfunction of liver clearance, the tpa level elevates inversely, while with normal or less elevated pai- in patients with cirrhosis, can cause proportion disequilibrium and final hyperfibrinolysis. in patients with acute liver failure, because of large elevation of pai- as acute phase reactive molecule, the activity of fibrinolysis decreases. on the contrary, taf- decreases by almost %, which induces elevation of fibrinolysis activity. although synthesis of partial coagulation factors and clotting dysfunction usually appear during the course of acute exacerbation of hepatitis b and development of severe-type hepatitis/liver failure, with manifestation of prolonged pt/decreased pta/elevated inr, some investigators had observed conflicting results recent years. specifically, although the mean inr was . ± . in acute liver injury/acute liver failure patients complicated by hepatic encephalopathy, concentrations of factor v and factor vii also decreased, the mean values of the indicators mentioned above detected by thromboelastography (teg) were normal, and also five teg parameters were normal in % patients. we think that the reason of normal clotting function detected by teg in patients with ali/alf might be the normal value of platelet count and fibrinogen quantitation. furthermore, more platelet and factor vii can be produced, and the levels of anticoagulant protein (protein c, protein s and antithrombase) decrease, which also compensates for the defect of the other coagulation factors. in total, clotting parameters such as inr, etc. can be considered as valuable indicators for predicting prognosis, although they cannot be used to reflect hemorrhage severity in patients with ali/alf. ammonia has been considered as a precipitating factor of hepatic encephalopathy for more than years. during severe liver dysfunction, carbamide synthesis is injured, and brain tissue becomes a major organ of ammonia detoxication. with glutamine synthetase, astrocytes in brain can convert glutamate to glutamine to remove accumulated ammonia in vivo by the amidation of ammonia. because the synthesis of glutamine consumes energy, the large consumption of atp can cause energy exhaustion. excess accumulation of glutamine in astrocytes induced by high blood ammonia can cause increasing osmotic pressure and brain cellular edema. this has been confirmed by mri, while with the recovery of liver antidotal function after liver transplantation, previous hepatic encephalopathy can be reversed. in patients with acute liver failure, the risk of cerebral hemorrhage increases rapidly when arterial blood ammonia is above μmol/l. about % patients can have acute intracranial hypertension when arterial blood ammonia exceeds μmol/l. bernal w, et al. in their study which involved patients with acute liver failure, observed that the level of arterial blood ammonia on admission was an independent risk factor of hepatic encephalopathy and intracranial hypertension, and the sensitivity was % for predicting severe hepatic encephalopathy with arterial blood ammonia above μmol/l. combining with the meld score can further increase specificity and sensitivity. the toxicity of ammonia is multiple. the accumulation of ammonia not only influences brain metabolism, injuries brain cellular organelle directly and indirectly, causes disequilibrium of brain internal inhibitory and excitatory neurotransmitter and injuries brain energy metabolism, but also changes a series of gene expressions that are important proteins maintaining brain function. autoregulation of cerebral blood flow is affected and the blood brain barrier broken down. latest studies have revealed that ammonia can injury many important functions of neutrophil, such as chemotaxis, phagotrophy and degranulation, and can also stimulate and produce large reactive oxygen species (ros), cause sirs, which then further aggravates the toxic effects of brain cells from ammonia resulting in a vicious cycle. the liver is an important site of amino acids metabolism. except for bcaa (leucine, isoleucine and valine) metabolism in skeletal muscle, almost all essential amino acid metabolism occurs in liver. because of this, patients with liver failure or cirrhosis, have large amounts of amino acids accumulate in the blood. fischer, et al. demonstrated that disequilibrium of plasma amino acids might be reason of encephalopathy, and further indicated that the molar ratio of valine+leucine+isoleucine and phenylalanine +tyrosine (bcaa/aaa) was closely related with the severity of hepatic encephalopathy. the analysis of plasma amino acids in animals with hepatic encephalopathy demonstrated that other concentrations of amino acids increased significantly except the concentration of arginine, which declined. analysis of brain homogenates from cases of fatal hepatic encephalopathy demonstrated that the concentration of aspartate, arginine and glutamate decreased significantly, and this was closely related with the severity of hepatic encephalopathy. other amino acids, especially the aromatic amino acids, such as tryptophan, phenylalanine and histidine, increased but with carrying amounts; thus, the concentration of the aromatic amino acids was closely related to the severity of hepatic encephalopathy, implying that these aromatic amino acids may play important roles in the pathogenesis of hepatic encephalopathy, although perhaps not as the primary driver. jiang y, et al. conducted a clinical study which enrolled patients with acute hepatitis, patients with chronic hepatitis, patients with severe hepatitis and cirrhosis patients. they observed the ratio of bcaa and aaa was normal in acute hepatitis, mildly lower in the chronic hepatitis (p > . ), significantly lower in the severe grade of chronic hepatitis (p < . ), and the lowest in the patients with severe-type or cirrhosis (p < . ). as for the child-pugh grading, the ratio of bcaa and aaa: c grade <b grade <a grade, with significantly different among the groups (p < . or p < . ). of the patients with severe-type hepatitis and cirrhosis, the ratio of bcaa and aaa in the patients with hepatic encephalopathy was lower than that without hepatic encephalopathy (p < . ); the ratio of bcaa and aaa was significantly lower in the non-survivors compared with survivors (p < . or p < . ). they concluded that detection of bcaa/aaa can reflect the degree of liver damage, the lower the ratio was, the worse the liver injury was, and with high risk to develop hepatic encephalopathy; the ratio might also have certain predictive value for prognosis. elevation of plasma free tryptophan has been correlated with aggravation of hepatic encephalopathy; while in patients with acute liver failure, serum methionine concentrations clearly increase, there is no overlap between acute hepatitis and acute liver failure, and this can be used for differential diagnosis between the two kinds of liver diseases. endotoxin is produced by gram-negative bacteria, and has many biological activities, which can cause endotoxemia, with multiple pathological changes, such as fever, shock, dic and granulopenia. critically ill patients even have acute respiratory distress syndrome (ards), acute renal failure (arf) and multiple organ dysfunction syndrome (mods)/multiple organs failure (mof). wang mr, et al. revealed that the incidence of endotoxemia is - % in severe-type hepatitis patients, . - . % in decompensated cirrhosis, . % in compensated cirrhosis, and % in acute viral hepatitis. it has been considered that the main mechanism of endotoxemia in patients with cirrhosis and severe-type hepatitis is: ( ) intestine mucosal nutritional disturbance, epithelial atrophy, shedding and ulceration, which cause injury of intestine mucosal barrier; ( ) intestine mucosal congestion, edema and increased intestinal wall permeability; ( ) alteration of intestinal flora resulting in translocation of bacteria from the intestinal tract and increasing endotoxin production and absorption; ( ) low ability of clearance of endotoxin absorbed by intestine or endotoxin into systemic circulation from bypass circuit. most studies have demonstrated that endotoxemia and liver injury are interrelated; the liver injury can be aggravated when accompanied by endotoxemia. the detection of endotoxemia is beneficial for predicting clinical course, disease severity and prognosis. medium molecule substance was discovered in patients with uremia accepting hemodialysis, and these molecules have molecular weight of - d. thereafter, it was confirmed to play an important role in hepatic encephalopathy. medium molecule substance can inhibit na + -k + -atp enzymatic activity of brain cells, which result in na + retention and cause cerebral edema. studies have confirmed that medium molecule substance in the blood increased when acute liver failure patient presented with hepatic encephalopathy, which indicated that an increasing amount of medium molecule substance was closely correlated with hepatic encephalopathy and could be used for predicting clinical severity and prognosis. as for the property of medium molecule substance, it has not been totally characterized. the clearance of human medium molecule substance is mainly by secretion and filtration from the glomerulus. according to clinic observations, the contents of medium molecule substance can be detected in the plasma when plasma creatinine (cr) concentration is above . μmol/l or creatinine clearance rate (ccr) is below ml/min. lactic acid is an intermediate product of glycometabolism. the concentration of lactic acid can reflect glycometabolism, peripheral circulation, tissue blood-supply and oxygen-supply indirectly. large studies has demonstrated that total blood lactic acid levels are correlated with disease severity and prognosis. the higher the lactic acid is, the worse the clinical condition is, and also for predicting a poor prognosis. some studies have revealed that the arterial blood lactic acid has a low predictive value for liver failure caused by hepatitis virus infection, toxin and immunity, but a better predictive value for acute and super-acute liver failure caused by acetaminophen over-dose. however, this is controversial. schmidt le, et al. showed that the predictive specificity of lactic acid was not high, even for liver failure caused by acetaminophen. furthermore, lactic acid cannot be used as an indicator for of liver transplantation, and if used, must be done in association with the current king's college hospital (kch) criteria. procalcitonin (pct) is a precursor of human calcitonin, with composed of amino acid residues, kd of relative molecular weight, and also has no hormone activity as a glucoprotein. pct mainly originates from the c cells of the thyroid. during pathologic conditions, multiple tissue and organs can become production areas, such as liver, lung and glandular tissue. the production of pct can be regulated by bacterial toxin and multiple inflammatory factors, and bacterial toxins are a major stimulating factor inducing production of pct. pct is the best ideal indicator for early diagnosis of systemic infection, and has high specificity. serum pct concentration is positively related with the degree of bacterial systemic infection, but can also descend to normal level gradually accompanied with control of bacterial infection and improvement of clinical condition. as for the cirrhosis patients with ascites, detection of serum and ascitic pct can be of benefit to the clinician for predicting early diagnosis and the curative effect and evaluation of spontaneous bacterial peritonitis (sbp). the pct levels can decrease significantly in cirrhosis patients with sbp upon effective antibiotics treatment. so, pct can be considered as a good predictive factor on diagnosis and differential diagnosis of cirrhosis ascites complicated by bacterial infection. it has been reported that the blood lactic acid and pct levels were elevated to different levels when facing patients with either acute liver failure or acute exacerbation of hepatitis b complicated by intestine endotoxemia. a persistently elevated compared to an or not decreasing level of lactic acid and pct are usually indicative of poor prognosis and therapeutic efficacy. combined detection of blood lactic acid and pct does have certain predictive value for acute exacerbation of hepatitis b. in patients with chb, peripheral blood tregs were positively related with serum virus load, while in patients with acute hepatitis b, peripheral blood tregs experienced process from low to high, and usually are normal in the acute, convalescence, and recovery stages respectively. for the patients their treg were rejected, the ability of ifn-γ secretion in the pbmcs increased significantly upon antigen stimulation. while the tregs isolated from the patients with hbv infection could inhibit proliferative response of hbv antigen treated with autologous pbmcs obviously, which might indicate that the peripheral blood and liver in the patients produced hbv antigen specific tregs. so, tregs not only play an important role on regulating immune response of hbv infection, but also influence the prognosis of the patients with hbv infection. natural killer cell (nk) and natural killer t cells (nkt cells) construct the first line of defense of body to defense pathogen invasion, which account for % and % respectively in peripheral lymphocytes, while higher with % and % in the liver. the content of nk cells account for % in liver lymphocytes. the accumulation of nk and nkt cells in the liver demonstrates that they have important function, which can directly eliminate virus by killing infecting cells or secreting cytokines. under the condition of virus specific igg antibody production, nk cells can mediate surface igg fc receptor (fcγriii), identify and kill target cells with complexed igg specifically by antibody dependent cell mediated cytotoxicity (adcc). furthermore, nk cells can also play a role in immunoregulation by secreting cytokines, such as ifn-γ, il- and tnf. nkt cells are a group of inherent t cells which can recognize lipoid and glycolipid antigen presenting by mhc-i molecules associated with antigen presentation, with cd d molecule restriction. nkt cells manifest phenotype of both t cells and nk cells (tcr and nk . [nkrp c], ly- ). nkt cells have two subtypes: type i nkt cells, also called inkt cells, and type ii nkt cells, also called non-inkt cells. the important characteristic of inkt cells is to produce th and th type cytokines rapidly by activation of tcr signal, and also activate multiple other cell types, such as dc, nk cell, b cells, and conventional t cells rapidly by activation of tcr. so, inkt cells can influence adaptive immunoresponse and series of host defense reaction and pathological course effectively. tian et al. demonstrated that nk cells exist abundantly in normal liver, which account for % of lymphocytes within liver and can play important roles on natural immunoresponse of anti-hbv infection. nk cells within the liver can be directly activated by virus or indirectly activated by other cells, such as nkt cells and antigen presenting cells (apc), and exert antiviral effects by their natural cytotoxicity and production of high level of antiviral cytokines. also they can regulate the function of lymphocytes, such as t cells, b cells, apc, and co-ordinate an adaptive immune response. after the binding of fasl expressed by ctl and nk cells and fas expressed by liver cell, the intracellular region of fas protein binds with fas associated death domain (fadd) protein by its c-terminal death domain (dd), then fadd protein activates caspase and lead to apoptosis. malhi h et al. showed that the role of nk cells and nk t cells was a major factor leading to mass death of hepatocyte during the clinical course of viral hepatitis. dendritic cells (dc) are antigen presenting cells with the most powerful function not only playing an antiviral role by secreting cytokine, such as il- and ifn-α, but also by promoting immunoclearance by t lymphocytes activated via antigen presentation. furthermore, dc also can regulate the equilibrium of th and th . so, the abnormity of number, phenotype and function of dc is bound to influence the outcome of hbv infection. some studies illustrated that during liver failure the number of pdcs and mdcs in the liver increased significantly; ifn-α produced by pdcs in the liver was correlated with the production of il- and il- ; the ability of ifn-α production by pdcs in the peripheral blood decreased and the degree correlated with disease severity, which indicated that the dcs in the peripheral blood had accumulated in the liver and activated when hbv associated acute-on chronic liver failure occurred. after dc activation, the role of antigen presentation was enhanced, and the immune response activated and even to a peak level. zh qian et al. observed that the dc of peripheral blood in patients with chronic severe hepatitis b manifested dysmaturity, dysfunction of cytokine excretion, especially the low secretion of il- which mediated cellular immunity, a high level of secretion of il- and tnf-α which mediated inflammatory reaction, and finally become an important factor of aggravating liver inflammatory reaction leading to severe hepatitis. th cell is a new discovered subgroup of cd +t lymphocytes, with significantly different biological function, cytokine expression and differentiation compared with th and th cells. th cells can mobilize, recruit and activate neutrophil leukocytes by secreting cytokines such as il- , il- and tnf-α, in order to mediate inflammatory reactions more effectively. zhang et al. detected the th cells in the peripheral blood and liver of chronic hepatitis b (chb) and hbv associated acute-on-chronic liver failure (aclf) patients. they observed the level of th cells and its associated cytokines increased in chb and aclf patients, and the increasing of th in the peripheral blood and liver is significantly positively associated with hbv dna load, serum alt level and histological index of activity. in vitro, il could promote mdcs and monocyte activation, increase its secreting ability of proinflammatory factors, such as il- , tnf-α and il- , which demonstrated that th might lead to aggravation of liver injury in chb patients, and th can be considered as one of the immunologic indicators reflecting clinical prognosis of hbv associated aclf patients. wu, et al. observed that the ratio of peripheral th in patients with ahb and severe hepatitis increased significantly compared with general patients with chb and health volunteers. th cells were significantly positively associated with serum alt increasing in severe hepatitis, while there was no correlation with hbv dna quantitation. furthermore, serum il- levels were significantly negatively associated with the ratio of th cells, which indicated that th cells correlated with the course of hbv and the severity of liver injury. ye et al. also observed that the ratio of secreting il- and secreting ifn-γ in the liver of the child-pugh c patients with hbv infection was significantly higher than that of child-pugh b; the numbers of the cells secreting il- in the patients with severe liver injury were more than the cells secreting ifn-γ; the degree of cell infiltration with il- secreting were significantly positively correlated with liver inflammation grading, expressing of il- in the liver and neutrophilic leukocyte infiltration. chen et al. observed that increasing serum ifn-γ levels was correlated with disease progression of chb. ifn-γ might be involved in inherent and acquired immunity after hbv infection; ifn-γ can be considered as a potential biomarker for predicting clinical severity of hepatitis b, especially for predicting the tendency of liver failure. il- is a suppressive cytokine which can be produced by th cells, kupffer cells and liver cells, with major functions of suppressing antigen presentation of macrophage, production of multiple proinflammatory cytokines and th response. zheng et al. observed that high expression of il- promoted active viral replication in chb patients, induced excessive immune response. il- can be used to evaluate disease severity and prognosis of severe type b hepatitis. serum il- levels decreased significantly in severe hepatitis b patients complicated by bacterial infection or overlap with other hepatitis virus infections, which indicated endotoxemia and concurrent viral infection play important roles on promoting occurrence of severe hepatitis b. tnf-α is mainly produced by active mononuclear macrophages. the concentration of tnf-α is low under normal conditions, with the function of regulating the immune response. persistent high level of tnf-α can induce liver apoptosis or necrosis. tnf-α has two surface receptors: tnfr and tnfr . du et al. observed that levels of serum tnf-α, tnfr and tnfr increased significantly in patients with acute fulminant liver failure compared with subacute fulminant type, acute type patients and health volunteers, and levels of tnfr were even higher in nonsurvivors. another study showed that expression of tnf-α and tnfr was obviously positively correlated with apoptosis liver cells in patients with fulminant liver failure; tnf-α might lead to widespread liver apoptosis by inducing fas gene transcription and other signal transduction system. except for inducing apoptosis, tnf-α can also induce production or release of other cytokines, such as il- , il- , and so on, to promote intrahepatic inflammatory reaction and aggravate liver cell necrosis by cascade amplified action. ip- is a recently discovered cxc chemokine, mainly induced by ifn. ip- has strong function of chemiotaxis and activation on t cells expressing specific receptor cxcr , and also plays important role on lymphocytic accumulating to inflammatory reactive site, which may be an important factor of inflammatory cell migration toward hepatic tissue and causing large area hepatic cell necrosis. detecting serum il- concentration can indirectly reflect expression of ip- in local hepatic inflammatory response. luo et al. observed that the levels of serum ip- in patients with severe hepatitis b were higher than that of chb group, indicating the higher degree of liver inflammation and injury, the higher serum il- levels. so, serum levels of il- might reflect the degree of inflammation in hepatitis b patients. trail is a new member of tnf α superfamily. a large number of studies indicated trail-r /dr play an important role on apoptosis. the trail induced apoptosis pathway may play an important role in the pathogenesis of severe type hepatitis. trail receptor/ligand system can specifically kill cells infected by virus in hbv infection. vander sloot am et al. observed that affecting hepg cell strain by soluble trail produced by cultural monocyte after lps stimulation can cause apoptosis; higher levels of soluble trail, more obvious apoptosis, which demonstrated that trail induced hepg apoptosis was dose dependent. further studies have shown that serum soluble trail increased significantly in patients with severe type hepatitis and was positively correlated with serum lps concentration and severity of liver injury, which illustrated that decrease of lps degradation accompanied with the degree of liver injury can stimulate mononuclear macrophage and dc to express more trail, shedding strail also increased, cause liver apoptosis and aggravated liver injury after its binding with receptors on liver cells. interleukin- (il- ) is one of the major cytokines secreted by th , and is a member of the il- cytokine family. th is one of the major cells secreting il- . osteopontin (opn) is a secretary phosphorylated glucoprotein which contains rgd (arginine, glycine and aspartic acid) protein with an integrin binding region discovered in . it was isolated from bone matrix by herring in who found that opn was a key cytokine which accumulates in immunocytes and initiates th cell immunity, and is involved in multiple pathological process of inflammatory reaction. some studies demonstrated that levels of peripheral blood opn were significantly high in patients with severe hepatitis b. it also confirmed that nkt cells can secrete opn, and opn also can enlarge activation of nkt cells, to further trigger neutrophil infiltration and activation. further studies on pathogenesis of opn in hepatitis demonstrated that the opn transgenic mice, after been treated with cona, manifested large necrosis of liver cells and monocytes infiltration, while with only minor liver injury on the controls. so it presumed that opn might cause immune disequilibrium of th and th , induce large hepatonecrosis. in patients with fulminant hepatitis, opn increased significantly, indicating that opn can initiate the effect of th cytokine network such as il- and ifn-γ by an autocrine pathway, to further cause macrophage activation and severe hepatonecrosis. as a specific receptor of endotoxin, toll-like receptor (tlr ) can recognize serum endotoxin, bind with lps/lbp/cd , and recruit myd , activate irak (il- r correlated kinase) and mapkkk (mapk kinase kinase) family, induce activation of nf-κb, activate target genes transcription, release a series of cytokines, and cause hepatocytes damage. in patients with severe hepatitis, because of intestinal microbial population derangement, bacterial overgrowth and intestinal mucosa congestion, the ability of endotoxin production and absorption increases, which is accompanied by severe hepatonecrosis, immune dysfunction and a decrease in kupffer cells clearance, which further causes a reduction of exdotoxin and exdotoxin immunocomplex clearance, and finally gives rise to endotoxemia. lipopolysaccharide (lps) is a major component of gram-negative bacteria, with important physiopathologic function. tlr is a key receptor for lps via signal transduction. regulating expressing of tlr should control lps related inflammatory reaction. xu, et al. designed and constructed tlr sirna expression vector, evaluated mouse phagocyte raw . gene silencing efficiency by transfection, and evaluated therapeutic efficacy of tlr gene silencing in the mouse hepatic injury model. it demonstrated that the expression of tlr mrna and protein decreased significantly in raw . cells; tlr sirna can inhibit obviously the up-regulation of tnf-α and mip- treated with lps; the activation of p -mapk and erk / by lps can down-regulate by tlr sirna. further studies demonstrated pretreatment of tlr sirna can be of benefit to control lps inflammatory reaction, reduce hepatic lesion of g bl/ mouse treated by d-ga n/lps, and reduce mortality of mice with acute hepatic injury. some studies also showed that the tlr protein expression on the surface of pbmcs and mrna were obviously higher in patients with chronic hepatitis b compared with the healthy controls; levels of tlr in severe-type patient of chb were significantly higher than that of chb, which indicated tlr played important roles during the course of liver injury with hbv infection, and with obvious increasing accompanied with clinical severity. fan jg, et al. observed the expression of tlr increased accompanied with the degree of endotoxemia and the prolong of stimulus duration; lps can positively up-regulate expression of tlr and enlarge biological effect of lps. so the severe hepatonecrosis and weakening of kupffer cell deactivation can accelerate the occurrence of endotoxemia, while endotoxin also can regulate liver cell metabolism or influence tlr mediated immune reaction to aggravate the clinical situation. the two aspects interact quite closely. all the studies mentioned above indicated high expression of pbmcs tlr in patients with chronic hepatitis, especially with severe-type presentation. monitoring dynamic changes of tlrs might be benefit for treatment. nitric oxide (no) is a biological regulatory factor with an extremely short halflife, produced by l-arginine catalyzed by nos (no synthetase), with extensive biological functions. it is now believed that no has potential antiviral inductive activity, and also might be one of the factors mediating liver injury. the studies of mouse model with inos defection demonstrated that the mouse model can tolerate ctl induced immunologic injury during the course of inhibiting virus replication because of no dyssynthesis, and also opposed against injection of fatal anti-fas antibody, further caused no obvious hepatonecrosis after treated with anti-fas antibody. in severe-type patients with hepatitis, serum no levels significantly increase during the early stage, then no levels decrease significantly with extenuation of inflammatory reaction, liver function recovery in the convalescence phase. these studies revealed that no is related to the degree of inflammation and clinical severity, and can play an important role in the clinical course of severe-type hepatitis. some studies demonstrated that inos gene was related closely with viral hepatitis. the expression of inos gene was extremely low in normal hepatic cells. once invasion of hepatic cells by virus appeared, it can induce hepatic cells to activate inos gene expression and produce large amounts of no, not only to kill liver tumor and pathogens such as virus, bacterium, parasites, play broad-spectrum antiviral function, but also injury to adjacent normal liver tissue by cell toxic action, and cause liver cell death. gong fj, et al. presumed that pre-s protein can downregulate inos transcriptive activity by its trans-regulation, further inducing the decline of no production and blocking the clearance of hbv in hepatocytes by no, and finally accomplishing long-term hbv chronic infection. so, expression of inos can be considered as an important factor of inflammatory injury. neutrophil gelatinase-associated lipocalin (ngal) was considered as a novel troponin-like biomarker, which was discovered in by kjeldsen, et al. who studied neutrophil matrix metalloproteinase (mmp- ) at that time. it was believed that ngal could be used to monitor acute kidney injury/acute renal failure. renal dysfunction is extremely common during the clinical course of severe liver disease such as decompensated cirrhosis and liver failure. serum cr levels can increase rapidly when acute renal failure occurs and is easily detected, while it is not easy detected or diagnosed when complicated by chronic kidney injury. furthermore, other diagnostic methods based upon glomerular filtration rate (gfr) are needed to detect clearance rate, and are not suitable for routine application. the detection of serum cr is simple, while is not accurate on detection of kidney injury based upon cirrhosis. gerbes al, et al. observed cirrhosis patients with ascites whose serum cr below . mg/dl, compared the levels of serum ngal and cr. the patients was divided into two groups, the gfr was ± ml/min and ± ml/min, and with serum cr of . ± . mg/dl or . ± . mg/dl respectively, with no obvious difference. the estimated gfr was ± ml/min and ± ml/min respectively, also with no difference. while in the study, serum ngal was ± ng/ml and ± ng/ml, demonstrating significant difference (p < . ). furthermore, levels of urinary ngal is similar with the blood ngal, especially the patients in the group , with urinary ngal above ng/ml. area under the curve (auc) analysis also revealed that ngal was better than serum cr assay when gfr < ml/min, with auc = . ( . ~ . ) of the former, and . ( . ~ . ) of the latter (p < . ). macrophage inflammatory protein (mip) is a novel protein first discovered in by wolpe, et al. which can be divided into five subtypes: mip- , mip- , mip- , mip- and mip- . mip- has a binding site for heparin, which can interact with heparin dextran sulphate in endothelial extracellular matrix, and enhance adhesion of leucocytes and vascular endothelial cells. mip- is involved in the whole process of inflammation by chemical chemotaxis and activation of inflammatory cells, with neutrophils as specific target cells. mip- can also activate neutrophils specifically and has been considered as an important proinflammatory cytokine during the early stage of inflammation. chen z, et al. observed that the levels of mip- in severe-type patients with hepatitis b was significantly higher than other groups. persistent high expression of mip- can be used to predict acute exacerbation of hepatitis b and clinical severity of patients with hepatitis b. thymosinβ (tβ ) is one of the important actin regulatory molecules in human body. it can bind with globular actin (g-actin), inhibit production of fibrous actin (f-actin), reduce formation of microthrombi and microcirculation dysfunction in patients with liver failure, and inhibit occurrence and development of mof. tβ can also reduce the levels of free radicals, slow lipid peroxidation and inhibit production of proinflammatory factors. patients with liver failure usually manifest a large amount of mononuclear macrophage infiltration, secretion, releasing proinflammatory factors and aggravate systemic inflammatory reaction. liu y, et al. observed that the levels of serum tβ decreased significantly in patients with liver failure compared with the cirrhosis group, chb group and healthy controls. dynamic changes of serum tβ can be considered as a novel predictive indicator for prognosis in patients with liver failure. t cell immunoglobulin mucin (tim- ) is a surface molecule correlated with function of t cells, which was discovered in . the studies on mouse model revealed that tim- can be expressed on the surface of th selectively as a negative regulatory molecule; the interaction of tim- and its ligand plays a down regulatory role on th cellular mediated immune response. sabatos ca, et al. observed that the levels of peripheral tim- in chb patients increased significantly, and it indicated the expression of tim- can be induced by regulating treg whose activity was modulated during the chronic infection period in patients with chb, and then inhibit specific th cell responses. zhou xq, et al. observed that the levels of tim- was significantly higher than other groups in severe-type patients with hepatitis b, which can be used to predict and monitor the clinical severity of hepatitis b patients. with the progression of chb, serum inos and ngal all increased, with the highest levels in severe grade of chb, while decreased inversely in the serum of the severe-type patients with hepatitis b, which can predict acute exacerbation of chronic hepatitis b. it was reported that the crosspoint of persistent elevation of tim and mip , and declining of inos and ngal, demonstrated important clinical significance on predicting acute exacerbation of chb. recent studies have demonstrated that genetic variation of hormone receptor gene was significantly different between chb patients with acute exacerbation and hbv carriers, which could explain the sexual difference on different clinical course of hepatitis b. clinical studies further demonstrated that male patients with hbv infection had higher incidence and ratio of chronicity; male patients with hbv infection had higher activity frequency, which indicated the sex hormone and its receptor might influence the process of hbv infection by regulating host immuno response and level of viral replication. it is now believed that immunoresponse of viral clearance might down-regulate male sex hormones and up-regulate female sex hormones. furthermore, steroid hormones might regulate replication of hbv mediated by androgen receptors. it has been proved that androgenic hormone can inhibit cellular and humoral immune reaction; estrogenic hormone can inhibit cellular immunoresponse and enlarge humoral immunoresponse inversely; progestogen can promote conversion from cellular immunity to humoral immuno reactivity. several studies have reported the action of estrogen receptor (esrs) in the diseases of multifactorial inheritance, and its roles have also been observed by geneticists and virologists. large number of studies have demonstrated that esr played important roles on regulation in patients with persistent hbv infection, and esrs genes were correlated with host hereditary susceptibility. because of low expression of esrs in hbv carriers, the reaction of immune system to sex hormone was insufficient, which make clearance of hbv difficult and can lead to hbv persistent infection. human esrs are divided into two types: estrogen receptor a (esr ) and estrogen receptor b (esr ). all the genetic variations can cause degression of estrogenic function, which may further cause different kinds of host hereditary susceptibility after hbv infection. of them, esr gene can mutate, with tendency of sex hormonal resistance. sex hormone mainly plays the role of binding esr , and esr gene is the true minor gene with host hereditary susceptibility after hbv infection. deng gh, et al. observed the relationship between esr gene polymorphism and chronic persistent hbv infection in a large sample study. they found that the susceptibility increased significantly compared with the individual of esr t/t genotype and the individual containing at least one c isoloci site (p < . ). linkage disequilibrium plotting analysis indicated that t c polymorphism included a linkage disequilibrium area sited from esr promoter to intron , which indicated that the esr t c convention originated from esr itself, and esr gene polymorphism in the chinese population was positively related with hbv persistent infection. the negative feedback regulation of hypothalamus-pituitary gland-genital gland axis can be performed by serum testosterone (t) and estradiol (e ) through central nervous system feedback, and control releasing of luteinizing hormone (lh) and follicle stimulating hormone (fsh) . some studies demonstrated that the levels of serum t and e decreased in male patients with hepatitis b cirrhosis, and the levels of lh and fsh increased correspondingly, which indicated liver injury might be an important reason of sex and hypophyseal hormone abnormality. until now, the studies on changes of sex and hypophyseal hormone in acute exacerbation of hepatitis b and severe-type hepatitis are still limited, while it is certain that precise detection of sex hormone and correlated hypophyseal hormone can reflect the severity of liver injury to a certain degree, and can be considered as a useful indicator evaluating the clinical course of hepatitis b, cirrhosis and its complication, hepatocellular carcinoma. it is now believed that the priming or control point of severe-type hepatitis occurrence and development is influenced by two aspects: virus (such as viral variation, viral protein and viral genotype) and host (such as cell immunity, cytokines and apoptosis), and of them, viral replication is the necessary condition and cause of severe-type hepatitis. some studies demonstrated that viral replication was relative vigorous during the early stage of severe-type hepatitis, which indicated viral factor occupy a central position in the occurrence and development of acute exacerbation of hepatitis b. the changes of host transcription factors and hbv genetic variation all possibly cause the changes of hbv transcription and replication, resulting in acute exacerbation of hepatitis b. hepatocyte nuclear factor (hnf ) is a protein with the function of genetic transcription and modulation, and is mainly found in liver. long ce, et al. observed that the hnf binding sites, located in enh i/xp and enhii/c promoter, is an important factor regulating c promoter activity and . kb mrna transcription, and has an important role on hbv replication and expression with high tissue specificity. previous studies demonstrated the expression of hnf increased in patients with chb, and the level of hnf is positively correlated with hbv replication. furthermore, from a study on liver biopsy specimens using gene array in patients with hbv infection, honda m, et al. observed hbv infection can cause more gene transcription of hnf , rxr/ppar and c/ebp, and possibly related with liver injury. so, the interaction of hnf and hbv genetic transcription and control might play an important role on the pathogenesis of severe-type hepatitis b. deng et al. studied cxcl snp variation on th immune response pathway, and observed that the polymorphism site g- a existed in cxcl gene promoter region was significantly related with disease progression in male hbv carriers. an emsa study revealed that the g- a site can change its binding ability with nucleoprotein; and reporter gene assays also revealed that this site can influence the transcriptional activity of cxcl gene promoter region; mrna real time quantitative experiments further revealed that the expression of susceptible allelotype - g in pbmcs was significantly higher than allelotype - a; elisa and immunohistochemical assays revealed a higher expression of cxcl protein in plasma and liver tissue of patients with chb that progressed than that of nonprogressive patients. all the studies mentioned above demonstrated that g- a is a new functional polymorphism site, cxcl protein involved in the process of inflammation and necrosis in liver tissue of patients with hepatitis b, and influence the progression of chb. furthermore, yan z, et al. reported that mutation of ifn-γ gene can influence expression of ifn-γ, which is a predisposing factor of chronicity of hbv infection; fulminant hepatitis patients with poor prognosis had high frequency of occurrence of - c, - a and tnf-βb allele in tnf-α gene promoter region; snp in il- gene promoter region was related with progression of chronic hbv infection; the frequency of haplotype expressed by down-regulated il- in the il- gene promoter region in patients with fulminant hepatitis was higher than the controls, while the frequency of haplotype gene occurrence by up-regulated il- expression was even lower. yan et al. also revealed the relationship between polymorphism of il- gene promoter in th immuno response pathway and chronic severe-type hepatitis, which provided new evidence on natural selection theory of il- promoter and sirs pathophysiology theory of acute liver failure. in a study of human fibrinogen-like protein (hfg ) gene regulation mechanism and network, han et al. observed that hbc protein and hbx protein all had the function of hfg activation, while hbs protein cannot activate the gene. a series of promoter deletion experiments demonstrated that the regulation sequence of activating hfg gene existed between the sits of hfgl gene promoter − ~ − , transcription factor c-ets- can bind with cis-acting element on hfg gene promoter, then activate the expression of the gene under the action of that viral protein; the phosphorylation level of p-jnk and p-erk was enhanced in patients with severe hepatitis b than healthy controls, which indicated jnk and erk signal pathways were activated. some studies in vitro revealed that jnk and erk signal pathways were activated respectively under the effect of viral protein hbx and hbc, and c-ets- expression and transposition were correlated with the activity of jnk and erk. all the studies mentioned above indicated that viral protein hbc and hbx can activate jnk pathway and erk pathway respectively, further activate transcription factor c-ets- , shift into the nuclear, and finally up-regulate the expression of hfg gene by binding with cis-acting element of hfg gene promoter. tnf-α is an important proinflammatory factor induced during liver failure. mfg /prothrombinase plays an important role on mouse fulminant hepatitis and acute on chronic liver failure. gao et al. constructed eukaryotic expression vector of mfg gene and tnfr gene, green color fluorescence fusion protein and its shrna interfere plasmid. it was more effective to improve the survival rate of mouse with severe type hepatitis using combination of fg and tnfr genes compared with single gene intervention, and improve serology and pathological changes of liver, which indicated that fg and tnfr genes might produce synergistic effect during the progression of severe-type hepatitis. hbv infection is a necessary condition and cause of occurrence and progress of severe-type hepatitis, while hbv gene transcription and replication are a key step of the hbv life cycle. beginning from the new point of genetic transcription, further exploring the effect and mechanism of acute exacerbation of hepatitis b on hbv genetic transcription and regulation, will provide important clue of host transcription factor, which can be useful to predict and monitor the occurrence, development of severe-type hepatitis, and to provide new targets for effective methods of treatment of severe-type hepatitis. genome-wide association study (gwas) has been available since , and have become the main aspect of complicated genome association study. the strategy does not need to select candidate genes, but to make genome-wide associations and analysis by selecting directly ten thousand single nucleotide polymorphism (snp) sites and copy number variation (snv) sites over the total genome. compared with the strategy of a candidate gene, gwas was a significant improvement in statistical efficacy, and also avoided the bias of group stratification and randomness of gene selection. in recent years, some studies on the aspects of viral hepatitis and hbv related hepatocyte carcinoma has gained great breakthrough using these approaches. several independent and large sample gwas in usa, europe and japan demonstrated that only rs site near the il- b gene had relationship with curative effect of interferon in the study on genetic correlation of ifn response in patients with hepatitis c (with p value of − ). guo et al. observed that gene variation of hlp-dp gene can play great influence on clinical severity of persistent chronic hbv carriers. karlsen et al. also reported the relation between hla-ii gene and hbv infection. he fc and zhou gq made gwas of hbv related hepatoma, and they discovered a related site p . , where one of the three genes (kif b, ube b, pgd) might be predisposing gene of hbv related hepatoma. the studies on genetic predisposition of severe-type hepatitis b are relatively less. only a minority of studies have examined a candidate gene-disease associated study strategy, and their study populations were all from asia. recently, under the support of the national program and key state science and technology project of infectious disease, chinese experts have taken affymetrix snp . array (including , snp sites and , cnv sites in the range of human total genome-wide) to make gwas in severe-type patients with hepatitis b and asymptomatic carriers. the project is now being analyzed, expecting to explore common genetic variance sites influencing severe-type hepatitis b under the point of host total genome. epigenetics mainly studies the change of herediable gene expression with no change of dna sequence. currently, epigenetic mutation mainly includes dna methylation, microrna (mirna) and chromatin remodeling. several functional studies also demonstrated that mirna participated and regulated the expression of multiple inflammatory signal factors. it was found that mir- and mir- might be linked with liver inflammatory activity of viral hepatitis. mir- mainly participated in regulating maturation of b lymphocytes, and also regulate the differentiation of th and th cells. mir- located in human x chromosome, and highly expressed specifically in granulocyte series cells. johnnidis, et al. observed the granular leukocytes increased twice in mir- gene knock out mouse, and these leukocytes were more sensitive when treated with antigenic stimulation outside (especial fungi); compared with mir- gene non-knock out mouse, the lung inflammation of mir- gene knock out mouse was more obvious, and the tissue injury induced by exdotoxin stimulation even more serious. the latest studies have discovered seven susceptible molecules correlated with acute exacerbation of hepatitis b: mir- a (down-regulation in severe-type twins patients with hepatitis b), mir- a (up-regulation in severe-type patients with hepatitis b), mir- (up-regulation in severe-type patients with hepatitis b), mir- (providing early warning signal on acute exacerbation of hepatitis b and disease severity), mir- (has obvious decline tendency with the development of liver failure, and can be considered as biomarker for predicting clinical severity and acute exacerbation of hepatitis b), mir- (its expression decreases gradually accompanied with viral clearance, can be considered a biomarker of turnover of acute exacerbation of hepatitis b, and provide early warning signal on acute exacerbation of hepatitis b and treatment), mir- (down-regulation has predictive value to liver inflammation). it has been finally shown that detection of specific mirna correlates with liver inflammation, by using mirna array technique in severe-type patients, can provide novel target on early prediction of severe-type hepatitis. qi zx, et al. observed the difference of th and th cells chromatinization status in il- gene promoter region, which interfered with the binding of promoter and transcription factors, and influenced gene transcription of il- . the distribution of methylation in patients with liver failure showed obvious differences between chb patients and healthy controls; concentrations of serum il- were negatively related with promoter methylation status, which indicated the methylation of il- gene promoter region might involve in the pathogenesis of liver failure, and it is also a reason of il- change. furthermore, some studies revealed that methylation of glutathione-s-transferase p promoter region can promote acute exacerbation of hepatitis b. of hepatitis b toll-like receptors (tlrs) mediated signal transduction pathway includes two forms: myeloid differentiation factor- (myd ) dependent form and myd independent form. tlr , tlr and tlr mediated signal pathway belongs to myd dependent form pathway; tlr mediated signal pathway belongs to myd independent form or trif independent form signal pathway. in the myd independent form pathway, tlrs recognized ligand, then activate nf-κb and mapk, induce release of multiple cytokines, up-regulate costimulatory molecules such as cd , cd , et al., then finally activate specific immune system. tlrs and tlr in particular, play important roles on the mechanism of acute exacerbation of hepatitis b, which can better predict tendency of the disease, and can be considered as indicators for evaluating prognosis of severetype hepatitis b. tlr is a key receptor in endotoxin transmembrane signal transduction; it is also a rate limiting factor in lps transmembrane signal transduction of mononuclear cell and macrophage. some studies demonstrated that the expression of tlr mrna in severe-type patients with viral hepatitis was higher than the healthy controls; and it is higher during the aggressive stage than the critical stage in the nonsurvivors and it is lower during improvement than the critical stage in the survivors. furthermore, expression of tlr mrna is positively correlated with the levels of endotoxin. deng m, et al. observed that the severe-type patients with viral hepatitis had tendency to endotoxemia, with endotoxin binding with tlr , then transmembrane signal activation of nf-κb through intracellular structure of the tlr , further induced release of inflammatory factors, such as tnf-α and il- , and produce secondary intrahepatic microcirculation disturbances. tlr is a pattern recognition receptor with extensive pathogen recognition abilities. tlr can recognize cell walls from multiple bacteria such as gram-positive bacteria, gram-negative bacteria, fungi, spirochaetes, mycobacterium tuberculosis and mycoplasma, and play important roles on acute and chronic infection. yan cg, et al. observed the expression of tlr of mouse with fulminant liver failure induced by d-gal/lps. they observed the high and persistent expression of tlr mrna, and accompanied with high expression of tnf-α, which indicated that tlr can up-regulate expression of downstream inflammatory response genes and release of cytokines, to involve in acute liver failure induced by d-gal/lps. metabolomics is a new technology, which studies various changes of endogenous metabolic products and biologic conditions in humans by observing biosystem changes qualitatively and quantitatively under various stimulations and changes of internal and external environment. the study object is the micromolecule compound with relative molecular mass < in the metabolic network. at present, the major techniques include nuclear magnetic resonance spectrum (nmr), gas chromatogram (gc), liquid chromatography (lc), capillary electrophoresis (ce), mass spectroscopy (ms), infrared spectrum and ultraviolet spectra. of them, nmr, gc-mc and lc-mc are used more extensively. yang et al. investigated the serological profile in acute hepatitis b patients with sudden aggravation of liver function by hplc-ms and pls analytical techniques. they observed the changes in individual atoms in lysophosphatidyl choline (lpc) and gcdca, which can predict early disease aggravation. feng b, et al. analysed the metabolic product in serum of rats with fulminant liver failure by gc-mc and pca software, and observed high content of -hydroxyindole acetic acid, glucose, β-hydroxybutyric acid and phosphate, which indicated the changes of these substances might become potential biomarkers of fulminant liver failure. yu, et al. set up a research platform based upon liver failure metabolomics by using gas chromatographic mass spectrometry (gcms) and super-high performance liquid chromatogram mass spectrometry technology, constructed a model for evaluating clinical severity of liver failure, and constructed serum specific metabolism spectrum by partial least squares discriminant analysis, which can be used to predict the prognosis, with sensitivity of . %. mao et al. also derived similar results with a high positive diagnostic rate of . % for liver failure, and also found that the changes of climax concentration of plasma citrate had potential diagnostic significance. li lj, et al. pointed out the low level of lysolecithin, high level of fatty amide and nonelevation of bile acid by using metabolomics were risk factors for predicting poor prognosis in liver failure patients with pretreatment and post-treatment by artificial liver support systems. by observing the trend changes of serum metabolic spectrum in patients with pretreatment and post-treatment with an artificial liver, we can predict the prognosis and assess curative effect of artificial liver. electrolyte disturbances were usually complicated in patients with liver failure because the patients typically have low renal blood flow and glomerular filtration rate (gfr). a majority of patients manifest hypokalemic, hyponatremia, hypomagnesiumemia and hypocalcemia. the occurrence of hyponatremia and its severity are positively related with prognosis in patient with liver failure. severe hyponatremia can cause acute low sodium syndrome and encephalopathy; and hyponatremia can also cause renal injury and promote or aggravate development of hepatic encephalopathy and hepatorenal syndrome (hrs), and increase mortality. some studies indicated that the mortality of the patients with severe hyponatremia (< mmol/l) is extremely high and up to . %. in patients with acute liver failure, hypochloraemia can be caused by insufficient intake of chlorine, vomiting, large excretion of aldosterone and acidosis, while water intake insufficiency, renal failure, renal tubular acidosis and potassium chloride overdose can cause hyperchloraemia inversely. furthermore, hypophosphatemia is more readily detected than hypocalcemia in most patients. acid base disorders (abd) are very common in patients with acute liver failure, especially alkalemia. li xm, et al. presumed that arterial carbon dioxide pressure depression and respiratory alkalosis are frequent manifestations of acid base disorders. abd is not related with basic pathological changes of severe-type hepatitis, whilst blood ph value is an important factor influencing the survival outcome of severe-type patients with hepatitis. in the latest years, some scholars think that pre-estimation of compensatory formula (pcf) for abd has practical value on differential diagnosis of partial simple and duality abd. partial patients with acute liver failure complicated by abd can be diagnosed by anion gap (ag), potential hco −· and cl − . elevation of ag is a reliable indicator of appearing potential metabolic acidosis, and is common in patients with hepatic encephalopathy, lactic acidosis caused by hypoxemia. during hepatorenal syndrome since reduction of acidum excretion of kidney ketoacidosis may appear. potential hco −· is the hco −· value exclusive of the masking hco −· by high ag metabolic acidosis, which can raise the existence of ag metabolic acidosis and metabolic alkalosis with triplicity abd. a mixed metabolic acidosis can be discerned by combination of ag and blood ci. series of primary abd can overlap, even to develop duplex and triplication abd. for example, if the respiratory alkalosis prolongs, accompanied with increasing of potassium excretion by renal, and with iatrogenic factors to develop metabolic alkalosis, and further develop multiple complications combined with metabolic acidosis, which finally develop triplicated abd. triplication abd is usually the end stage of acute liver failure, with worst prognosis. so, prevention and active treatment of abd is an important parameter for rescuing patients with acute liver failure. liver biopsy is a good method to acquire pathologic changes of liver in multiple liver diseases including liver failure, which have important value on directly evaluating the severity of liver damage. liver biopsy is generally divided into percutaneous biopsy, transvenous biopsy, surgical/laparoscopic biopsy and plugged biopsy. of them, percutaneous biopsy is safe and simple, without special instrument or equipment, and being used extensively. transvenous biopsy is more suitable to the patients with large amount of ascites, clotting dysfunction, diminution of liver volume and increasing of liver hardness because of cirrhosis, difficulty to determine hypochondria because of obvious obesity, and with high hepatic vein pressure. the contraindication of liver biopsy are as the follows: ( ) excessively prolonged pt, excessive declining of pta, which might cause haemostasis difficult even mild injury; ( ) obvious cavity bleeding or large area of petechia; ( ) patients with unconsciousness, who might not be able to fit the operation; ( ) medium dose ascites, especially patients complicated by abdominal infection; ( ) patients with obvious pleural effusion and severe heart and lung disease. it has been proved that liver biopsy is safe to a great extent under careful preparation, attentive operation, even in the liver failure patients with obvious clotting dysfunction. what we need to point out is that although liver biopsy has important value on recognizing liver tissue degeneration, necrosis and degree of inflammatory infiltration, the pathologic change of the biopsy tissue might not be able to reflect entire pathologic change of the liver because of the limited liver tissue obtained. so, it is very important for clinicians to collect and combine various forms of clinical data, laboratory and pathogenetic results from the patients, for getting an accurate judgment. the incidence of cerebral edema in severe-type patients is high, with ratio of to %, and a quarter of them can appear cerebral hernia with poor prognosis. so, measurement of intracranial pressure plays important role on diagnosis and treatment of cerebral edema. traditional lumbar puncture manometry can measure intracranial pressure only one time; it cannot be observed that the changes of intracranial pressure dynamically and exactly. on the other hand, lumbar puncture on patients with acute intracranial hypertension might cause or aggravate cerebral hernia. furthermore, under the condition of cerebral hernia, cranial cavity and spinal cord cavity can not intercross, so the pressure detected by lumbar puncture cannot represent actual intracal pressure. so, persistent detection of intracranial pressure retrieves shortage of lumbar puncture, and cerebral ventricle catheterization is still an extensively used method, which is considered as gold standard of intracranial pressure monitoring by clinicians. intracranial hypertension is defined as persistent elevation of intracranial pressure with levels above mmhg. according to the difference of intracranial pressure, intracranial pressure can be divided into four stages: ( ) normal: ~ mmhg; ( ) mild: ~ mmhg; ( ) moderate: ~ mmhg; ( ) severe: above mmhg. if intracranial pressure is close to mean arterial pressure (map), which illustrates almost no brain perfusion, with nearly close to brain death. currently, mmhg is defined as the critical value of applying treatment of anti-intracranial hypertension. complications of intracranial pressure measurement include infection, intracranial hemorrhage, iatrogenic acute intracranial hypertension and brain parenchyma injury. although laboratory parameters can reflect pathological changes and liver functions during the clinical course of severe-type hepatitis and acute exacerbation of hepatitis b sensitively and in time, and can provide detailed reference data for clinical typing, grading and curative effect evaluation, in total, the extensive and large numbers of indicators listed above are still under study and experimental evaluation, and their availability and utility need to be checked by more clinical practice. until now, laboratory parameters have been extensively used and applied clinically but are still limited. the laboratory parameters conventionally tested in the majority of hospitals are mainly pta, inr, serum bilirubin, alt/ast, pre-protein and blood ammonia. furthermore, some studies also reveal that serum afp, serum sodium and phosphate are closely related with prognosis of liver failure. although some indicators such as arterial blood lactic acid, pct, nagl and inos are closely related with severe-type hepatitis or liver failure, they also score as abnormal in other severe diseases or conditions such as septic shock and mods, so these parameters are not specific for severe-type hepatitis b and liver failure. it is the same on the indicators related with immunity or inflammatory cell or factors, with high sensitivity, while without specificity. other indictors related with heredity, metabolomics, genomics, proteomics and glycomics are still in research, or needing expensive instruments, and still difficult to the routine clinical practice in the short term. what we need to point out is that the various kinds of prognosis scoring systems and mathematical models constructed and tested in the latest years, such as royal school of medicine standard in united kingdom, model for end-stage liver disease (meld) and child-pugh scores, have higher accuracy and utility on predicting prognosis of severe-type hepatitis and liver failure, although some of them still need additional supplement and improvement in the future. because of diversity and individualization for particular causative factors, clinical types, clinical course, complications and clinical treatments, until now, there are still many challenges reflecting prognosis of liver failure. it is believed that more and more novel biomarkers will come into the clinic in the near future, and with development of studies of mechanism and clinical characteristics on acute exacerbation of hepatitis b, therapeutic efficacy of severe-type hepatitis and liver failure will be improved. pathogenesis of aechb is generally considered to include viral factors (hbv genotype, viral replication and viral mutation, etc.), host factors (host genetic characteristics, immune injury, cell apoptosis and necrosis) and interaction between these two aspects. currently, there are at least ten different hbv genotypes and several subtypes distributed all over the world (see table . ) based on the differences of whole genome sequences > % for the major genotypes or - % at the sub-genotype level. genotype a is mainly distributed in northern europe (a ), west africa (a ), and sub-saharan africa (a ). in asia, genotype b and c are most common. genotype b have six subtypes (b -b ), in which, b was found in japan, b ~ b were found in east asia, and b was found mainly in the arctic regions, such as alaska, northern canada and greenland. genotypes b -b are also regarded as recombinants with genotype c. genotype c, including five subtypes (c ~ c ), is mainly distributed in east asia and southeast asia. genotype d, including subtypes d ~ d , prevails in africa, europe, mediterranean and india. genotype e is only found in west africa. genotype f, having four subtypes, is found in central and south america. genotype g has been reported popular in france, germany and the united states. genotype h is found in central america [ ] [ ] [ ] . genotype i was recently newly found in vietnam and laos, but this new genotype is under controversial [ ] . the newly found genotype j in japan has a close relationship with the orangutan's genotype and human genotype c. population migration, promotion of antiviral therapy and host immune selection pressures result in increased risk of hbv gene recombination or mutation [ ] . with the development of genetic testing methods, even more new genotypes could still discovered. due to the distribution differences of hbv genotypes, the study on the relationship between hbv genotypes and severe hepatitis b is quite limited. comparisons have only been conducted in a few genotypes. a study from united states on patients with hbv related acute liver failure suggested that outcomes of these patients were not associated with genotypes [ ] . a multicenter study from japan involving patients with acute hbv infection has compared genotypes (ae [a ],ba [b - ],bj [b ],ce [c ],cs [c ],dandg). multi-factor regression analysis indicated that subtype bj(b ) is one of the independent predictors for fulminant hepatitis. subtype ae(a ) is more related to hbv persistence but not self-limiting hepatitis b [ ] . another multicenter study from japan showed that patients with genotype c accounted for and % in patients with acute-on-chronic liver failure and acute liver failure, respectively. these rates are much higher than that in patients with chronic infection, suggesting that patients with genotype c are more likely to suffer from fulminant liver failure [ ] . a study from china included hbv carriers, patients with chronic hepatitis b, patients with aclf, hbv related liver cirrhosis and patients with hbv related hcc. data showed that genotype b ( . %) and c ( . %) were the main genotypes in these patients. compared with genotype b, genotype c was seen more frequently in those with severe liver diseases, was accompanied with high levels of hbv replication, indicating that genotype c is associated with high hbv replication and severe liver disease [ ] . however, results from another study showed no difference in genotype composition among patients with chronic hepatitis b and those with chronic severe hepatitis [ ] . in a study involving hbv-infected pediatric patients, in which patients had been treated with nucleos(t)ide analogues, genotype c and b were found to be the most prevalent subtypes ( . and . %). compared with genotype b , genotype c is more likely to cause severe hepatitis in hbeag positive pediatric patients [ ] . the association between various genotypes and the pathogenesis of severe hepatitis needs further studies. hbv uses reverse transcription to copy its dna genome and lack of proof-reading capability permits the emergence of mutations in the genome. every day, approximately viral particles are produced and released to maintain a stable level of virus in blood. the average mismatch rate of hbv polymerase is from : to : , potentially resulting in a large amount of mutants in the circulation [ ] [ ] [ ] [ ] . for single individuals infected with hbv, hbv genome mutation either naturally occurs naturally or is selected out by antiviral drugs or the change of internal host environment. the probability of hbv mutations varies in different regions of the whole genome. generally, mutations are more likely to occur in the basal core promoter (bcp), pre-c region and neutralization determinants of the viral envelope, but they can also be found in other regions. some of these mutations have important clinical significance, but most of them are silent mutation without biological significance. pre-c region encoding hbeag, is composed of amino acids. the most common mutation is a guanine (g) to adenine (a) substitution at nucleotide , which creates a premature stop codon at codon , and prevents the translation of the hbeag [ ] . although synthesis of hbeag is inhibited, the hbv replication still continues, manifesting as hbeag negative hbv infection. hbeag expression is not necessary for viral replication, its role in the hbv life cycle remains unclear. in the immune system, hbeag may act as bait, which induces immune tolerance, especially in the newborn babies whose mothers have high level viremia. the hbeag-induced immune tolerance can prevent the attacks on the virus-infected hepatocytes by ctl on hbcag, thus the hbv-infected hepatocytes are not able to be cleared. the hindered synthesis of hbeag may facilitate the ctl to damage infected liver cells, which might be one of the mechanisms of severe hepatitis. it is reported in japan that patients, especially children, infected with this mutant, are more likely to suffer from severe hepatitis. other pre-c mutations include point mutations generating other termination codons, for example g a generating uga codons. point mutations may change the initiation codon of p or the specific amino acid for cleavage and insertion of key signal peptide, particularly between and . all of these mutations can affect the hbeag production. the pre-c mutation can reduce hbeag expression, increase hbv replication and aggravate liver damage [ ] . a number of studies have suggested the association of pre-c mutation and development of severe hepatitis. the hbcag encoded by c region, contains a t-cell-dependent/independent epitopes and induces the host humoral and cellular immune responses. the c gene mutation may cause deletion of the cell surface antigen, and consequently the lack of humoral immune response by the host against hbv. however, the cytotoxicity of ctls was not affected resulting in potentially massive necrosis of liver cells and eventually leads to severe hepatitis. core promoter directly activates pgrna transcription, and plays a central role in hbv replication. core promoter is composed of basic core promoter (bcp) and its upstream regulatory elements. the bcp region partly overlaps with the 'end of x gene and the ' end of pre-c gene and can independently start the transcription of pre-mrna and pgrna. the core promoter mutation, which is associated with nt - that are at the upstream of the starting point of pre-c mrna, reduces the synthesis of hbeag, but has no effect on hbcag. the development of severe hepatitis is often accompanied with hbv core promoter mutation, especially the , double set of mutations [ ] . other common mutations in the core promoter region of patients with severe hepatitis include nt c-t, nt t-a and cluster mutations, including nucleotide insertion, deletion and substitution. the bcp mutations can induce changes of two codons (l m and v i) in x protein, and generate the hnf (hepatocyte nuclear factor ) binding sites. insertion of bases into the core promoter produces a new binding site of hnf , which can enhance viral replication and lead to severe hepatitis [ ] . in-vitro studies have confirmed that bcp trans-activating transcription causes x protein replacement, which downregulates the transcription of pre-c region and pgrna [ ] . however, the double-mutant can upregulate the transcription of pgrna, increase hbcab production, thereby enhancing viral replication. previous studies have shown that a t and g a mutation are more prevalent in hbv genotype c, which partially explains the stronger pathogenicity of genotype c than genotype b. recent studies have shown that a t and g a mutations in hbv genotype b may be associated with severe hepatitis, but in hbv-infected pediatric patients, a t and g a mutations show no significant difference in genotype b and c. overall though, compared with wildtype hbv, bcp double mutation is more commonly associated with severe liver diseases, especially liver cirrhosis and hcc [ ] . the mutations in pre c/bcp region may alter the biological characteristics of the virus, and induce the development of severe hepatitis through impacts on host immune responses, being more vigorous in hbeag-negative chb [ ] . however, the biological significance of mutations in different sites or forms, the dynamic interaction between virus mutation and host immunity, the influence of different genotypes and viral quasispecies on mutation are still not fully elucidated. immune evasion or vaccine failure related pre-s point mutation or deletion mutation does not affect the viral replication. the pre-s , s recombination, including deletion mutation and promoter mutation, have been regularly found in patients either with chronic or fulminant hepatitis. these mutations frequently occur after hbeag seroconversion or interferon treatment, suggesting the host immune selective pressure during their selection. the pre-s region mutations may play a role in hbv persistent infection, and may also cause liver damage. it has been reported that the pre-s mutation is related to fulminant hepatitis, and the pre-s mutation is a strong risk factor for the development of hbv-related liver cancer [ , ] , presumably as a result of an accumulation of viral envelope proteins inside the cell.mutations of the surface antigen protein, particularly aa mutation (sg r), result in conformational change of the major antigen epitope 'a'. this change disables the immunological recognition and surveillance of the host immune system for the mutant can also result in failure of the clinical vaccination and might be one of the precipitations for the exacerbation of hepatitis b [ ] . the p gene mutation in key catalytic domains indicates the hbv resistance against nucleos(t)ide analogues (na). the current five nas in clinical application include lamivudine, adefovir, telbivudine, entecavir and tenofovir. the clinically extensive application of nas leads to rapid selective drug resistance. the selection of drugresistant mutation depends on the following factors: ( ) the long half-life of hepatocytes and intrahepatic cccdna; ( ) the capacity of hbv replication and mutation; ( ) antiviral drug pressure; ( ) genetic barrier to resistance. for example, the lamivudine resistance is closely associated with the hbv reverse transcriptase gene ymdd motif mutation, presenting as rt m i and rt m v mutation, with or without rt l m mutation. the replication capacity of rt m i/v mutants is weaker than the wild strains in the absence of drug. the rtl m mutation can restore the hbv replication capability. in addition, the single or combined mutation of rt l i, rt l m, rt f l, rt v l, rt a v and rt v i may compensatorily restore the replication capability of rtm i/v mutant [ ] . the resistance rate is up to % after -years of lamivudine therapy, and more than % after -year mono-therapy. the -year resistance rate of telbivudine is about %. the rtn t site mutation (threonine substitute asparagine) is mainly seen in adefovir resistance, and the -year resistance rate is about %. the rta t/v (valine/threonine substitution alanine) site mutation is found in all the above three antiviral drugs. entecavir resistance mutations include and substitutions, combined with substitutions at codons , and . the -year resistant rate in patients initially treated with entecavir is less than %. for patients that have been previously treated with lamivudine, the resistance rate of entecavir increased significantly. so far, there have been no reports on the primary drug resistance for tenofovir mono-therapy. the rta t mutation was found in the combination therapy of tenofovir and lamivudine [ ] . the entire s gene is included in the p gene, and the rt region overlaps with s gene, thus the rt mutation may cause the s gene mutation. double mutations in these two regions can change the viral replication capacity. the hbv drug-resistance mutation occurs in patients with chronic hepatitis b may reduce the hbeag seroconversion rate, reverse the histological improvement, increase the disease progression rate, aggravate liver cirrhosis, and increase the death risk of liver transplant patients. rt mutations may also cause the s protein epitope changes and affect the hbs antibody and the function of ctl, suggesting its role in the development of liver failure [ , ] . as salvage therapy for lamivudine resistance, lamivudine combined with adefovir dipivoxil has higher rate of viral suppression and lower rate of adefovir resistance compared with switching to adefovir dipivoxil monotherapy [ , ] . tenofovir is a potent antiviral drug for lamivudine-resistance salvage therapy, and showed a better effect than switching to adefovir dipivoxil monotherapy [ ] . in contrast, switching to entecavir is not an optimal choice for lamivudine-resistance [ ] . telbivudine resistance is associated with the rtm i mutation, and has crossresistance with lamivudine. therefore, telbivudine could not be an alternative for lamivudine-resistance [ ] . treatment for telbivudine resistance is similar to that for lamivudine resistance [ ] . treatment for adefovir-resistance is determined by the virus mutation types and antiviral medication history [ , ] . lamivudine has been proved to be effective on inhibiting rtn t adefovir resistance mutations [ ] . in vitro data has also suggested the effectiveness of telbivudine. additionally, entecavir might be a reasonable choice for rtn t mutants. patients with rtn t mutation are suggested to ( ) switch to or add entecavir; or ( ) add lamivudine or telbivudine; or ( ) switch to tenofovir. compared with that on hbv wild strain, lamivudine becomes less effective on a v adefovir resistant strain. in vitro studies show that tenofovir has reduced sensitivity to the rta t mutation. clinically, entecavir and tenofovir can effectively inhibit the replication of a t adefovir resistant mutants [ ] . patients with rta t mutation are suggested to ( ) switch to or add entecavir; or ( ) switch to tenofovir. under this circumstance, lamivudine should not be suggested in case it increases the risk of cross-resistance [ , ] . currently, there has been no data from large sample clinical trials that can guide the treatment for entecavir resistance. in-vitro studies and case reports suggested that adefovir and tenofovir are effective for entecavir-resistant patients. based on expert opinions, patients with entecavir resistance are recommended to add tenofovir or adefovir [ ] [ ] [ ] . in summary, the virus mutation in above regions may be associated with the pathogenesis of severe hepatitis. however, the severity of hepatitis depends on key factors of virus and host. the same mutant may lead to different clinical outcomes in different hosts. thus except for virus mutation, factors including host immune status, cytokines and hla might also account for the development of severe hepatitis. tests for hbv infection hbv surface antigen/pre-s protein/pre-s protein hbsag is the major coat protein of hbv with antigenicity but not infectivity. in a broad sense, hbsag contains the major protein, middle protein and large protein. narrowly, hbsag simply refers to the major protein, which appears earliest and has the highest titer. thus it is considered as an important marker for early diagnosis of hepatitis b. for typical acute hepatitis b, hbsag appears during the incubation period, followed by the clinical symptoms and abnormal liver function in - weeks. hbsag stays in the blood for - months, and disappears in the recovery period. persistence of hbsag more than months indicates the development of chronic hepatitis. hbsag can also be detected in hbv carriers and patients with hbv-related liver cirrhosis or liver cancer. a rapid reduction of quantitative hbsag within months can predict the efficacy of antiviral drugs, but no changes in the quantitative hbsag level after months is not considered as a good predictor [ , ] . pre-s protein, which appears early and is significantly associated with hbeag and hbv dna, can be used as a marker of acute hepatitis b early in infection. the pre-s protein has a strong immunogenicity, and includes the important site where hbv attaches to and invade the hepatocytes, the sodium taurocholate co-transporting peptide (ntcp). it is also a reliable reflection of hbv replication level. the synergy from pre-s protein is also important for hbv invasion. most patients with acute exacerbation of chronic hepatitis, or chronic active hepatitis, or acute hepatitis developing to chronic hepatitis have persistent pre-s protein expression in serum. therefore, serum pre-s protein can be used to estimate the activity and infectivity of hbv in chronic patients [ ] . pre-s and pre-s protein, which can induce and regulate humoral and cellular immune response of the host, provide important immune defense for eliminating virus in blood circulation and preventing healthy liver cells being infected. hbsab, which is a protective antibody, can eliminate the virus and prevent hbv infection. hbsab appears in the late stage of acute infection, just before hbsag becomes negative, and will gradually rise to the peak levels in - months. this antibody can last for a long time, but the titers will gradually decline after years. a small number of cases do not produce hbsab after hbsag becomes negative. in acute hepatitis b infection, appearance of hbsab suggests recovery of the disease. patients with severe hepatitis often present with high titers of hbsab, which forms the immune complexes with hbsag, which can induce flares of hepatitis that lead to liver cell necrosis. after the hepatitis b vaccination, hbsab (> iu/l) means the successful vaccination and development of immunity. in the blood, hbcag is mainly located in the core of the dane particles or virions. the only small amount of free hbcag is also combined by high titers of hbsab and presents as immune complexes. thus, it cannot be detected unless treated by detergent. hbcag on the surface of hepatocytes is considered to be the main target antigen of the host ctls. hbcag is a direct evidence of hbv infection and replication, and also a marker for evaluating the efficacy of antiviral drugs. hbcag is strongly immunogenic, so that hbcab can be detected in most patients with hbv infection. hbcab often emerges in the early stage after infection, is present in high titer in blood and can persist for a very long time. titer of hbcab above : , together with abnormal alt level, can be used for the diagnosis of hepatitis b infection. for occult hepatitis b, high titer of hbcab is also valuable for the diagnosis. hbcab consists of igm and igg antibodies. anti-hbc-igm, which suggests hbvresulted liver damage, is the main evidence for the diagnosis of acute hepatitis b. it may become positive during the active phase of chronic hepatitis b and turn negative during remission. it also appears during the flares of chronic hepatitis b, mostly in a week after infection, and disappears within months. anti-hbc-igg appears late but is sustained for many years or even a lifetime. in patients with acute hepatitis b, the titer of anti-hbc-igm is higher than anti-hbc-igg, while in those with chronic hepatitis b the situation is opposite. both antibodies show high titers in fulminant hepatitis. hbeag is a soluble antigen, which appears later than hbsag. sustained expression of hbeag suggests persistence of hbv infection. in patients with chronic hepatitis b, hbeag acts as an important immune tolerance factor leading to a low immune response to hbv infection. it is a valuable marker for evaluating the efficacy of antiviral drugs. seroconversion refers to the loss of hbeag and development of anti-hbe. approximate - % patients have spontaneous seroconversion every year. studies have shown that spontaneous seroconversion occurs earlier in patients with genotype a, b, d and f than those with genotype c. appearance of hbeab demonstrates the decrease or termination of viral replication and low infectivity. recent studies showed that, after year since the spontaneous hbeag seroconversion, viral load more than iu/ml increased the incidence of fulminant hepatitis. for hbv chronic carriers and patients with hcc, hbeab does not mean the recovery, or elimination. in contrast, hbv dna integration is often found in these patients. x protein is capable of transactivating the expression of numerous cellular and viral genes, and is vital for virus replication. x protein can be detected in some patients with chronic hepatitis, so it is used as an auxiliary diagnostic marker of hbv infection [ ] . x protein plays a central role in hbv-related hcc progression and stimulation. thus, follow-up is necessary for patients active and persistent hbv replication. serum hbv dna is the direct evidence of active hbv infection, reflecting the level of viral replication and infectivity. the quantitative detection for viral genes is a very important marker for treatment decision, efficacy prediction and observation. long-term high load of hbv dna is an independent risk factor for predicting the development of liver cirrhosis and hcc. numerous studies have shown that viral load is the most reliable marker to predict the development of hcc [ ] . the differences of hbv whole genome sequence are approximately - %. the diverse variants that are genetically linked through mutation are known as quasispecies. quasispecies contain a large number of mutated genes serving as a reservoir for viral selection under the pressure of immune response and antiviral treatment. when changes occur in the environmental conditions, the quasispecies structure responds by rebalancing its composition. the predominant sequence may shift by selection of a variant that is better adapted to the new environment, in the classic darwinian process of survival of the fittest [ ] . this feature gives the virus strong adaptability, and makes it difficult to prevent and control. study on the relationship between quasispecies and different clinical outcomes will provide valuable information for exploring anti-hbv treatment strategies. . quasispecies evolution under host immune pressure. in chronic hbv infection transmitted via perinatal transmission, the different immune phases of chronic hbv infection confer different environments on hbv quasispecies. thus, the characteristics of hbv quasispecies may differ. a preliminary study on the differences of full-length hbv quasispecies between mothers and their progeny showed that, after years of evolution, the dominant sequence of hbv quasispecies became different between mothers and daughters. the characteristics of hbv quasispecies in various gene regions are different in mothers and daughters with different treatment responses or disease status. among these genes, the prec/c gene had the highest substitution rate [ ] . under antiviral drug selection pressure, hbv mutants are selected from the preexisting pool of quasispecies and over time become the dominant species [ ] . the probability of resistant mutations depends on the effectiveness of antiviral drugs. low potent drugs have almost no selective pressure on the virus, thus lead to low probability of viral resistance; on the contrary, drugs that completely inhibit viral replication also rarely result in resistance. only medium potent drugs have the highest rates of drug resistance. a study showed that during the -week lamivudine therapy, distinct patterns of quasispecies evolution are found between responders and non-responders; the structures of viral quasispecies tended to be simpler in responders, but more complicated (higher diversity) in non-responders. similar phenomenon was also observed during entecavir therapy. another study detected the full length sequence of resistant virus in lamivudine and adefovir sequential therapy, and found that variation of nucleotide or amino acid sequence usually occurs in hbv hbsag or rt region. using single strand conformation polymorphism (sscp) and dna sequence analysis, researchers found that the complexity of hbv quasispecies in patients with cirrhosis was more than those in patients with chronic hepatitis b, suggesting that complexity of hbv quasispecies is associated with disease status [ ] . researchers from china had also used the same methods to analyze the difference of quasispecies complexity in s region among patients with chronic severe hepatitis b, patients with chronic hepatitis b and hbv carriers. it is found that the quasispecies complexity in the s region increases along with disease progression in chronic hbv infection. analysis on quasispecies in acute hepatitis b, chronic hbv carriers, chronic hepatitis b and chronic severe hepatitis by full-length hbv genomic clone and bioinformatics methods also discovered the positive correlation between hbv quasispecies complexity and disease severity [ ] . on one hand, complex evolution of quasispecies may lead to persistent infection and continuous liver damage, and increase opportunities for the emergence of new hbv variants. on the other hand, enhanced virulence of mutated virus and change of antigen epitope may cause excessive immune response and severe liver damage. however, correlation between hbv quasispecies complexity and disease severity still needs dynamic large sample research to confirm [ ] . the dynamic change of quasispecies during na antiviral therapy may be related to the antiviral efficacy and drug resistance. results from a study on patients receiving lamivudine for weeks suggested that the baseline quasispecies heterogeneity is not associated with antiviral efficacy, and the changes of quasispecies complexity at an early stage may predict antiviral efficacy and drug resistance more accurately than the change of hbv dna level during lamivudine therapy. another study on dynamic changes of quasispecies in patients receiving entecavir antiviral therapy suggested that, compared to the partial responders, quasispecies complexity is reduced but dispersion is increased in complete responders after weeks of treatment [ ] . in these two studies, the quasispecies dispersion decreased in lamivudine responders but increased in entecavir responders after weeks of treatment. this might be because entecavir has stronger antiviral effect than lamivudine, thus generates more selection pressure. in addition, entecavir can still induce complete response even when some mutations occur since it has a higher drug resistance barrier. therefore, the early changes in hbv quasispecies complexity may act as a predictor of sustained antiviral effect of nucleos(t)ide analogues. the resistant strains typically already exist before antiviral therapy, and become the predominant strains under selection pressure of antiviral drugs. a study showed that adefovir treatment for weeks in patients with chronic hepatitis b selected resistant virus strains [ ] . study on gene heterogeneity of hbv reverse transcriptase suggested that lamivudine monotherapy is liable to induce the quasispecies that affect response rate of salvage therapy with adefovir for virologic breakthrough in lamivudine-treated patients, and reduce the sensitivity to other nucleos(t)ide analogues [ ] . under different drug selection pressure, non-responders have similar quasispecies evolution patterns, suggesting that this pattern may be associated with viral resistance mechanisms. from the perspective of quasispecies, drug selection pressure changes the relative ratio of viral populations, and leads to population drift. dna sequence analysis is the most direct and reliable method to detect gene mutations. it is also the gold standard to detect nucleic acid sequence mutation. however, the high cost of this method prevents its application in large sample research. dna sequencing, which can be divided into direct sequencing and cloning sequencing, is built on the basis of high-resolution denaturing polyacrylamide gel electrophoresis. direct sequencing detects the nucleotide sequence of dominant strains; clone sequencing method can find out changes of nucleotide in other strains. rflp is a technique that applies to detect variations in homologous dna sequences. it refers to a difference between samples of homologous dna molecules from differing locations of restriction enzyme sites, which may be naturally formed or brought in by pcr mismatch. the dna sample is broken into pieces and digested by restriction enzymes and the resulting restriction fragments are separated according to their lengths by gel electrophoresis. mismatched pcr is a modified pcr, which changes one or a few nucleotide bases in designed pcr primers to make the synthetic dna meet special requirements. here, a restriction enzyme recognition sequence is introduced into the pcr amplified fragment in order to change the target dna restriction map and distinguish the mutants and non-mutants. nucleic acid hybridisation is the pairing of complementary single-stranded nucleic acids (dna or rna) to produce dna-dna or dna-rna hybrids. when the target dna obtained by pcr amplification combines with the probe labeled by radioactive or non-radioactive labels, any mismatch between the probe and target dna can be detected. this helps to distinguish the wild and mutant strains. nucleic acid hybridization applies to high frequency point mutation and is suitable for large sample detection, but the high requirement for hybridization temperature makes it difficult to popularize. the principle of sscp is based on conformational difference of single-stranded nucleotide sequences of identical length. this property allows sequences to be distinguished by means of gel electrophoresis so as to determine whether mutations exist. it applies to the single base substitution and screening of dna fragment mutation. however, sscp is neither stable nor practical [ , ] . primer extension starts from the ′ end in pcr amplification, and only succeeds when the ′ end of the primer is an exact match to the template dna. based on this phenomenon, the primer with ′ end containing mutated bases is used to detect the mutation of target dna. the ′ base-specific pcr technology is simple but with low sensitivity, and the results are prone to be false-negative. the incomplete block of pcr amplification also leads to false-positive results [ ] . melting curve analysis is an evaluation of the dissociation-characteristics of doublestranded dna during heating, the temperature at which % of dna is denatured is referred to the melting point [ ] . in chronic hbv infection, the peak numbers of dna melting curve in patients with moderate and severe hepatitis are significantly more than those in hbv carriers and mild hepatitis; the peak number of melting curve in patients with severe hepatitis is significantly more than that in moderate hepatitis. the melt-curve analysis is less sensitive than sscp, but is more accurate on analyzing genetic variation. the strong operability and high cost-effective make it a preferable method for genetic variation analysis. the high demand for low-cost sequencing has caused the development of highthroughput sequencing-the next-generation sequencing, which includes massively parallel signature sequencing (mpss), solexa sequencing, solid sequencing, pyrosequencing, heliscope single molecule sequencing, etc. these methods apply to genome sequencing, genome resequencing, rna-sequencing, chip-sequencing and epigenome characterization [ ] [ ] [ ] . current dna sequencing methods under development include microscopy-based techniques, macromolecule and nanotechnology that can distinguish the base signal and directly read the sequence without the use of biological or chemical reagent. the third-generation sequencing methods include: ( ) non-optical microscope imaging: the dna sequence can be read out if the resolution of image is high enough to differentiate the four kinds of bases on dna when visualizing the spatial linear arrangement of nucleotides. this idea is based on non-optical microscope at the atomic level, for instance, scanning tunneling microscopy (scanning tunneling microscope, stm) [ ] . ( ) nanopore dna sequencing: it is based on the readout of electrical signals occurring at the single stranded dna or rna molecules passing by alpha-hemolysin pores covalently bound with cyclodextrin [ ] [ ] [ ] . those methods still need further validation and improvement. the pathogenesis of hbv infection is determined by the interplay between both virus and host. different outcomes after infection are related to different host immune responses and viral mutations. however, the biological significance of viral mutation has not been fully elucidated because of: ( ) the limited samples and lack of comparison between different groups; ( ) the overlap of each hbv genomic coding regions; ( ) the limitation of detection technology. with the rapid development of detection technology, the large sample, long-term, multi-level studies will help to understand more about the host-virus interaction and potential mechanisms. zhi the indications for liver transplantation are changing over time. before , the major indication for liver transplantation was primary malignancy of the liver, especially hepatocellular carcinoma (hcc) which significantly influenced the therapeutic efficacy of liver transplantation and postoperative survival. currently, not only are acute or chronic liver diseases nonresponsive to other medical treatment and surgery now the major indication for liver transplantation but also liver diseases that markedly affect quality of life. the major indications for liver transplantation are shown in table . . viral hepatitis has and continues to be a major public health problem in china. it is the most common liver disease and most common cause of liver failure. the proportion of patients with hepatitis b-induced liver failure among patients receiving liver transplantation in china is significantly higher than in other countries. hepatitis b has diverse clinical manifestations and may progress to chronic hepatitis, liver failure (including acute liver failure, acute on chronic liver failure, and chronic liver failure), hepatic cirrhosis (compensated and decompensated), and even hepatitis b-related liver tumors. whether liver transplantation is necessary for patients with hbv infection and the timing of transplantation must be carefully considered. early liver transplantation may increase patient risk and be an economic burden because patients could recover and even survive for a long time in the absence of liver transplantation. however, presently it is difficult to predict patient outcome to hbv infection. in contrast, if liver transplantation is performed in the late stages of disease, numerous complications of hepatitis b-induced liver dysfunction may either increase the risk of transplant surgery resulting in a poor prognosis. thus, for patients with severe hepatitis b, the indications for and the timing of liver transplantation are crucial and should be carefully evaluated. in general, the following conditions are indications for liver transplantation in patients with chronic hepatitis b: . obvious manifestations of liver failure including sustained elevation of serum bilirubin to > mg/dl; prothrombin time > s longer than the reference range; plasma albumin < . g/dl; and liver failure which is nonresponsive to active and/or symptomatic therapy (such as infusion of fresh plasma and albumin) or if the patient continues to deteriorate clinically despite optimal medical therapy. . when there are complications related to either liver failure or portal hypertension such as the presence of severe hepatic encephalopathy, disturbances of coagulation function, refractory bleeding due to rupture of esophageal or gastric varices, refractory ascites, repeated episodes of spontaneous bacterial peritonitis, or the development of hepatorenal syndrome. . when hepatitis b influences the quality of life including the development of severe lethargy, uncontrollable itching, metabolic bone diseases or the development of bacterial cholangitis and . the development of hepatocellular carcinoma. although there are some well defined indications for liver transplantation, the ultimate determination of the timing and specific indications for liver transplantation remains a challenge in clinical practice for hepatitis b patients with liver failure. the introduction of the model of end stage liver diseases (meld) and some derivative scoring systems are now frequently used to determine whether a patient should be listed for transplantation and the urgency of liver transplantation. in , the mayo clinic proposed that meld should replace the child-pugh grading system for the determination of urgency of liver transplantation. meld is calculated using the following calculation: meld = . × log e (creatinine [mg/dl]) + . × log e (bilirubin [mg/ dl]) + . × log (inr) + . × (causes: for biliary and alcoholic; for others). meld was originally used for the evaluation of prognosis of hepatic cirrhosis patients receiving transjugular intrahepatic portosystemic shunt. as described above, the meld score is determined on the basis of total bilirubin, international standardization of bleeding time, serum creatinine, and etiology. compared to the child-pugh grading system, meld employs objective parameters, which are helpful for the comparisons of patients from different centers. while the meld score changes with the alteration of the liver disease, the child-pugh grade has only three levels and is unable to meet the requirements of accurate and objective evaluation. saab et al. investigated the prognosis of patients receiving liver transplantation and found that the -year survival rate was , , , , and % in patients with preoperative meld scores of ≤ , - , - , - , and ≥ , respectively. however a higher meld score negatively affected the -year survival rate. for example patients with a meld score ≤ had a year survival rate of whereas patient survival was reduced to % in patients with a meld score > . despite the benefits and the widespread application of meld in clinical practice, there remain some imperfections in the evaluation of timing of liver transplantation and the assessment of prognosis and therapeutic efficacy. one of the major imperfections is the failure to consider complications such as infection, hepatic encephalopathy, hepatorenal syndrome, and disturbances in fluid and electrolytes which may significantly influence the prognosis and timing of liver transplantation but are not included in the meld scoring system. hence inclusion of sodium (na) in the meld scoring system: meld − na [meld + . × ( − na)], imeld [meld + ( . × age) − ( . + na) + ]; and meso [meld/na (mmol/l)] × has improved outcomes of patients requiring liver transplantation. those scoring systems, however, also fail to consider other complications. including the development of hepatic encephalopathy or ascites and therefore, their applicability is limited in china. in china, liver transplantation was initiated relatively late compared with other countries; regulations regarding liver transplantation remain imperfect; and smooth communication among transplantation centers in different regions of the country are lacking. these drawbacks markedly influence the timely and fair distribution of donor livers. determination of waiting times for transplantation is not scientific. therefore, the comprehensive evaluation of a patient's condition of hepatitis b-induced liver failure, determination of the timing of liver transplantation, and optimization of the rational and fair distribution of donor livers needs to be carefully considered by chinese clinicians in the field of hepatology going forward if improvements are to occur. there are currently additional scoring systems which have been used by chinese clinicians. one of these was developed by professor ke wm in the department of infectious diseases of the affiliated third hospital of sun yat-sen university. in that system, hepatic encephalopathy, serum creatinine, prothrombin activity, serum total bilirubin, liver size (determined by ultrasonography), amount of ascitic and pleural fluid (determined by ultrasonography), and infection (peripheral white blood cell count, proportion of neutrophils, and inflammatory findings from thoracic imaging examinations) are taken into account and objectively and conveniently scored on a scale of - according to their severities as described in tables . , . , and . . [ ] these newer scoring systems and the meld scoring system can favorably predict the mortality of acute on chronic liver failure patients with hepatitis b. the area under receiver operator curves of scores determined with a new scoring system and meld scoring system was . [ % confidence interval (ci), . - . ] and . ( % ci, . - . ), respectively. there was no overlap in % cis between the two, and they were significantly different (p < . ) as illustrated in figs. . , . , and . ). wenzhou medical college in china subsequently proposed a scoring system in which the total score is calculated using the following equation: x = . + . × hepatorenal syndrome (hrs) + . × lc − . × hepatitis b e antigen (hbeag) − . × alb − . × pta. that scoring system has been used in several centers for liver diseases, and its effectiveness and accuracy have now been confirmed in these centers. in summary, to establish a new scoring system on the basis of china's national status of liver disease and to further validate this system, large multicentered studies with a large number of patients is crucial before they are adopted in china. it is important that any new system developed under these conditions be subjected to rigorous study, standardization and scientific validation. for liver transplantations in patients with hepatitis b-induced liver failure, perioperative therapy is crucial, impacting the recurrence of hepatitis b virus reinfection following liver transplantation. in fact, perioperative therapy can make a major fig. . receiver operator curve of a new scoring system and meld scoring system difference in patients with hepatitis b compared to patients with other liver diseases (e.g., tumor recurrence). it is well appreciated that antiviral therapy is necessary in hepatitis b virus patients as discussed below. the guideline for the prevention and treatment of chronic hepatitis b in china ( ) recommends that ( ) for patients with hepatitis b planning to receive liver transplantation, oral lamivudine be administered within - months before liver transplantation when hepatitis b virus dna is detectable ( mg q h). hepatitis b immune globulin (hbig) should be administered during the anhepatic stage in surgery and long-term use of lamivudine and low-dose hbig ( iu q h for the first week postsurgical, iu q week, then iu q month). the dose of hbig and interval between two treatments [generally, trough hepatitis b surface antibodies (anti-hbs) are > - miu/ml and anti-hbs is better if > miu/ml within months of surgery) should be determined according to the anti-hb levels. for patients nonresponsive to lamivudine, other nucleos(t)ide analogues effective for resistance mutation should be used and for patients with a low risk for recurrence of hbv infection (i.e., hbv dna negative before liver transplantation and absence of hbv infection recurrence within years after liver transplantation), lamivudine plus adefovir should be considered for the prevention of hbv infection recurrence. while patients are waiting for liver transplantation, an artificial liver support system (alss) may be of use as a bridge to transplantation. liver failure may significantly compromise the detoxication, synthesis, and metabolism activities, resulting in a significant accumulation of toxins and deficiency of some important factors (such as coagulation factors). unfortunately, some patients die while waiting for either their liver function to improve or for liver transplantation. alss have been generally used to partially substitute the liver's normal activities, such as clearing some toxins and supplement some missing factors, which is helpful for life maintenance while awaiting subsequent liver transplantation or spontaneous recovery. thus, for patients with liver failure secondary to hepatitis b, alss maybe an important therapeutic strategy while patients are waiting for liver transplantation. alss can be classified as a mechanical artificial liver, biological artificial liver, or mixed artificial liver. mechanical alsss mainly utilizes nonbiological materials to clear toxins in the body and supplement some missing factors. biological alsss use biological materials to substitute the liver's activities. the core feature of the biological alss systems is to simulate hepatocyte function. however, the source and biosafety, as well as their location (bioreactor), are important factors limiting the development of biological alsss. finally, mixed alsss employ the hemodiafiltration, plasma exchange, and hemoperfusion that are able to detoxify substances (nonbiological alss) and human or porcine "hepatocytes" in the bioreactor. currently, nonbiological alsss are the most widely used in clinical practice, and biological and mixed alsss are still undergoing investigation and have not been widely applied in clinical practice. in addition to its use before liver transplantation, alss post liver transplantation may be used to correct ongoing renal failure, brain edema, severe water and electrolyte disturbances, and severe infections. another factor to consider is that liver donors are rare. as a result, the inability to perform liver transplantation in a timely fashion results in reduced survival rates. under those conditions, pre transplant complications including multi organ failure may persist. in such cases, alss therapy may prove to be beneficial. for patients with hepatitis b-related liver failure, perioperative therapies are more important than in patients receiving elective liver transplantation (such as liver tumor patients without liver failure). liver failure causes a variety of complications that directly contribute to transplantation failure and a high-risk status after surgery. clinicians need to address problems such as portal hypertension, upper gastrointestinal bleeding, severe jaundice, ascites, spontaneous peritonitis, hepatic encephalopathy, hepatopulmonary syndrome, portal pulmonary hypertension, kidney dysfunction, hepatorenal syndrome (among others) through active treatment/management in specialized liver units. preserving liver function and maintaining a normal physiological status have been shown to reduce the risk of therapeutic failure in patients receiving liver transplantation due to hepatitis b-related 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from immunosuppressed patients with severe liver disease probable implication of mutations of the x open reading frame in the onset of fulminant hepatitis b association between the various mutations in viral core promoter region to different stages of hepatitis b, ranging of asymptomatic carrier state to hepatocellular carcinoma number of mutations within ctl-defined epitopes of the hepatitis b virus (hbv) core region is associated with hbv disease progression pre-s defective hepatitis b virus infection in patients with fulminant hepatitis associations between hepatitis b virus mutations and the risk of hepatocellular carcinoma: a meta-analysis potential threat of drug-resistant and vaccine-escape hbv mutants to public health early changes of hepatitis b virus quasispecies during lamivudine treatment and the correlation with antiviral efficacy hepatitis b virus variant with the a t substitution within reverse transcriptase before and under adefovir and tenofovir therapy restoration of 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cross-resistance profile than the nucleotide analog inhibitors adefovir and tenofovir management of treatment failure in chronic hepatitis b resistance to adefovir dipivoxil therapy associated with the selection of a novel mutation in the hbv polymerase selection of a hepatitis b virus strain resistant to adefovir in a liver transplantation patient susceptibility to antivirals of a human hbv strain with mutations conferring resistance to both lamivudine and adefovir impact of hepatitis b virus rta v/t mutants on hepatitis b treatment failure management of drug-resistant chronic hepatitis b treatment of patients with lamivudine-resistant and adefovir dipivoxil-resistant chronic hepatitis b virus infection: is tenofovir the answer? successful treatment of an entecavir-resistant hepatitis b virus variant how to diagnose and treat hepatitis b virus antiviral drug resistance in the liver transplant setting prevention and management of drug resistance for antihepatitis b treatment is there an association between the measurement of qualitative hbsag and virologic response in chronic hbv infection serum hepatitis b surface antigen quantitation can reflect hepatitis b virus in the liver and predict treatment response serological and molecular diagnosis hbv induced hcc: major risk factors from genetic to molecular level new insight in the pathobiology of hepatitis b virus infection quasispecies structure, cornerstone of hepatitis b virus infection: mass sequencing approach clinical implications of evolutionary patterns of homologous, full-length hepatitis b virus quasispecies in different hosts after perinatal infection hepatitis b virus mutations associated with antiviral therapy hepatitis b virus s gene mutants in a patient with chronic active hepatitis with circulating anti-hbs antibodies hepatitis b virus genomes of chronic hepatitis patients do not contain specific mutations related to acute exacerbation molecular virology and the development of resistant mutants: implications for therapy evolutionary patterns of hepatitis b virus quasispecies under different selective pressures: correlation with antiviral efficacy complex dynamics of hepatitis b virus resistance to adefovir emergence of hepatitis b virus quasispecies with lower susceptibility to nucleos(t)ide analogues during lamivudine treatment optimization of nonradioisotopic single strand conformation polymorphism analysis with a conventional minislab gel electrophoresis apparatus evaluation of the genetic variability of orchid fleck virus by single-strand conformational polymorphism analysis and nucleotide sequencing of a fragment from the nucleocapsid gene an improved allele-specific pcr primer design method for snp marker analysis and its application product differentiation by analysis of dna melting curves during the polymerase chain reaction next-generation sequencing in aging research: emerging applications, problems, pitfalls and possible solution next-generation dna sequencing methods applications of next-generation sequencing technologies to diagnostic virology fast dna sequencing via transverse electronic transport single-nucleotide discrimination in immobilized dna oligonucleotides with a biological nanopore rapid sequencing of individual dna molecules in graphene nanogaps detecting single stranded dna with a solid state nanopore peri-operative therapies of patients receiving liver transplantation due to severe hepatitis establishment of a scoring system for evaluating the severity of hepatitis b patients with acute-on-chronic liver failure key: cord- - ioiwsd authors: varghese, praveen mathews; tsolaki, anthony g.; yasmin, hadida; shastri, abhishek; ferluga, janez; vatish, manu; madan, taruna; kishore, uday title: host-pathogen interaction in covid- : pathogenesis, potential therapeutics and vaccination strategies date: - - journal: immunobiology doi: . /j.imbio. . sha: doc_id: cord_uid: ioiwsd abstract the current coronavirus pandemic, covid- , is the third outbreak of disease caused by the coronavirus family, after severe acute respiratory syndrome and middle east respiratory syndrome. it is an acute infectious disease caused by severe acute respiratory syndrome coronavirus virus (sars-cov- ). the severe disease is characterised by acute respiratory distress syndrome, septic shock, metabolic acidosis, coagulation dysfunction, and multiple organ dysfunction syndromes. currently, no drugs or vaccine exist against the disease and the only course of treatment is symptom management involving mechanical ventilation, immune suppressants, and repurposed drugs. as such the severe form of the disease has a relatively high mortality rate. last months have seen an explosion of information related to the host receptors, virus transmission, virus structure-function relationships, pathophysiology, co-morbidities, immune response, treatment and most promising vaccines. this review takes a critically comprehensive look at various aspects of host-pathogen interaction in covid- . we examine genomic aspects of sars-cov- , modulation of innate and adaptive immunity, complement-triggered microangiopathy, and host transmission modalities. we also examine its pathophysiological impact during pregnancy, in addition to various gaps in our knowledge. the lessons learnt from various clinical trials involving repurposed drugs have been summarised. we also highlight the rationale and likely success of the most promising vaccine candidates. receptor on enterocytes in the small intestine and is consistent with clinical reports of gastrointestinal symptoms and viral shedding in faeces ( , ) . this has been further resolved with the comprehensive identification of host cells/tissues expressing both ace and tmprss ( figure ). thus, likely targets for sars-cov- primarily include secretory goblets of the nasal mucosa, lung type ii pneumocytes and absorptive erythrocytes of the small intestine ( ) . of note, this study also showed that the ace receptor is an interferonstimulated gene in sars-cov- infection in the cells of the human upper nasal epithelium and lung, predominatly mediated by ifn-α and ifn-γ ( ) . moreover, bystander cells are subject to interferon-mediated effects (upregulation of ace receptor) rather than sars-cov- infected cells, suggesting a mechanism of enhanced viral targeting and entry during pathogenesis and a possible avenue for therapeutic intervention ( ) . analysis of genetic variation in the ace gene has identified single nucleotide polymorphisms (snps) that differ in frequency globally among the human population, particularly between males and females ( ). characterising these snps more fully with epidemiological and clinical data on covid- will in time shed light on the precise molecular mechanisms of transmission and disease. furthermore, in the sars-cov- viral s protein, amino acid substitutions have been described, although these occurred outside the rbd that directly interacts with ace ( ) . of paramount importance is characterising the genetic variation and its consequences in the s protein and its rbd, as this will determine whether the sars-cov- virus is evolving and is likely to be a seasonal infection with new variants for the human population. undoubtedly, variation in the s protein and ace , the central interface of hostpathogen interaction in covid- will have evolved from natural selection contributing to the pathogenesis of this disease. proteins and transported into the endoplasmic reticulum (er). these proteins are processed via the secretory pathway and are transported into the er-golgi intermediate compartment ( , ) , where the full-length viral genomes are packaged with the nucleocapsid n protein, budding from the membrane, and thus forming the enveloped mature virion ( ) . the n protein has two domains that can bind the rna genome, with the aid of nsp protein, and attaching it to the rtc, facilitating the packaging of the virus ( ) ( ) ( ) . the viral m protein has three transmembrane domains and is responsible for the majority of protein-protein interactions needed for virus assembly, including membrane curvature and binding the nucleocapsid ( , ) . pseudo-virus particles can also only be formed when there is a co-expression of m protein and e protein, indicating the requirement of both these two proteins to form the coronavirus envelope ( ) . the viral e protein is also involved in structural shaping of the viral membrane envelope and in inhibiting m protein aggregation, as well as a role in pathogenesis ( ) ( ) ( ) ( ) . after the assembly of the mature virions, they are transported in vesicles, where they are released from the infected cell via exocytosis ( ) . unlike sars, covid- patients had the highest viral load near presentation, which could account for the fast-spreading nature of this epidemic. in a study involving covid- patients in hong kong recorded high viral load on presentation with the onset of symptoms and also when the symptoms are mild ( ) . sars cov- viral rna load was detected in the deep throat (posterior oropharyngeal) saliva samples for days or even longer. the peak of the viral load correlated positively with age. viral load in posterior oropharyngeal saliva samples was higher during the first week of symptom onset, which gradually declined. thus, the location of sample collection and the timing for the onset of symptoms both are important factors to be considered for the detection of sars cov- positive cases. in the same study, most of the patients showed rising antibody titres days after symptom onset, though the serum antibody levels did not show correlation with clinical severity ( ) . the patient's antibody to sars-cov- viral nucleocapsid protein using infected cell lysates was identified on the th day after symptom onset by western blot ( ) . in another study involving patients with covid- , all were tested positive for antiviral igg within days after symptom onset. both igg and igm titres reached a plateau within days after seroconversion ( ) . in wuhan tongji hospital, around convalescent patients tested positive for the igg against the virus, while patients tested negative for igm, where igg titre was higher comparatively. both the antibody titres showed a decrease when tested weeks apart ( ) . thus, titres of sars-cov- antibodies can reflect the progress of viral infection and can be a vital component to understand the development and prognosis of the disease and similarly timing of antibody seroconversion is also crucial for determining the optimum duration for collecting serum specimens for antibody diagnosis. as previously mentioned, several other studies also confirmed the presence of sars-cov- nucleic acids in the faecal, urine samples and rectal swabs of covid- patients and thus it becomes essential to ascertain viral load dynamics in such samples too ( ) ( ) ( ) . transmission sars-cov- is transmitted through "respiratory droplets", which are large droplets of virusladen mucus or through close contact with infected individuals ( ) ( ) ( ) ( ) . at the same time virus has also been reported to spread via asymptomatic but infected individuals in several countries, including china, germany, usa, and india ( , ( ) ( ) ( ) ( ) . a systematic review and meta-analysis of observational studies with no randomised controlled trials and relevant comparative studies in health-care and non-health-care settings revealed transmission of virus decreased as physical distancing increased to metre or more ( ) . eye protection, n or similar respirators in health-care settings and - -layer cotton or surgical masks in the community were found to greatly control the transmission ( ) . studies have also established that the median half-life of the aerosolised virus is ~ . hours under lab conditions, similar to the sars-cov. however currently, no evidence supports real-world airborne transmission of the virus through aerosols ( ) . sars-cov- was found to remain viable for up to hours on copper surfaces, up to hours on cardboard surfaces, and up to hours on plastic and stainless-steel surfaces. thus, there exists a possibility of contact transmission to occur, although no confirmed cases of contact transmission have been reported ( ) . the virus was also found in the faeces of infected patients showing that the virus can survive and replicate in the digestive system ( ) . this suggests that there may be a possibility of an oral-faecal route of transmission, though again no confirmed cases have been reported ( ) . the royal college of obstetricians and gynaecologist uk have reported that transmission from mother to baby antenatally or intrapartum is possible although this requires further study for confirmation; there appears to be no evidence supporting vertical transmission to the foetus ( ) ( ) ( ) . additionally, as reported by who and cdc, the virus has not been found to be transmitted by breastfeeding and has not been found in breastmilk of covid- mothers ( , ) . covid- was found to have low severity and mortality than sars, but it is highly contagious and affecting comparatively more men than women ( , , ) . the difference in fatality rate between males and females may probably be explained by the fact that as ace is located on the x chromosome. there may be alleles that confer resistance to covid- , at the same time, oestrogen and testosterone sex hormones have different immunoregulatory functions that may contribute to protection or severity of the disease ( , ) . the disease has also been found to disproportionately affect older aged persons and people suffering from social deprivation, diabetes, severe asthma, cardiovascular disease, obesity, haematological malignancy, recent cancer, kidney, liver, neurological or autoimmune conditions ( ) . studies have also reported that members of minority communities such as the black and south asian populations, are at a higher risk of the disease ( ) . the incubation period of the disease ranges between to days and the median incubation period is approximately - days before symptom onset ( , , , ) . during the onset of the illness, the common symptoms that most patients exhibited were fever and cough. other symptoms include conjunctivitis, myalgia (muscle pain) or fatigue, headache, dyspnoea (short of breath), chest pain, diarrhoea, nausea, rhinorrhoea (runny nose), vomiting, loss of appetite, abdominal pain, gastrointestinal bleeding, autoimmune haemolytic anaemia, and sometimes haemoptysis (coughing of blood) ( , ( ) ( ) ( ) ( ) ( ) . patients have also reported anosmia (loss of smell), dysgeusia (distortion of the sense of taste) ( ) ( ) ( ) ( ) . for sars-cov- asymptomatic patients, anosmia, hyposmia, or dysgeusia are symptoms that were suggested for screening ( ) . in addition to these, neurological manifestations such as dizziness, headache, impaired consciousness, acute cerebrovascular disease, ataxia, seizure, nerve pain, skeletal muscular injury manifestations, intracerebral haemorrhage, central nervous system vasculitis, encephalopathy, encephalitis, cranial neuropathies and psychosis were reported predominantly in older people ( ) ( ) ( ) . in paediatric patients, an autoimmune and autoinflammatory disease, paediatric inflammatory multisystem syndrome (pims), also known as multisystem inflammatory syndrome in children (mis-c), has been reported to occur after sars-cov- infection ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . cutaneous manifestations of covid- have also been reported ( ) ( ) ( ) . a case report from strasbourg, france reported purpuric lesions in the lower extremity ( ) . an italian study reported patients presenting with an erythematous rash, urticaria and chickenpox-like vesicles mainly in the trunk with little or no itching that did not correspond to disease severity ( ) . the prolonged use of personal protective equipment and repeated washing have also led to an increase in dermal conditions such as pressure injury, contact dermatitis, itch, pressure urticaria, and exacerbation of pre-existing skin diseases ( ) . the first step of infection is the inhalation of viral particles present in respiratory droplets from an infected host. once inhaled, the virion enters the nasal cavity of a healthy host and likely binds to goblet and ciliated cells in the nose that express ace ( ) . at this time, a limited innate immune response may occur, and the virus replicates and moves further down the respiratory tract via the conducting airways. as the virions proliferate and spread towards the upper respiratory tract, usually a robust innate immune response is triggered by the detection of the virions by pattern recognition receptors (prrs) like toll-like receptors, rig- , and mda- . this may present several symptoms starting from dysphonia (hoarseness), ulceration of the epiglottis and subglottis, and profound oedema and granulations in the subglottis and also in the upper trachea ( ) . in a few patients, mild tachypnoea and coarse breath sounds were also observed while the virus is in the upper airway ( ) . furthermore, the detection by prr leads to the expression of type interferons (ifn) in the early stages of infection, which helps establish an anti-viral state in the cells by producing inflammatory cytokines and chemokines. the sars-cov produces an enzyme that adds ' o-methyl group to viral rna, which helps it evade detection by mda- , thereby delaying the induction of ifn. studies have established that unlike an early ifn response, a delayed ifn response causes an inability to control viral replication, leading to cellular damage of airway epithelia and the lung parenchyma and an eventual lethal inflammatory cytokine storm ( ) ( ) ( ) . the sars-cov- papain-like protease, which is essential to generate the rtc, has been shown to preferentially cleave the ubiquitin-like protein isg from interferon responsive factor (irf ), attenuating type i interferon responses ( ) . the c-terminus of the sars-cov- non-structural protein was reported to bind to the s ribosomal subunit and block the mrna entry tunnel ( ) . this obstruction effectively inhibits the rig-i-dependent innate immune response ( ) . accordingly, no significant expression of ifn was detected up to hours post-infection with sars-cov- . only il- , which correlates with respiratory failure, acute respiratory distress syndrome (ards), and adverse clinical outcomes were upregulated. monocyte chemoattractant protein- (mcp ), c-x-c motif chemokine (cxcl) , cxcl , and cxlc , were also upregulated h post-infection with sars-cov- ( ) . the suppression of innate immune activation and annihilation of t cells can help explain the mild or even the lack of symptoms in many infected patients. the increased viral replication efficiency in the respiratory tract early on leads to the highly efficient person-to-person transmission of the virus in the community ( ) . the virions further migrate towards the lower respiratory tract and reaches the alveoli where it binds to the type pneumocytes and begins replication. as the type pneumocytes undergo apoptosis after viral release, they secrete inflammatory mediators like cxlc proteins that attract macrophages and neutrophils ( figure ) ( ) . the stimulated macrophages further secrete cytokines such as il- β, il- and tumor necrotic factor α (tnf-α). the released cytokines trigger a "cytokine storm", which stimulates the release of vascular endothelial growth factor (vegf), monocyte chemoattractant protein- (mcp- ), il- , and additional il- , as well as reduced e-cadherin expression on endothelial cells causing vasodilation and increase capillary permeability ( ) . this causes the plasma to leak into the interstitial spaces and alveoli, increasing interstitial and alveolar oedema. the increased alveolar oedema decreases the level of surfactant in the alveoli. this causes an increase in the surface tension in the alveoli, which leads to alveolar collapse. oedema and alveolar collapse may present as multiple peripheral ground-glass opacities in subpleural regions of both lungs, which is observed in many patients ( ) . chest ct scan of patients also revealed bilateral multifocal infiltrates and mediastinal and hilar lymphadenopathy in some patients ( ) . these decrease the gas exchange efficiency causing hypoxemia and increased work of breathing presenting as dyspnoea (shortness of breath), culminating in ards ( ) . abnormal coagulation parameters, mainly elevated d-dimers seem to be associated with a higher risk of development of ards in covid- patients ( ) . the aberrant wound healing may even lead to fibrosis than other forms of ards ( ) . stimulated neutrophils secrete reactive oxygen species (ros) and proteases which destroy both infected and uninfected type and type pneumocytes, leading to further reduced gas exchange and alveolar collapse, respectively ( ) . furthermore, the dead pneumocytes slough off into alveoli filling them up with fluid, protein deposits, cell debris, macrophages, and neutrophils. this causes pulmonary consolidation, which leads to altered gas exchange and causes hypoxemia ( , ) . the consolidation also leads to productive cough. the hypoxemia can further trigger chemoreceptors that stimulate the sympathetic nervous system (sns) that causes tachycardia (increased heart rate) and tachypnoea (increased respiratory rate) ( , ) . the central nervous system (cns) is also affected by the high concentrations of il- β, il- and tnf-α in the blood, as these cytokines stimulate the hypothalamus to release prostaglandins such as pge , which causes an increased body temperature leading to fever ( ) . studies have also reported elevated levels of myeloperoxidase (mpo)-dna and citrullinated histone h (cit-h ), which are markers used to detect neutrophil extracellular traps (nets), in the serum of covid- patients ( ) . furthermore, control neutrophils treated with covid- patient serum exhibited netosis ( ) . nets, while protecting the host from invasive pathogens, have been attributed to play a role in many autoimmune and vascular diseases. for example, nets are known to contribute to ards, pathogen-induced acute lung injury, thrombosis and can contribute to further cytokine release lading to the inflammation ( ) . an increased frequency of neutrophils, eosinophils and monocytes was reported in severe covid- positive patients; severe patients showed further increase in neutrophils though their activation status had not altered. there was no significant change in the immature granulocyte frequencies. however, there was an inverse correlation between frequency of immature granulocytes in moderate and severe patients with the duration since the appearance of symptoms. severe patients exhibited lower percentages of both conventional and plasmacytoid dendritic cells (dc) ( ) . response syndrome (sirs). the spread of the inflammation from the lungs into the circulatory system causes increased capillary permeability within the systemic circulation. this leads to a decrease in blood volume along with increased vasodilation of systemic arteries, leading to decreased peripheral resistance. the decreased blood volume, along with peripheral resistance, causes hypotension (decreased blood pressure), which decreases perfusion to other organs leading to multisystemic organ failure (mof) ( - ). the cytokine storm has also been shown to trigger autoimmune haemolytic anaemias (aiha) (with warm or cold antibodies) ( , , ) . most of the studies report manifestation of aiha early, during the active phase of covid- (within to days), a timeframe matching that of the cytokine storm ( , , , ) . as a result of ards, sirs and mof, patients suffering from severe sars-cov- infection exhibit significantly elevated levels of, il- , il- , il- , g-csf, gm-csf, mip- α, crp, and ddimer, in addition to il- , il- β and tnf-α ( ). there are reports suggesting that in addition to the lungs, sars-cov- infection may induce the multiorgan injury in patients involving brain, heart, liver, kidney, intestine and eyes ( , ) . covid- associated neurological complications the neurological pathologies observed in covid- are similar to those observed in previous coronavirus epidemics ( ) . myoclonus and demyelination are reported in a few cases ( , , ) . a study conducted in wuhan, china involving covid- patients, reported that patients developed neurological manifestations ( ) . in another study from strasbourg, france where effectively patients were recruited for an observational study, reported agitation in % of the patients, confusion in %, and % of the patients had corticospinal tract signs ( ) . a systemic review and meta-analysis of literature databases for psychiatric and neuropsychiatric presentations in coronavirus infections reported transient encephalopathies with features of delirium and psychosis ( ) . the study also reported cognitive dysexecutive syndrome and delirium with agitation in a few cases ( ) . there is also a reported case of autoimmune encephalitis with the typical clinical features of opsoclonus and myoclonus, and another case of autoimmune encephalitis with a radiological imagery showing typical limbic encephalitis ( ) . the exact mechanism for encephalopathy may be multifactorial (effect of sepsis, hypoxia, and/or cytokine storm) ( ) . a few cases of guillain-barré syndrome (gbs) associated with sars-cov- have been reported from italy ( ) . however, further epidemiological and mechanistic study is required to confirm the incidents of gbs in covid- . the binding of the virus to the ace- receptors on endothelial cells causes extravasation of red blood cells leading to cerebral microbleeds ( , ) . there have also been reports of severe strokes in covid- patients, but further study is required to determine its association with covid- ( ) . magnetic resonance imaging (mri) revealed abnormalities such as meningeal enhancement, ischaemic stroke, perfusion changes, microhaemorrhages, medial temporal lobe signal abnormalities similar to that seen in viral or autoimmune encephalitis ( , ) . very few cases have been reported where sars-cov- was detected in csf and its supportive histopathological features; no reports of the virus in the brain exist yet ( , , ) . thus, it is important to establish whether the above-described syndromes may be caused due to either direct viral injury, hyperinflammation, vasculopathy and/or coagulopathy, autoantibody production to neuronal antigens, sepsis and hypoxia, or a combination of these ( ) . out of the first patients diagnosed with covid- in wuhan, of them had myocardial injury associated with the sars-cov- , which mainly manifested as an increase in highsensitivity cardiac troponin i ( ) . the hemogas analysis showed hypoxia; laboratory tests showed elevation of c-reactive protein, transaminases and lactate dehydrogenase, and lymphopenia ( ) . several patients showed abnormal myocardial zymogram, showing high levels of creatine kinase ( ) . because of an excessive inflammation, hypoxia, immobilisation and diffuse intravascular coagulation (dic), covid- patients may predispose to both venous and arterial thromboembolic disease ( , , ) . it has also been observed that concomitant acute thrombosis of the abdominal aorta and pulmonary embolism induces cardiovascular complications in covid- patients, suggesting an association of hypercoagulable condition with the disease ( ) . covid- patients with abnormal liver function were also documented, where patients had alanine aminotransferase (alt) or aspartate aminotransferase (ast), bilirubin, acute phase recants (apr) like crp, fibrinogen and il- above the normal range ( , ) . sepsis, hypovolaemia, and nephrotoxins were found to be important contributors to kidney damage in covid- patients. cardiorenal syndrome, particularly right ventricular failure, might lead to kidney congestion and acute kidney injury in covid- patients ( ) . symptoms such as olfactory and gustatory dysfunctions were also found to be related to covid- ( ) . sars-cov- , facilitated by tmprss and tmprss , was found to infect and reproduce in ace- + mature enterocytes ( ) . however, the virions released into the intestinal lumen were inactivated by stimulated human colonic fluid and no infectious virions were recovered in stool samples, in spite of the presence of viral rna in stools. this study thus established the intestine as a site of viral replication and its effect on local and systemic illness and overall covid- progression ( ). as in the case respiratory infections by respiratory syncytial virus and sars-cov, the eyes have been shown to act as a portal of entry for the virus. while there have been no reports of sars-cov- transmission in humans via ocular tissues, further studies are required to exclude the eyes as a source of infection and as a portal of entry. moderate conjunctivitis could be the first sign of severe respiratory distress in covid- patients ( ) . studies from china on patients with covid- reported conjunctivitis and other ocular manifestations, such as epiphora, conjunctival congestion, or chemosis in patients with severe covid- ( ) ( ) ( ) ( ) . the studies also reported a few patients with positive conjunctival swab for covid- determined by rt-pcr ( , , ) . similar results were also reported in a study conducted by the national institute for infectious diseases in rome, italy ( ) . in addition to the conjunctivitis, the ocular swabs were positive for sars-cov- even when nasopharyngeal swabs tested negative for the virus. this suggests that the conjunctiva may sustain viral replication for an extended period of time ( ) . there are reports from france, italy, united kingdom and the united states of america, suggesting the presentation of autoimmune and auto inflammatory diseases in children, especially in children of african descent, such as paediatric inflammatory multisystemic syndrome (pims), also known as, multisystemic inflammatory syndrome in children (mis-c) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . this syndrome includes kawasaki-like disease, kawasaki disease shock syndrome, toxic shock syndrome, myocarditis and macrophage activation syndrome ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . the exact cause for kawasaki disease remains unknown; however, it is believed that it is caused by an apparent atypical immune response to pathogens in genetically predisposed individuals ( ) ( ) ( ) . previous studies have implicated the pathogenesis of kawasaki disease with the infection of certain members of the coronavirus family ( , ) . the temporal association between the beginning of covid- , sars-cov- infection and the onset of pims suggest a causal link ( ) . this is further supported by the fact that in most cases, the patients exhibiting pims tested positive for igm or igg sars-cov- antibodies ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . the presence of igg antibodies clearly indicates a delayed onset of pims following sars-cov- infection ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . the onset of pims occurred - weeks after acute covid- ( ) . the patients presented with fever, diffused skin rashes, rash/oedema of hands and feet, conjunctivitis, dry cracked lips, cervical lymphadenopathy and arthritis. the kawasaki-like disease caused by covid- exhibited a few differences in both clinical and biochemical features from patients suffering from kawasaki disease without sars-cov- infection. clinically, the patients suffering from covid- associated kawasaki-like disease were older and the disease occurred in both sexes, unlike the classical kawasaki disease that occurs in younger male children ( ) . the covid- associated kawasaki-like disease also had a higher incidence of abdominal pain and/or more frequent diarrhoea, meningeal and respiratory involvement, and a strikingly different myocarditis severity and frequency when compared to classical kawasaki disease ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . biochemically the patients exhibited leukopenia with thrombocytopenia, increased ferritin, elevated myocarditis markers and high levels of procalcitonin, crp and cytokines were observed when compared to classical kawasaki disease ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . nearly % patients also showed resistance to the initial treatment with intravenous ivig infusion, and required a second infusion for successful treatment ( , ) . while the children exhibited the devastating effects of the cytokine storm associated with covid- , such as heart failure, pneumonia, gastrointestinal, neurological and renal manifestations, the paediatric patients in the french study rarely had respiratory manifestations ( ) . this suggests a different host immune response in children compared to adults. treatment for pims involves the administration of il- receptor antagonist, il- receptor blockers such as tocilizumab or sarilumab, ivig, and steroids or biologics to control inflammation ( , ) . covid- is known to affect older members of the population disproportionately, with adults over the age of years making up to % of hospitalization and having a -fold greater risk of death ( , ) . one possible explanation for this could be the increased baseline inflammation, called inflammaging, commonly observed in individuals over the age of ( ) . studies have shown increased baseline serum concentrations of crp and cytokines such as il- and il- ( ) . inflammaging could be the result of the accumulation of mis-folded proteins, compromised gut barrier, obesity and impaired clearance of dead or dying cells ( , ) . senescent nonlymphoid cells have been known to secrete inflammatory cytokines, chemokines, growth factors, and matrix metalloproteinases ( , ) . this increased baseline inflammation inhibits antigen-specific immunity affecting the efficacy of many vaccines ( ) . studies have shown that treatment with rapamycin, mapk inhibitor or steroids reduces this excessive inflammation and enhances vaccine efficacy ( , , ) . in case of covid- , this baseline inflammation may itself not be detrimental but contributes to the initiation of an inflammatory cascade that ends in the deadly cytokine storm ( ) . furthermore the accumulation of senescent cells in the lungs of older patients could inhibit t cell response, induce nkr ligand expression, which marks the cells for elimination by infiltrating t cells expressing nkrs ( ) . as observed in the case of vaccines against other respiratory viruses, inflammaging may reduce the efficacy of covid- vaccinations in this already disproportionately affected group. as with any infection, both the innate and adaptive arms of the immune system are required to mount a successful defence against a viral incursion. in case of covid- , a decrease in the circulating t helper cells (cd + cells), cytotoxic t cells (cd + cells), b cells, natural killers cells, lymphocytes, monocytes, eosinophils and basophils has been reported ( , , , ) . a retrospective, single-centre study involving patients revealed a significant decrease in the total number of regulatory t cells, memory t cells and suppressor t cells ( ) . the study also reported an increase in the percentage of naïve t cells ( ) . as naïve t cells help respond to novel pathogens that the immune system has not yet encountered by managing release of cytokines, this may help explain the hyperinflammation ( ) . the lower levels of memory t cells reported may also explain the relapses reported in covid- convalescent individuals ( ) . direct infection of thp- cells, human peripheral blood monocyte-derived macrophages and dendritic cells by mers-cov and infection of t cells and macrophages by sars-cov has been reported ( , ) . hence, it can be speculated that sars-cov- may also infect monocytes and macrophages by a mechanism that is yet to be elucidated ( ) . receptors such as cd on the surface of t cells and other immune cells may mediate viral entry ( ) . the clinical trial with anti-cd monoclonal antibody, meplazumab, showed promising efficacy and safety in covid- patients ( ) . however, cd did not show a direct interaction with the s protein of sars-cov- ( ) . similarly, lymphopenia can be attributed to sars-cov- direct infection and lymphocyte death, destruction of the lymphatic organs, and/or high levels of the programmed cell death protein (pd- ) on cd + t cells (which is known to trigger t cell exhaustion) ( , , ) . lymphocytopenia, neutrophilia and neutrophil-to-lymphocyte ratio are being used as a predictor for the severity of the illness during early stages of infection and a poor outcome in covid- ( , , , ) . this further alludes to the hyper-inflammatory nature of covid- . furthermore, covid- patients were reported to have elevated serum levels of highsensitivity c-reactive protein and procalcitonin, whose levels have been associated with high risks of mortality and organ injury ( ) . lower percentage and count of cd + , cd + , and cd + lymphocytes populations serve as a prognostic marker for mortality, organ injury, and severe pneumonia ( ) . sars-cov exposed as well as a subset of non-exposed people exhibit a cross-reactive t cell repertoire ( ) . studies have also reported the presence of sars-cov- spike glycoprotein-reactive cd + t cells in peripheral blood of a subset of donor who were not infected with sars-cov- ( , ) . these s reactive cd + t cells were found to primarily react with the c-terminal of the s epitope ( ) . this binding preference could be attributed to the presence of overlapping human coronavirus mhc-ii epitopes in the c-terminal domain. hence, these cd + t cells are cross reactive clones generated during previous infections with endemic human coronavirus ( ) . a long-term information and knowledge for ageing -camden' (linkage) sub-study is currently underway to study if pre-existing antibodies and specific t cells contribute to the devastating effect observed in old people ( ) . the b cell response occurs alongside the t helper cell response (~ week post infection) in covid- patients and helps mount a humoral response via antibodies that would help neutralise the virus ( ) . characterisation of the transcriptome during the recovery stage of the disease revealed significantly lower levels of naive b cells, while plasma b cell levels had increased in peripheral blood mononuclear cells ( , ) . it was found that a certain subset of patients who contract the disease may not develop long-lasting antibodies against the pathogen; it is possible that these patients may be susceptible to the re-infection ( ) . immune cell profiling of covid- patients in the recovery stage by single-cell sequencing has identified several new b cell-receptor changes such as ighv - and ighv - , and isotypes used earlier for vaccine development including ighv - , ighv - , and igkv - ( ) . the strongest pairing frequencies, ighv - -ighj , has been suggested to indicate a monoclonal state associated with sars-cov- specificity ( ) . antibodies analysed from the serum of covid- patients revealed no cross-reactivity with the s subunit of the sars cov spike antigen, while some reactivity was observed between the nucleocapsid antigens of sars-cov and sars-cov- ( ) . the rbd-specific igm and igg antibodies were significantly elevated in the severe and recovered patients ( ) . investigations conducted on covid- recuperating rhesus macaque models, re-infected with sars-cov- , reported no measurable viral spreading, clinical manifestations, or histopathological changes associated with covid- ( ) . the study found lower viral loads in nasopharyngeal or anal swabs or days after reinfection, compared to the recorded viral loads or days after the initial infection with sars-cov- at similar sites. similarly, increased levels of leukocytes and neutrophils were recorded days after reinfection, compared to the levels measured during the initial infection. significantly higher specific antibody titres were recorded day post reinfection. there were also increased activation of cd + t cells, changes in cd + tcm cells and memory b cells. thus, increased production of neutralising antibodies protected the primates against covid- re-infection ( , ) . a study on covid- convalescent individuals revealed that plasma collected after days of symptom manifestation had a variable half-maximal pseudovirus neutralizing titres of less than : in %, below : , in %, and only % showed titres above : , ( ) . interestingly, in spite of the low titres reported, antibodies specific to three distinct epitopes on the rbd of the sars-cov- s protein neutralized at half-maximal inhibitory concentrations as low as single digit ng/ml ( ) . hence, a vaccine that can elicit the production of such highly potent antibodies, or monoclonal antibodies raised against the rbd of the sars-cov- s protein, may be highly protective. however, studies on sars-cov and mers-cov revealed that neutralizing antibodies to s protein can potentially augment severe lung injury by exacerbating inflammatory responses ( , ( ) ( ) ( ) . hence, therapeutic antibodies should be carefully studied to minimise any unwanted pro-inflammatory activity while retaining maximum virus neutralizing capacity. additional specific insights on the intracellular life cycle have also been gained from nextgeneration sequencing (ngs) studies on the transcriptome and epi-transcriptome profile of sars-cov- virus and infected host cell. this fundamental approach has given an insight into the specific molecular dialogue between the pathogen and the host cell. this dialogue is complex. the sars-cov- transcriptome has been studied in high resolution. it has revealed its highly complex nature, mainly as a result of numerous discontinuous transcription events, revealing canonical and non-canonical rna transcripts with rna modifications ( ) . in addition to the canonical full-length genome and other sgrnas, this study also found numerous non-canonical rna transcripts of unknown orfs that contained rna modifications. putative rna medications were identified at an aagaa motif. these previously unknown orfs represent the epi-transcriptome of sars-cov- and has revealed numerous viral transcripts that may be involved in pathogenesis ( ) . another study looked at transcriptome profiling in the primary human lung epithelium and compared differences between sars-cov- and sars-cov infection and identified several pathways potentially involved in pathogenesis and gender-specific differences in clinical presentation ( ) . among the genes that were upregulated were a cluster involved in the cytokine-mediated signalling pathways, and in particular, the il- signalling pathway ( ) . specifically, cytokine pathways driven by nuclear factor kappa-light-chain-enhancer of activated b cell (nf-κb), toll-like receptors (tlrs), mitogen-activated protein kinase (mapk), bone marrow stromal cell antigen (bst ), il- , tnf alpha induced protein (tnfaip ), tnfaip interacting protein (tn p ), intercellular adhesion molecule (icam- ), intercellular adhesion molecule (icam- ), matrix metallopeptidase (mmp ), baculoviral iap repeat containing (birc ), and rho family gtpase (rnd ), were significantly upregulated during sars-cov- infection, suggesting a significant role in pathogenesis ( ) . moreover, rela (nf-κb p subunit) seems to be significantly upregulated in sars-cov- infection, leading to il- involvement ( ) . of note is the expression of oestrogen receptor (esr ), which was also enhanced under sars-cov- infection, suggesting sex hormones may be involved in differential expression during viral infection and may have implications for the differences in clinical severity seen between genders ( ) . additionally, over and hour post-infection, cxcl- was significantly upregulated in sars-cov- infection compared to sars-cov ( ) . a recent study using single-cell rna-seq in human, non-human primate and mouse tissues/cells was able to resolve further the host cellular targets for sars-cov- and their abundance in specific tissue/cell types ( ) . the study identified ace and tmprss co-expressing cells (lung type ii pneumocytes, ileal absorptive enterocytes and nasal goblet secretory cells) and also determined that that ace is induced by interferon-stimulated gene, suggesting a possible mechanism for enhanced viral infection ( ) . the clinical pathways of covid- disease severity may also depend on host-specific factors that may contribute to the 'cytokine storm', or cytokines release syndrome (crs), which is the massive release of pro-inflammatory cytokines including cytokines (il- β, il- , il- , il- , il- , and tnf-α) and chemokines such as cxcl and ccl in the lungs ( , ) . these genomic approaches also shed light on the specific genetic host factors that predispose individuals to this severe clinical presentation. proteomic and transcriptomic studies on bronchoalveolar lavage (bal) samples from covid- patients have also revealed considerable insights into the expression of sars-cov- receptors, co-receptors, immune responses, as well as risk factors for severe disease e.g. age and co-morbidities. asthma, chronic obstructive pulmonary disease (copd), hypertension, smoking, obesity, and male gender status were all associated with higher expression of ace and cd in bal, as well as bronchial biopsy and blood from covid- patients ( ) . furthermore, there was a positive correlation between the expression of cd -related genes in bal and the age and body mass index (bmi) of covid- patients ( ) . in another study on bal from covid- patients, an association was observed between covid- severity and enhanced levels of certain cytokines, e.g. ccl /mcp- , cxcl /ip- , ccl /mip- a, and ccl /mip b ( ) . this study also found that sars-cov- triggered apoptosis and the p signalling pathway in lymphocytes, probably causing additional lymphopenia in these patients ( ) . a comparison of transcriptome profiles between patients with covid- and influenza a virus infection revealed an absence of significant type i interferon/antiviral responses with sars-cov- infection, with enhanced expression of genes involved in metabolic pathways e.g. haem biosynthesis, oxidative phosphorylation and tryptophan metabolism, suggesting an important role for mitochondria during sars-cov- infection ( ) . furthermore, a meta-analysis on bal data from covid- patients also revealed an excess for neutrophils and chemokines ( ) . in meta-transcriptomic sequencing of bal from covid- patients, the expression of proinflammatory genes, especially chemokines, was significantly elevated in these patients compared to community-acquired pneumonia patients and healthy controls, suggesting hypercytokinemia ( ) . it also revealed enhanced dendritic cell and neutrophil activity ( ) . in contrast to sars-cov, which induces an ineffective interferon response, sars-cov- was found to strongly initiate expression of numerous interferon stimulated genes, which are thought to significantly contribute to immunopathogenesis ( ) . similarly, an analysis of rna-seq data sets of bal from covid- patients identified upregulation of neutrophil, inflammatory genes and chemokines, which may be involved in immunopathology, e.g. tnfr, il- , cxcr , cxcr , adam , gpr , mme, anpep, and lap ( ) . chronic co-morbidities for covid- patients include cardiovascular disease, hypertension, diabetes, stroke and malignant tumour ( ) . it was also found that parameters such as older age, underlying hypertension, high cytokine levels (il- , il- , and tnf-α), and high lactate dehydrogenase level were significantly associated with severe covid- during hospital admission ( ) . in a study involving icu patients with covid- pneumonia, all of them showed an incidence of thrombotic complications such as symptomatic acute pulmonary embolism (pe), deep vein thrombosis, ischemic stroke, myocardial infarction or systemic arterial embolism ( ) . approximately, one-third of patients experienced gastrointestinal symptoms. during hospitalization, a substantial proportion of patients presented cardiac injury, liver, and kidney dysfunction, and hyperglycaemia. icu covid- patients had higher plasma levels of il- , il- , il- , gscf, ip , mcp- , mip- , and tnf-α, compared to non-icu patients. majority of icu patients diagnosed with covid- were found to be at highest thrombotic risk ( ) . patients with severe covid- likely developed ards and died of respiratory failure. biopsy samples at autopsy from a patient who died from severe covid- showed bilateral diffuse alveolar damage with cellular fibromyxoid exudates, and mononuclear inflammatory lymphocytes in both lungs ( , ) . diffuse alveolar damage with fibrin rich hyaline membranes are pathological results of covid- . in a study, covid- -infected cancer patients were found to have underlying diseases, such as hypertension, diabetes and chronic obstructive pulmonary disease ( ) . cancer patients with accompanying covid- infection showed deteriorating conditions and poor outcomes, and thus it was recommended to avoid treatments causing immunosuppression ( ) . the complement system is an integral part of the innate immune response. it consists of a group of plasma proteins produced mainly by the liver or membrane proteins expressed on cell surface. these proteins interact in a cascade that leads to the opsonization of pathogens and the induction of inflammatory responses. the complement system comprises of three distinct activation pathways, i.e. classical, alternative or lectin (mbl). the activation of these pathways is based on different molecules present on the pathogen surfaces. the classical pathway is initiated by the binding of c q to the pathogen surface or antibody complex. the initiation of the alternative pathway is triggered by the binding of a spontaneously activated complement component to pathogen surface. the binding of the mbl to mannose-containing carbohydrates on pathogens triggers the initiation of the lectin pathway. the early events of three pathways eventually converge to generate a protease called, c convertase, which is covalently bound to the pathogen. the c convertase then cleaves c , present in plasma, into c a and c b. the c b binds to the pathogen and targets it for destruction by phagocytes. furthermore, c b binds with the c convertase to form c convertase, which produces c a and c b. c b triggers the late events of the complement cascade, which are a series of polymerization reactions where c , c , c and c interact with each other to form the membrane attack complex (mac). the mac can damage the membrane of certain pathogens by creating a pore in it. the c a and c a produced are important small peptide mediators of inflammation [reviewed in ( ) ]. studies in c -/-(gene-deficient) mice infected with sars-cov revealed the presence of c activation products such as c a, c b, ic b, c c, and c dg day post infection ( ) . the c deficient mice showed significantly less respiratory dysfunction and lower weight loss as compared to control. the mice also showed significantly lower levels of neutrophils and monocytes compared to the control. lower il- , tnf-α and il-  levels were reported in the lungs of the c deficient mice ( ) . the study also reported lower weight loss in mice deficient in factor b or c . in view of the critical role of the complement system in sars-cov infection since the first day of infection, it raised possibility for complement involvement in sars-cov- . levels of the terminal component of the complement system (mac) and c a are increased in patients with ards ( , ) . mac is known to damage endothelial cells, and thus, regulation or inhibition of mac by its known regulators such as cd or clusterin could be a potential treatment for endothelial dysfunction/damage in ards or covid- ( ) ( ) ( ) . considering the lectin pathway of the complement system, mbl was shown to bind sars-cov in vitro and inhibit its infectivity ( ) . the n-protein of sars-cov and sars-cov- has been shown to interact with mbl-associated serine proteases- (masp ), which is known to initiate the lectin pathway ( ) , leading to over-activation of the complement system. this same study also highlighted excess complement proteins found in post-mortem covid- patient lungs ( ) . furthermore, deletion of masp or perturbance of the masp- -n protein interaction was found to reduce lung injury. these studies, along with human proteomic studies, demonstrate the activation of multiple complement pathways during a coronavirus infection. in case of covid- , the alternative and lectin pathways of the complement system seem to be preferentially activated ( ) . increased levels of plasma c a and mac were recorded in patients with moderate and severe covid- ( ) . a post-mortem study of lung and skin vasculature in covid- patients showed significant deposits of mac and c d that colocalized with the sars-cov- s-protein, and masp in the micro-vasculature. this study did not find prominent classical features of ards such as hyaline membranes and inflammation in the histopathological examination ( ) . a recent study reported an increase in levels of c a, which correlated with increased covid- disease severity, as well as high levels of expression of c ar in blood and pulmonary myeloid cells of covid- patients ( ) . furthermore, use of anti-c ar monoclonal antibodies in human c ar knock-in mice was found to successfully prevent c a-mediated myeloid cell recruitment and activation, thereby inhibiting acute lung injury ( ) . a recent genetic study in covid- patients as reported that gene variants associated with complement regulatory protein, cd (decayaccelerating factor, which accelerates the decay of complement proteins, and thus inhibits complement activation) is associated with increased risk in clinical outcome (odds ratio . - . ); gene variants that map to c showed some protective effect (odds ratio . - . ) ( ) . neutrophils along with the complement system are another important component in the defence of the host against invading pathogens. neutrophil infiltration in pulmonary capillaries, acute capillaritis with fibrin deposition, extravasation of neutrophils into the alveolar space, and neutrophilic mucositis have been reported in the case of sars-cov- infection ( ) . the neutrophilic extracellular traps (nets) and the neutrophils activated by sars-cov- infection contain c , factor b and properdin, triggering the alternative pathway of the complement system ( ) . while this is usually beneficial, the sustained activation and nets formation leads to a hyper-inflammatory immune response that damages and destroy surrounding tissue. this aberrant behaviour, in concert with the abnormal complement activation, leads to the well recorded clinical manifestations observed in the case of covid- such as ards and pulmonary inflammation ( ) . additionally, nets have been reported to induce the production of excessive thrombin and subsequently generate c a ( ) . hence, it has been speculated that feedback loop that begins with complement activation leading to netosis causing an increases in thrombin production, that further stimulates the complement activation causing enhanced net formation ( ) . microangiopathy refers to a disease of the small blood vessels. the term is used when small blood vessels pathologically thicken or weaken, which leads to impaired flow of blood as well as leaking of cells and proteins. sustained inflammation in the vascular system due to the sars-cov- infection leads to thrombosis and microangiopathy ( , ) . this is supported by reports of increased lactate dehydrogenase, bilirubin, activation of platelets, elevated d-dimer levels and hyper-fibrinolysis ( ) . a post-mortem case series of patients with covid- found thrombotic microangiopathy, which was restricted to the lungs, along with diffuse alveolar damage could have contributed to causing death ( ) . another such study of cases found similar diffuse alveolar damage with significant capillary congestion and microthrombi despite anti-coagulation therapy ( ) . due to the presence of severe pulmonary vascular dysfunction in ards, it has been argued that ards is a type of vascular microthrombotic disease with lung phenotype involvement. this argument is supported by the association of mortality in ards with thrombocytopenia and mof as a result of disseminated intravascular coagulation ( ) ( ) ( ) . in recent times, a couple of theories on the pathogenesis of ards in sepsis have evolved: the 'two-path unifying theory' in which certain homeostasis mechanisms lead to microthrombogenesis, and the 'twoactivation theory of the endothelium' in which the complement mac leads to inflammation via cytokines and microthrombogenesis via platelet activation ( , ). the complement system plays a key role in the pathogenesis of thrombotic microangiopathy. this is a syndrome characterised by thrombocytopenia (low platelet count), microangiopathic haemolytic anaemia and systemic organ damage. atypical haemolytic uremic syndrome (ahus) is an example of such a disorder that typically leads to kidney damage. it is caused by excessive activation of the alternative pathway due to mutations in complement regulators factor h (common), factor i, membrane-cofactor protein, or c . analysis of renal tissue morphology from autopsies of covid- patients revealed strong c b- staining (via the alternative pathway) on the apical brush border of tubular epithelial cells with minimal deposition on glomeruli and capillaries of the kidney ( ) . treatment with eculizumab (c inhibitor) dramatically improved outcomes of survival in ahus. features similar to complement-mediated thrombotic angiopathy such as kidney and cardiac injury increased lactate dehydrogenase and d-dimer, and decreased platelets were observed in covid- ( , ) . eculizumab was used successfully as part of management of four covid- patients with severe pneumonia or ards in the intensive care unit, and this preliminary data is being used to conduct further full-fledged clinical trials with eculizumab ( ) . considering the overlap with complement-mediated thrombotic angiopathy in covid- , few studies are underway to test the effectiveness of complement regulators. a recent case study demonstrated a favourable outcome for the compstatin-based c inhibitor amy- . the study, which involved a -year-old caucasian male with severe pneumonia and systemic inflammation, found that amy- was safe and had a favourable outcome in improving the clinical presentation of the patient significantly ( ) . furthermore, treatment with a recombinant c a antibody on male covid- patients aged and years showed significant benefit in suppressing complement hyperactivation, which contributes to the excessive immune response causing aggravated inflammatory lung injury, a hallmark of sars-cov- pathogenesis and lethality ( ) . one of the many challenges includes determining patients who have a dysregulated complement activation. c bound to erythrocytes has been detected in patients with covid- ( ) , which may prove to be a useful blood marker as well as in identifying patients who potentially merit intervention with complement regulators ( ) . in covid- , endothelial injury has been found to be a key pathophysiological feature. a case series found direct evidence of viral infection of endothelial cells and endothelial inflammation, leading to endothelial cell death ( ) . in covid- patients, endothelial cell abnormalities were recorded in the kidney, lung, heart, small bowel, and liver. of deceased covid- patients were found to have suffered endothelial cell swelling with variable foamy degeneration in the glomeruli and an additional patients were found to have severe injury to the endothelium due to segmental fibrin thrombi in glomerular capillary loops ( ) ( ) ( ) . mac deposition has been observed in the endothelium of covid- patients ( ) . such studies have led to notion that in covid- , there are strong vascular and inflammatory components as well, which play a significant role in the pathophysiology of the illness ( ) . consistent with endothelial injury, the significantly elevated levels of von willebrand factor found in the patient with severe covid- has led to the idea that the infection of the ace expressing endothelium by sars-cov- induces injury and activates the complement , which sets up a feedback loop that maintains a state of inflammation ( , ( ) ( ) ( ) . it is worth noting that ards may occur in covid- despite well-preserved lung gas volume, which could indicate a key role for inflammatory processes, leading to vascular constriction and subsequent low oxygen levels in the blood ( ) . furthermore, d-dimer (a fibrin degradation product) levels are also found to be elevated in covid- and are associated with poorer prognosis ( , ) . these factors add to the importance of understanding vascular changes in this disease, including microangiopathic processes and coagulopathies in patients with covid- . pregnancy is associated with several maternal adaptations in both immune function (immunosuppression) and cardiovascular physiology (increased cardiac output, physiological anaemia, cardiac hypertrophy) that would likely alter susceptibility to viral respiratory infections including sars-cov- . maternal death occurred in % of patients admitted to the icu for covid- and in % of those who required invasive mechanical ventilation ( ) . to date, the literature consists of case reports, case series and retrospective studies. the most common presenting symptoms of maternal covid- are fever, cough, dyspnoea, and gastrointestinal symptoms ( ) . clinical findings of respiratory manifestations were similar to those seen in the non-pregnant populations, with similar ct findings together with elevations in c-reactive protein with decreased white blood cell counts ( ) . although the portal of entry is inhalational, there are widespread systemic effects. the immobility, hypoxia and acute inflammation lead to a prothrombotic hypercoagulable state, and indeed, elevated ddimers are correlated with disease severity ( ) . covid- is thus associated with venous or arterial thromboembolism ( ) . the mechanism by which this occurs is currently thought to be as a result of inflammatory cytokines ( ) inducing production of tissue factor with subsequent thrombin activation. the elevations of d-dimer (often seen in acute phases of infection) may be related to this increased thrombin generation. while serious maternal morbidity has been seen, the vast majority of pregnant women with sars-cov- infection remained asymptomatic for respiratory symptoms ( ) ( ) ( ) . pregnancy is coupled to physiological changes in cardiorespiratory status ( ) which might be expected to alter susceptibility to a respiratory upset. nevertheless, the evidence suggests that this is less prevalent than thought. however, the changes in immune function and coagulation in pregnancy appear to increase some complications. similarly, the coagulopathies seen in covid- in the non-pregnant population might be expected to have deleterious effects in pregnancy, which is already a prothrombotic state. covid- has been seen to be associated with preeclampsia ( , ) with one non-peerreviewed report suggesting a causal link ( ) . the placenta has also been reported as having vascular malperfusion and thrombosis ( , ) , which may provide an underlying explanation for the preeclampsia, a disease, associated with poor placental perfusion and altered vascular function ( ) . evidence of increased clotting at the placental surface suggests that this mechanism may be responsible (in part) for the increased incidence of preeclampsia. sars-cov- virions have been seen in the syncytiotrophoblast, the part of the placenta responsible for the angiogenic imbalance seen in preeclampsia and effects on the release of known factors associated with the disease (sflt- -soluble fms like tyrosine kinase and plgf -placental growth factor) are unknown. the disease is also linked to preterm premature rupture of membrane (pprom) ( , ) , and preterm labour ( ) , both of which are linked to inflammation. the underlying mechanism by which pprom occurs is not entirely elucidated. however, reports have suggested that activation of the coagulation system and thrombin causes fetal membrane weakening and subsequent rupture of membranes ( , ) . the alterations in clotting and thrombin seen in covid- may well provide a mechanism for this. similarly, thrombin has been related to the induction of preterm labour and weakened fetal membranes by induction of decidual colony-stimulating factor (csf)- ( ). at present, there are no drugs, therapeutics, or vaccines approved for curing, preventing, or treating sars-cov- specifically. as of june , a total of ( in human trails, in preclinical development) therapeutic drugs are under development against covid- . the current treatment for sars-cov- patients involves the management of clinical symptoms and providing supportive care. while research into developing new drugs and treatments against sars-cov- are ongoing, much of the effort currently focuses on the repurposing existing medicines used against viruses, multiple sclerosis, arthritis, blood plasma derivatives and malaria. moreover, although immunosuppressive treatments, e.g. corticosteroids have shown promise for covid- , there is considerable concern about possible side effects. other immunotherapeutic approaches given as adjunct therapy and based on neutralizing inflammatory cytokines and other immunomodulators, passive viral neutralization to reduce lung pathology and viral load, could be a promising approach ( ) . a number of these approaches are discussed below. the antiviral drug remdesivir, developed by gilead sciences, is an adenosine analogue, which incorporates into nascent viral rna chains and results in premature termination, effectively inhibiting viral rna synthesis ( ) . it was developed for the treatment of ebola and marburg virus infections ( ) , and animal studies have shown that it is effective against the other coronavirus ( ) . in vitro studies have established its efficacy against sars-cov- ( ) . an open-label trial across the united states, europe, canada and japan showed clinical improvement of of the covid- patients who were treated with a -day course of remdesivir on a compassionate basis ( ) . however, a follow-up multi-centre, randomized, double-blinded, placebo-controlled trial of patients showed that the drug was not associated with a difference in time to clinical improvement. compared to the placebo, the drug was found to have a non-significant but, numerically faster time to clinical improvement in patients with a symptom duration of days or lower ( ) . currently, japan and the usa have allowed the use of the drug under emergency use authorization for the treatment of covid- . in a randomized, open-label, multi-centre phase clinical trials, a -day course remdesivir brought about a significant clinical improvement compared to standard alone in patients with moderate covid- . this clinical study assessed the effect of -day (n= ) and -day (n= ) investigational remdesivir courses plus standard of care, versus standard of care alone (n= ) on clinical improvement on day ( ) . in case of patients with severe disease, both day and courses of the drug have been found to have similar clinical outcomes, but as the study lacked placebo control, the magnitude of benefit cannot be determined ( ) . umifenovir, marketed as arbidol, is a derivative of indole carboxylic acids used for the treatment of influenza a and b virus infection, and other arboviruses ( ) . it functions by incorporating into cell membranes and interfering with the hydrogen bonding network of phospholipids, blocking both the fusion of the virus to the cell membrane and the virusendosome fusion ( ) . in vitro studies have established anti-viral efficacy against ebola virus, human herpesvirus , hepatitis c virus, tacaribe arena virus, sars-cov and sars-cov- ( ) ( ) ( ) . a retrospective study on sars-cov- patients treated with umifenovir did not reveal any improvement in clinical prognosis or accelerated viral clearance ( ) . currently, two randomized and open-label trials to determine the safety and efficiency of the drug are ongoing in china. favipiravir, another anti-viral drug, developed by fujifilm toyama chemical (as avigan) and zhejiang hisun pharmaceutical, is a pyrazinecarboxamide derivative. it is converted into an active phosphoribosylated form (favipiravir-rtp) in cells and is recognized as a substrate by viral rna polymerase, thereby blocking the activity of rna-dependent rna polymerase. it was developed as a treatment against influenza. the drug is currently approved for the treatment of sars-cov- in china and italy. a study with sars-cov- patients treated using the drug has reported that better efficacy was observed in anti-viral activity and lower adverse reactions compared to the control group that was treated with lopinavir/ritonavir ( ) . another prospective, multi-centre, open-label, randomized superiority study with sars-cov- infected patients was conducted at three hospitals. they showed faster recovery from clinical symptoms when compared to the controls that were treated with umifenovir, even though similar numbers required the use of ventilators and oxygen ( ) . there are currently six trials ongoing in china evaluating the efficiency of this drug against other antivirals for the treatment of covid- and a phase clinal trial to assess its effectiveness and safety is scheduled in japan and usa. anti-malaria drugs, chloroquine and hydroxychloroquine, are lysosomotropic agents that function by increasing late endosomal and lysosomal ph, which results in impaired viral release from the endosome or lysosome ( ) ( ) ( ) . in vitro studies have shown antiviral activity against sars-cov- with hydroxychloroquine, a weak diprotic base, to have higher potency against the virus ( , ) . in sars-cov- , chloroquine, along with its lysosomotropic activity, is believed to reduce glycosylation of ace affecting the binding of the virus to the cells ( ) . furthermore, chloroquine is also shown to block the production of proinflammatory cytokines such as il- preventing ards ( ); hydroxychloroquine was found to possess an anti-inflammatory effect on th -related cytokines (il- , il- and il- ) ( ) . initial clinical studies in china involving sars-cov- infected patients, who were treated with chloroquine, showed amelioration of pneumonia, shortened disease progression, increased resolution of lung lesions on ct, and a better virus-negative conversion ( , ) . hydroxychloroquine and combination therapy with azithromycin was found to reduce viral load in a french open-label non-randomised clinical trial and in an observational pilot study ( , ) . nevertheless, these studies were plagued with several limitations, such as small sample size, very short observation period, no randomisation, lack of reports on clinical progression, poorly described inclusion and exclusion criteria, and low national early warning score ( , , ) . another trial with sars-cov- infected patients treated with hydroxychloroquine for seven days in china and a study with effectively sars-cov- patients, revealed no significant difference in the nasopharyngeal viral carriage when compared to the controls that were provided with the local standard care ( , ) . a third randomized clinical trial conducted in china with patients exhibiting mild sars-cov- when treated with hydroxychloroquine were found to have recovered faster from cough and fever when compared to the placebo. however, the result of this study cannot be extrapolated to patients with severe sars-cov- ( ). a retrospective cohort study of random sample of inpatients with laboratory-confirmed sars-cov- admitted to hospitals in the new york city was conducted. it did not find any significant differences in in-hospital mortality associated with the treatment with hydroxychloroquine, azithromycin, or both, compared to the controls where the patients were given neither of the drugs ( ). the us fda and european medicines agency (ema) and many other countries like india and poland have authorized emergency use of hydroxychloroquine to treat sars-cov- infected patients. however, the fda and ema have issued warnings against the reported side effects of the drugs. these include abnormal electrical activity that affects the heart rhythm (qt interval prolongation, ventricular tachycardia, and ventricular fibrillation), particularly when taken at high doses or in combination with the antibiotic azithromycin. other side effects reported are liver and kidney problems, nerve cell damage that can lead to seizures and hypoglycaemia ( , ) . around clinical trials have been registered to study the effects of hydroxychloroquine and chloroquine independently or in combination with each other on sars-cov- have been registered in the usa and china ( ) . another anti-parasitic drug, ivermectin, has been shown to be effective against sars-cov- in vitro ( ) . a clinical trial to assess the efficiency of ivermectin against sars-cov- has been planned to take place in japan soon. the corticosteroid, dexamethasone, functions as an immunosuppressant. the drug is believed to modulate the lung injury caused by a dysregulated immune system, thereby reducing the progression to respiratory failure and death ( ) . in a randomized, controlled, open-label, adaptive, platform trial, , patients treated with mg of dexamethasone (orally or intravenously) for days were found to have a significantly reduced day mortality rate among those receiving mechanical ventilation by . %, and by % among those receiving oxygen without mechanical ventilation, compared to , patients treated with standard care ( ) . treatment with the drug did not provide any benefit to patients who did not require oxygen or mechanical ventilation, hinting at possible harm. the use of corticosteroid in the case of severe respiratory infections requires the use of "the right dose, at the right time, in the right patient" ( ) . this is because a high or an early dose may help the virus proliferate by suppressing the immune system, instead of reducing inflammation. in case of covid- , the peak of viral shedding is higher early in the disease. the benefit of dexamethasone when patients require respiratory support or after the first week of the disease suggest that this stage is dominated by an irrepressible immune response versus active viral replication ( ) . dexamethasone is the first drug found to reduce mortality in covid- ( ). lopinavir/ritonavir is a drug combination. lopinavir is a protease inhibitor, developed by abbott laboratories against hiv- that functions by blocking essential viral proteases ( ) . due to poor pharmacokinetics, it is administered exclusively in combination with ritonavir which increases lopinavir's plasma half-life through inhibition of cyp a-mediated metabolism of lopinavir, thereby increasing its exposure and improving the anti-viral activity of the drug ( ) . in vitro studies have revealed that lopinavir inhibited the replication of the sars-cov- virus in vero e cell ( ) . in a randomized, controlled, open-label trial with patients with laboratory-confirmed sars-cov- infection, no benefit was observed with lopinavir-ritonavir treatment beyond standard care ( ) . another single-blind randomised controlled trial in china treated patients with mild/moderate covid- for days, or umifenovir or standard care with no antiviral ( ) . in the study, no differences were found in the time taken for viral clearance, as assessed by pcr of nasopharyngeal swabs, fever, cough, or lung ct findings. clinical status deterioration to severe/critical from mild or moderate clinical status and gastrointestinal side effects was seen highest in patients treated with lopinavir/ritonavir when compared to umifenovir treated or those treated with standard care and no antivirals ( ) . both these randomised clinical trials suffer from small sample sizes and lack of blinding. a multi-centre, prospective, open-label, randomised, phase trial in hong kong with sars-cov- infected patients involved treatment for days with only lopinavir-ritonavir (control), or with a combination of lopinavir-ritonavir, ribavirin, an oral hepatitis c virus drug, and ifn-. it found that the combination treatment was effective in reducing symptoms and viral shedding faster, as well as duration of hospital stay ( ) . currently, about a dozen trials are studying the effect of the drug against sars-cov- . one such study is a phase randomized controlled trial in china in which the effectiveness of lopinavir-ritonavir against influenza drugs, umifenovir and oseltamivir, is to be studied. another south korean trial is looking to compare the efficacy of lopinavir-ritonavir against hydroxychloroquine. the who solidarity trial and uk-based recovery trial is looking to study the effectiveness of lopinavir-ritonavir independently; the who solidarity trial also looks to the explore the drug in combination with interferon-. another second-generation protease inhibitor against hiv- , darunavir, has shown significant inhibition of sars-cov- replication (in vitro). according to a press release by johnson & johnson, an unpublished single-centre, open-label, randomized, and controlled trial in china in which sars-cov- patients were treated with darunavir and cobicistat was not effective in treating sars-cov- ( ). however, a further three clinical studies in china are scheduled. other drugs currently being tested against sars-cov- include tocilizumab, a monoclonal antibody against il- developed by roche, which is used for the treatment of moderate to severe rheumatoid arthritis by blocking il- activity. the drug was found to have helped cure of covid- patients in a trial conducted in china ( ) . another open multi-centre randomized controlled trial french study awaits publication, in which patients were split into two groups, i.e. routine treatment with and without tocilizumab: in the group treated with tocilizumab, the combination of ventilation requirement (mechanical or non-invasive) or death was achieved in a significantly lower proportion of patients ( ) . a phase trial to test its efficacy in treating patients with severe covid- has been authorised by the fda. moreover, an italian multi-centre, retrospective study of patients with severe covid- pneumonia, revealed that the use of tocilizumab given either intravenously or subcutaneously was associated with reduced risk of mechanical ventilation and death ( ) . anakinra is a recombinant il- receptor antagonist that has shown promise in treating severe covid- disease. in a retrospective cohort study of patients with covid- and ards that were managed with non-invasive ventilation (outside the icu), their treatment with high-dose anakinra was observed to be safe and associated with clinical improvement in % of patients ( ) . another study has also described the early use of anakinra in covid- patients with cytokine storm syndrome (css) and acute hypoxic respiratory failure (ahrf) which may be beneficial in preventing the need for mechanical ventilation ( ) . these results have encouraged further clinical trials to validate its safety and efficacy ( ) . approaches targeting inhibition of bruton tyrosine kinase (btk) has also shown promise. btk plays a significant role in human innate immune responses. tlrs recognize ssrna of viruses like sars-cov- and induce signalling via btk-dependent activation of nf-κb, initiating a pro-inflammatory response ( ) ( ) ( ) ( ) . btk also plays a key role in the activation of the nlrp inflammasome, resulting in maturation and secretion of il- β, a key pro-inflammatory cytokine ( ) ( ) ( ) . thus, btk seems a favourable target against the cytokine storm in covid- . in one study, acalabrutinib (a selective inhibitor of btk) was given to patients with severe covid- and clinical improvements were observed over a -week treatment period, with reduced biomarkers of inflammation (c-reactive protein and il- ) to normal levels ( ) . other dual inhibitors e.g. ibrutinib which target btk/il- -inducible t-cell kinase (itk) signalling have also shown promise ( ) . in one study of patients given ibutinib for treatment of b-cell malignancies and chronic graft-versus-host disease (cgvhd), there was evidence that ibutinib may also protect against pulmonary injury in covid- , which these patients subsequently had, suggesting ibutinib as a possible prophylactic for vulnerable patient groups ( ) . similar findings demonstrating a possible protective role of btk inhibitors in cancer with covid- have also been subsequently described ( ) ( ) ( ) . these promising findings now merit a controlled randomised trial to demonstrate efficacy and drug safety of these btk inhibitors. intravenous immunoglobulin (ivig) is a pooled preparation of normal igg obtained from several thousand healthy donors. it is generally used in the immunotherapy of several autoimmune and inflammatory diseases, ( ) , and thus has been investigated for treating covid- to mitigate the css. ivig therapy has shown promise through several studies, although careful selection of covid- patients and timing of ivig administration appear to be the key for good clinical outcome. preliminary findings from one multi-centre study showed that early administration of high dose ivig improved the prognosis of critical patients with covid- ( ) . similarly, patients with severe covid- who received high-dose ivig made a satisfactory recovery ( ) . in another study, the use of ivig as an adjuvant treatment for covid- pneumonia within hours of admission to the icu reduced the use of mechanical ventilation, icu and hospital time, and the -day mortality rate of patients with severe covid- pneumonia ( ) . in a case study of a covid- patient with respiratory failure and shock, treatment with plasma exchange before ivig treatment resulted in prompt recovery without the need for mechanical ventilation and may be an additional early treatment step to treat critically ill covid- patients ( ) . ivig treatment of severely-ill covid- patients on mechanical ventilation has also shown promise. in one study of patients, treatment with ivig improved clinical and respiratory outcome, particularly saturation o levels, resulting in earlier extubation of the patients ( ) . furthermore, ct graphs obtained after ivig therapy also revealed improvements in pulmonary lesions of these patients ( ) . convalescent plasma therapy (cpt) is another classical adaptive immunotherapy used for the treatment of infectious disease for over a century. it has currently been approved for covid- by the fda under compassionate use rules. the treatment involves the transfusion of high neutralizing antibody titre containing blood plasma from sars-cov- recovered patients. this provides immediate short-term immunity. this is accomplished by binding of the pathogen to the antibody, which results in the activation of the immune system causing cellular cytotoxicity, phagocytosis, or direct pathogen neutralisation. five clinical studies, conducted involving covid- patients who were treated with cpt, revealed significantly lower viral titres, increased levels of neutralizing antibody, improved clinical symptoms such as apyrexia, resolved ards and unassisted breathing ( ) ( ) ( ) ( ) ( ) . among the cpt-treated patients, no fatalities were recorded, and no severe adverse reactions or treatment complications associated with cpt were reported ( ) ( ) ( ) ( ) ( ) . while providing with valuable initial data, these studies suffer from several limitations such as lack of proper control groups, non-randomized evaluations, concomitant drug treatments, poor participant selection, lack of proper cpt dosage, and duration of therapy ( ) . three clinical trials are currently being evaluated by the fda to test the safety and efficiency of cpt in patients who have been exposed to the virus and are at high risk of developing severe covid- , patients who are admitted in hospital with acute respiratory symptoms, and for covid patients under mechanical ventilation ( ) . further trials are also planned or ongoing in china, columbia, iran, mexico and the netherlands ( ) . early safety indicators of covid- cpt were evaluated in a study of , patients and showed that the mortality rate was not unduly high and concluded that transfusion of convalescent plasma appears safe in hospitalized patients with covid- ( ) . while the repertoire of antivirals and repurposed drugs tested against sars-cov- are expected to help manage the disease, the development of a safe and effective vaccine would help cut down the overall number of deaths and prevent the population from getting the disease in the first place. a recent study suggested that mandatory bcg vaccination can possibly be associated in flattening the curve in the spread of covid- . it analysed the rate of day-wise increase in positive cases in countries and deaths in countries for the first -day period ( ) . while arguments for the potentially beneficial effects of pre-existing vaccines have been sporadically made, including giving mmr (mumps, measles and rubella) vaccines to elderly population, generating a sars-cov- specific vaccine seems a logical and obligatory choice. as of july , the who landscape document reports candidate vaccines developed on various platforms ( figure ) in preclinical stages of development: only are under clinical evaluation. mrna- vaccine is a sequence optimized mrna/lnp expressing a perfusion stabilized form of sars-cov- s- p a transmembrane anchored protein with the native furin cleavage site, developed by moderna in collaboration with the national institute of allergy and infectious diseases vaccine research center ( , ) . the vaccine is undergoing an openlabel phase clinical trial that started in march, with healthy adult ( - -year-old) volunteers for six weeks in three dose cohorts ( µg, µg and µg) as two doses approximately days apart via intramuscular injection in the upper arm. three cohorts of - -year-old volunteers and three cohorts of healthy volunteers aged and above are being enrolled in addition to the initial volunteers. the volunteers will be followed through months after the second vaccination to assess safety data, common vaccination symptoms, review trial data and advise niaid ( ) . a phase ii trial with healthy participants in two cohorts ( - years old adults and adults aged years and above) treated with a placebo, a μg or a μg dose has begun from may, th , . the in vivo studies in murine models suggested the vaccine to be immunogenic and could elicit igg a and igg subclass s-binding antibodies. mrna- immunized mice splenocytes showed higher secretion of ifn- than il- , il- or il- upon re-stimulation with peptide pools (s and s ). a dose of μg of mrna- was found to induce robust cd + t cell response to the s peptide pool with balanced th /th ab isotype response in mice. thus, a μg dose of vaccine has been decided for human trial in phase study, which is equivalent to μg dose induced in mice ( ) . the fda has granted fast track designation for the vaccine. the pfizer licensed biontech's bnt vaccine development programme has developed four coronavirus vaccine candidates ( ) . two of the vaccines contain mrna coding for the spike protein of sars-cov- , while the other two contain only the rbd of the spike protein ( ) . furthermore, the four vaccine candidates are made of three different mrna formats. two of the vaccine are based on nucleoside modified mrna (modrna), which incorporates modified nucleosides in the mrna ( ). this suppresses intrinsic immune activation and the production of anti-drug antibodies against the mrna itself ( ). the suppressed immune activity against the therapeutic mrna helps produce the antigenic protein for longer periods ( ). the next vaccine candidate is based on the optimised unmodified mrna (urna) format ( ). urna uses uridine in the mrna, making it more immunogenic ( ). finally, the last vaccine candidate uses self-amplifying mrna (sarna) ( ). it is based on the principle of viral replication. the sarna, in addition to encoding a protein of interest, also encodes, replicase ( ). this enables the self-amplification of the mrna inside the cell ( ). the dsrna intermediate created during the replication of the rna triggers an immune response making sarna a potent activator of the immune system ( ). a phase / , randomized, placebo-controlled, observer-blind, dose-finding, and vaccine candidate-selection study to evaluate the safety, tolerability, immunogenicity, and potential efficacy of the candidate in healthy adult volunteers is ongoing ( ). another frontrunner among the candidates is cansino bio's ad -ncov ( ) . it is a genetically engineered vaccine candidate with the replication-defective adenovirus type (live virus) as the vector to express sars-cov- spike protein. this would help the body to produce neutralizing antibodies against sars-cov- . it has been shown to induce a strong anti-viral activity against sars-cov- in animal and in vitro studies. a single-centre, non-random, open, and dose-escalation phase i clinical trial for recombinant novel coronavirus vaccine (adenoviral vector) in healthy adults aged between and years were conducted. the vaccine has been administered as a liquid formulation intramuscularly in the deltoid muscle ( ) . three different doses were chosen: (a) low dose of x viral particles/ . ml; (b) intermediate dose of . x viral particles/ml; and (c) high dose combines both low and intermediate dose (one in each arm). the volunteers are assessed for a period for months to study any adverse reactions or other relevant outcomes ( ) . most common systematic adverse reaction observed were fever, fatigue, headache and muscular pain but with no serious adverse effect were noted within days. participants showed four-fold increase in anti-rbd antibodies in all the groups; neutralizing antibodies increased gradually being highest at days post vaccination. ad neutralizing antibody titres were boosted significantly postvaccination. il- and tnf- were detected and polyfunctional memory cd + t cell phenotypes were higher than cd + t cells. this suggested ad vectored covid- vaccine to be immunogenic and capable of stimulating both b and t cell response. for phase clinical trial, intermediate dose was chosen and is expected to be completed by january ( ) . the vaccine may have some negative effects in older age people thus in the nd clinical trial participants above years will be included. t cell response peaked earlier from th day after the st shot of vaccine whereas the antibodies production level peaked at th day post vaccination. the study also highlighted the possibility of negative effect on vaccine elicited immune response due to pre-existing ad immunity ( ) . chadox -ncov is being developed by oxford university, uk ( ) . it is a replication deficient simian adenovirus vector chadox , containing full length s-protein of sars cov- along with a tissue plasminogen activator leader sequence. the vaccine is reported to be effective in inducing an antiviral response in animal models ( ) . chadox -ncov was found to be immunogenic in mice mounting robust anti-viral response. single dose of this vaccine was capable of inducing humoral and cellular immune response in rhesus macaques ( ) . a phase i/ii single-blinded, randomised, multi-centre study to determine efficacy, safety and immunogenicity of the vaccine in about healthy adult volunteers aged - years was initiated on april rd , ( ). the volunteers have been subjected to either one dose of x vp of chadox ncov- , an additional booster dose of . x vp of chadox ncov- , or a control of menacwy vaccine delivered intramuscularly ( ). the volunteers were assessed for a period for months to study any adverse reactions or other relevant outcomes ( ) . the results showed increase in s-specific antibodies with a single dose by th day and increase in neutralizing antibodies with booster dose in all participants. chadox -ncov was also capable of inducing heightened effector t-cell response quite earlier than antibody response. t cell response peaked on day th and sustained up to days. the results showed chadox ncov- vaccine to be safe, tolerant and immunogenic, which further supported phase trial which is now underway ( ) . picovacc is a purified inactivated sars-cov- vaccine candidate which is capable of inducing neutralizing antibodies in mice, rats, and nonhuman primates specific to sars-cov- . cn strain of sars cov- virus was chosen to develop picovacc which was inactivated with β-propiolactone. this inactivated vaccine candidate was able to produce about -fold higher s-specific antibody titres in murine model when compared to covid- recovered patients. efficacy of picovacc was also tested in rhesus macaques with an intramuscular low ( . µg), medium ( µg) and high ( µg) dose administered three times ( , th and th day) and on day nd sars cov- cn strain was inoculated through intratracheal (lungs) route. all vaccinated macaques showed protection towards sars cov- infection and their viral loads declined significantly. no notable haemato-and histopathological changes were observed; human clinical trials are awaited ( ) . a group of us scientists have come up with a series of prototype dna vaccines expressing variants of the sars-cov- spike protein. the efficacy of the dna vaccine candidates was evaluated in rhesus macaques ( - -year-old). intramuscular dose ( mg) of dna vaccine was administered, followed by booster dose on rd week and antigenic challenge ( . x viral particles) on th week (both intranasal and intratracheal route). dna vaccine was found to be protective with dramatic reduction of viral replication and enhanced production of sspecific binding as well as neutralizing antibodies compared to controls. the study has not yet addressed the possibility of mutations that may emerge in escaping neutralizing antibodies, though it seems to be protective in primates against sars-cov- ( ). j o u r n a l p r e -p r o o f ino- , developed by inovio, is a dna vaccine candidate ( ) . the optimized spike protein of sars-cov- virus dna plasmids are introduced into cells by the use of a proprietary platform, cellectra®, via electroporation ( ) . once inserted, the plasmids are expected to strengthen the body's own natural response. a phase i open-label study to evaluate the safety, tolerability and immunogenicity of ino- as a prophylactic vaccine against sars-cov- in healthy volunteers aged - years is ongoing ( ) . the volunteers will be treated with either one or two doses of mg of vaccine administered intradermally followed by electroporation the following day ( ) . the volunteers will be assessed for a period for year to study any adverse reactions or other relevant outcomes ( ) . once the initial safety and immunogenicity of the vaccine are satisfied, phase ii efficacy studies are planned. qualitative and quantitative properties of cd + and cd + t cell responses in covid- and prophylactic vaccine development necessitate identifying viral regions and potential epitopes. thus, a total of peptides ( -to -mer), which span the full proteome of the sars-cov- excluding orf- , were designed and used to assess the memory t cell responses upon challenge on patients following recovery from covid- . peptides were identified containing cd + and/or cd + epitopes. the memory of t cell responses from convalescent individuals with covid- was found to be greater in severe covid- cases compared to mild ones. immunodominant epitope clusters and peptides were most markedly observed to belong to spike, m, and orf proteins. in about % of study groups, strong cd + t cell responses specific to the np protein were observed, suggesting the possibility of inclusion of non-spike proteins in future covid- vaccine design ( ) . in another study, a comprehensive immunogenicity map of the sars-cov- virus was carried out; peptide sequences ( -mers) were generated based on computational algorithms. a single -mer peptide containing multiple epitopes that can possibly present on hla class i and class ii across majority of population and provide long-term immunity in most people acting as b and t cell epitopes had been identified. this in silico analysis needs further evaluation for safety and efficacy as a vaccine ( ) . in an unprecedented short span of time and speed since the beginning of the covid- pandemic, significant progress has been made in our understanding of the pathogenesis of sars-cov- infection. however, there are endless unanswered questions; hopefully and most likely, they will be answered in near future. why there are a huge population that are asymptomatic carriers? what are the genetic contributors to susceptibility and resistance to developing covid- ? how pregnant women are so resilient to developing covidsymptoms; for that matter young children as well! what happens during the period of latency, i.e. between being infected and showing symptoms? how far the lung surfactant system gets affected during severe symptoms? what triggers thrombotic microangiopathy in addition to complement activation. on the adaptive immune aspects, what variations exist within population in terms of the proportion of neutralising antibodies? persistence of neutralising antibodies and recall memory magnitude following second infection (on vaccination trials) will yield serious information about how to finetune the dose, duration and vaccination strategies. in this acute crisis, a number of existing drugs have been repurposed empirically; clinical trials have yielded variable results. it is becoming clear that combination therapies are more likely to be successful. deciphering, at high resolution, the mechanisms and consequences of hostpathogen interactions in covid- will lead to novel therapies and preventative vaccination strategies. primary cellular host and co-receptor for sar-cov- . ) attachment and entry of sar -cov- requires priming by transmembrane serine protease (tmprss ) which cleaves the s protein into s and s portions, facilitating, ), s targeting and binding of the receptor angiotensin-converting enzyme (ace ), followed by receptor-mediated endocytosis of the virion into the host cell. j o u r n a l p r e -p r o o f tmprss is the key protease involved in priming sars-cov- , which forms a receptorprotease complex with ace on the host cell surface, thus facilitating viral targeting and entry to the host cell. co-expression of aec and tmprss has been found in proximal as well in distal airways. the nasal cavity has the highest expression of both the receptors in ciliated and secretory (goblet) cells compared to lung bronchi (ciliated and secretory cells) and lung parenchyma (alveolar type progenitor cells, at ). structural conformation of receptor-binding domain (rbd) present in s region of sars-cov- spike protein is capable of influencing the ace -binding affinity. in case of sars-cov- , the rbd contains a four-residue motif glycine-valine/glutamine-glutamate/threonineglycine which enables the binding loop to take a different conformation. it can undergo two possible conformational changes, a "lying down state" which has low affinity towards aec and a "standing up state" with high binding affinity. sars-cov- rbd is found mostly in lying down state, and thus being less accessible to aec . this hidden conformation of rbd in the spike protein can possibly be a masking strategy for immune evasion by sars-cov- . 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hydroxychloroquine decreases th -related cytokines in systemic lupus erythematosus and rheumatoid arthritis patients a systematic review on the efficacy and safety of chloroquine for the treatment of covid- breakthrough: chloroquine phosphate has shown apparent efficacy in treatment of covid- associated pneumonia in clinical studies clinical and microbiological effect of a combination of hydroxychloroquine and azithromycin in covid- patients with at least a six-day follow up: a pilot observational study hydroxychloroquine and azithromycin as a treatment of covid- : results of an open-label non-randomized clinical trial no evidence of rapid antiviral clearance or clinical benefit with the combination of hydroxychloroquine and azithromycin in patients with severe covid- infection a pilot study of hydroxychloroquine in treatment of patients with common coronavirus disease- (covid- ) efficacy of hydroxychloroquine in patients with covid- : results of a randomized clinical association of treatment with hydroxychloroquine or azithromycin with in-hospital mortality in patients with covid- in new york state fda drug safety communication: fda cautions against use of hydroxychloroquine or chloroquine for covid- outside of the hospital setting or a clinical trial due to risk of heart rhythm problems chloroquine diphosphate in two different dosages as adjunctive therapy of hospitalized patients with severe respiratory syndrome in the context of coronavirus (sars-cov- ) infection: preliminary safety results of a randomized, double-blinded, phase iib cl hydroxychloroquine and covid- the fda-approved drug ivermectin inhibits the replication of sars-cov- in vitro effect of dexamethasone in hospitalized patients with first drug to reduce mortality in hospitalised patients with respiratory complications of covid- found epigenetic gene silencing underlies c-fiber dysfunctions in neuropathic pain remdesivir, lopinavir, emetine, and homoharringtonine inhibit sars-cov- replication in vitro a trial of lopinavir-ritonavir in adults hospitalized with severe covid- triple combination of interferon beta- b, lopinavir-ritonavir, and ribavirin in the treatment of patients admitted to hospital with covid- : an open-label, randomised, phase trial lack of evidence to support use of darunavir-based treatments for sars-cov- clinical trial analysis of -ncov therapy registered in china le tocilizumab améliore significativement le pronostic des patients avec pneumonie covid moyenne ou sévère | aphp tocilizumab in patients with severe covid- : a retrospective cohort study interleukin- blockade with high-dose anakinra in patients with covid- , acute respiratory distress syndrome, and hyperinflammation: a retrospective cohort study use of anakinra to prevent mechanical ventilation in severe covid- : a case series emerging pharmacotherapies for covid- macrophages eat cancer cells using their own calreticulin as a guide: roles of tlr and btk bruton's tyrosine kinase is required for apoptotic cell uptake via regulating the phosphorylation and localization of calreticulin bruton's tyrosine kinase regulates tlr / -induced tnf transcription via nuclear factor-κb recruitment btk operates a phospho-tyrosine switch to regulate nlrp inflammasome activity bruton's tyrosine kinase is essential for nlrp inflammasome activation and contributes to ischaemic brain injury human nacht, lrr, and pyd domain-containing protein (nlrp ) inflammasome activity is regulated by and potentially targetable through bruton tyrosine kinase inhibition of bruton tyrosine kinase in patients with severe covid- btk/itk dual inhibitors: modulating immunopathology and lymphopenia for covid- therapy the btk inhibitor ibrutinib may protect against pulmonary injury in covid- -infected patients btk inhibitors in cancer patients with covid- : "the winner will be the one who controls that chaos treating leukemia in the time of covid- protective role of bruton tyrosine kinase inhibitors in patients with chronic lymphocytic leukaemia and covid- ivig-mediated effector functions in autoimmune and inflammatory diseases clinical efficacy of intravenous immunoglobulin therapy in critical patients with covid- : a multicenter retrospective cohort study high-dose intravenous immunoglobulin as a therapeutic option for deteriorating patients with coronavirus disease effect of regular intravenous immunoglobulin therapy on prognosis of severe pneumonia in patients with covid- successful treatment with plasma exchange followed by intravenous immunoglobulin in a critically ill patient with covid- recovery of severely ill covid- patients by intravenous immunoglobulin (ivig) treatment: a case series treatment of critically ill patients with covid- with convalescent plasma effectiveness of convalescent plasma therapy in severe covid- patients treatment with convalescent plasma for covid- patients in wuhan use of convalescent plasma therapy in two covid- patients with acute respiratory distress syndrome in korea treatment with convalescent plasma for critically ill patients with severe acute respiratory syndrome coronavirus infection convalescent plasma transfusion for the treatment of covid- : systematic review the resurgence of convalescent plasma therapy early safety indicators of covid- convalescent plasma in , patients mandated bacillus calmette-guérin (bcg) vaccination predicts flattened curves for the spread of covid- world health organization. draft landscape of covid- candidate vaccines - phase i, open-label, dose-ranging study of the safety and immunogenicity of -ncov vaccine (mrna- ) in healthy adults sars-cov- mrna vaccine development enabled by prototype pathogen preparedness a close look at the frontrunning coronavirus vaccines as of may (updated) chinese clinical trial registry. a randomized, double-blinded, placebo-controlled phase ii clinical trial for recombinant novel coronavirus ( -ncov) vaccine (adenovirus vector) safety, tolerability, and immunogenicity of a recombinant adenovirus type- vectored covid- vaccine: a dose-escalation, open-label, non-randomised, first-in-human trial chadox ncov- vaccination prevents sars-cov- pneumonia in rhesus macaques a study of a candidate covid- vaccine (cov ) -full text view -clinicaltrials.gov safety and immunogenicity of the chadox ncov- vaccine against sars-cov- : a preliminary report of a phase / , single-blind, randomised controlled trial development of an inactivated vaccine candidate for sars-cov- dna vaccine protection against sars-cov- in rhesus macaques tolerability and immunogenicity of ino- for covid- in healthy volunteers broad and strong memory cd + and cd + t cells induced by sars-cov- in uk convalescent covid- patients identification of sars-cov- vaccine epitopes predicted to induce long-term population-scale immunity key: cord- -nsdhvc w authors: maki, dennis g. title: sars revisited: the challenge of controlling emerging infectious diseases at the local, regional, federal, and global levels date: - - journal: mayo clin proc doi: . / . . sha: doc_id: cord_uid: nsdhvc w nan i n , a blue-ribbon panel was commissioned by the institute of medicine of the national academy of sciences to advise the us government on emerging infectious diseases that posed a threat or potential threat to the health of people living in the united states and assist in the federal allocation of public health resources. , emerging infections were defined as infections caused by newly identified human pathogens-such as the legionella bacillus, the reemergence of previously controlled pathogens-such as measles, or the appearance of anti-infective resistancesuch as methicillin-resistant staphylococcus aureus, the incidence of which had increased significantly within the past decades or threatened to increase in the near future. since the outbreak of severe pneumonia in us veterans attending a convention at the bellevue-stratford hotel in , found months later to have been caused by a previously unknown and remarkably ubiquitous waterborne bacterium, legionella pneumophila, more than dozen human pathogens have been identified as agents of emerging infectious diseases in the united states (table ) . the most recent and perhaps most fearsome emerging infections are the appearance of west nile virus encephalitis in new york city in and its rapid spread westward ; inhalation anthrax, deriving from use of bacillus anthracis spores as a biologic weapon against the us civilian population in ; the global outbreak of severe acute respiratory syndrome (sars) in ; and the looming threat of pandemic influenza, especially global disease caused by the highly virulent avian subtype a (h n ). [ ] [ ] [ ] during the past months, mayo clinic proceedings has published reviews of diseases caused by of these emerging pathogens, west nile virus, the sars coronavirus (sars-cov), and avian influenza virus. efforts to better inform readers about the consequences of emerging infectious diseases continue in this issue of the proceedings, in which chiang et al report a study of cases of nosocomial sars acquired in taipei hospitals during . most of the patients, of whom were followed up for at least months, were health care workers, and because few had underlying diseases, all except survived; thus, this study provides some of the best data on the long-term effects of sars on the lung. the clinical features and natural history of sars encountered by chiang et al are similar to those reported in much larger cohorts. , all their patients had fever, and most had cough and dyspnea as well; however, % had diarrhea, and % had myalgias, indicative of the severe systemic immunoinflammatory response to this unique new human infection. , similarly, all their patients had lymphopenia, and most had elevated levels of lactate dehydrogenase, now well-defined surrogate laboratory markers for patients presenting with sars. , however, chiang et al also found that patients with a very high c-reactive protein or lactate dehydrogenase level at the outset were far more likely to have progression to a severe stage of disease requiring mechanical ventilatory support, information of value to clinicians who might be called on to manage patients with sars in the not-too-distant future. most interestingly, chiang et al show that whereas bilateral fibrotic changes were demonstrable by high-resolution computed tomographic imaging to months after the acute infection, most of the survivors showed near-normal spirometric lung volumes (forced vital capacity, forced expiratory volume in second), albeit one third with reduced diffusion capacity, but none required home oxygen. it is likely that most of these individuals will have recovery of normal lung function. these findings are very similar to the well-documented long-term pulmonary effects of garden-variety acute respiratory distress syndrome, stemming from overwhelming pneumonia, gastric aspiration, near drowning, trauma, pancreatitis, or systemic sepsis, in which most survivors have gratifying recovery of lung function in the early years after the acute episode. [ ] [ ] [ ] because sars is such a unique human viral infection and induces such an unusually severe systemic inflammatory response, , it will be important to closely follow survivors of severe sars for considerably longer to be certain that latently expressed progressive pulmonary fibrosis does not occur. between the first reports from the world health organization and the centers for disease control and prevention (cdc) that defined sars as a global threat in march , and control of the epidemic in southeast asia and north america months later, more than persons in the republic of china, hong kong, singapore, vietnam, taiwan, and canada became infected, and ( . %) died of the infection ; mortality exceeded % in patients older than years. , since the last communication on sars in the proceedings in april , there have been extraordinary advances in our understanding of the disease in terms of its pathogenesis, epidemiology, management, and control: editorial • like many of the emerging human pathogens such as borrelia burgdorferi (mice, deer), us hantavirus (mice), west nile virus (wild birds), variant creutzfeldt-jakob prions (cattle, potentially free-ranging cervids), monkeypox virus (exotic african rodents and primates, us prairie dogs), and avian influenza virus a (h n ) (edible birds), infection by sars-cov can legitimately be considered a zoonosis, and wild mammals formed the initial reservoir of the virus-sars-cov has been isolated from palm-top civets and raccoon dogs in the live animal markets in southern china, and purveyors of these animals commonly show asymptomatic sars-cov seropositivity. , the genomic evidence that sars-cov is an animal-human recombinant is compelling, [ ] [ ] [ ] [ ] and sars-cov appears to have had a biologic origin remarkably similar to the human influenza a viruses and human immunodeficiency virus. although the sars virus originated in animals and made the leap to humans, once established in the human population, the virus has spread rapidly person-toperson, and the major reservoir of human disease is humans in the early stage of infection, before they have been diagnosed as having the virus and placed in respiratory and barrier isolation. • elegant molecular epidemiology has traced the origins of sars-cov to foshan in guangdong province, southern china, from whence it spread to beijing, then to hong kong, and from hong kong to vietnam, singapore, and canada. , a -year-old chinese physician who traveled to hong kong on march , , where he spent only day, appears to have been the source of sars-cov that ultimately resulted in thousands of cases of sars in countries and continents. • sars-cov is a new human pathogen to most of the world. studies by the cdc have shown no serologic evidence of past infection in more than specimens from us residents collected long before the sars epidemic. • as a new human pathogen, there is, understandably, little if any natural immunity. virtually all persons who became infected by sars-cov became symptomatic, and studies of exposed health care workers show that less than % to % of those infected experience mild or subclinical infection. • sars spreads almost exclusively person-to-person by respiratory droplets, rarely by the airborne route ; the roles of contact or fecal-oral transmission are less clear but probably occur. , the puzzling large outbreak in amoy gardens in hong kong was recently traced to virus-laden aerosols generated from sewage, using sophisticated airflow-dynamic studies and computational fluid-zone modeling. however, whereas the outbreak in the amoy gardens complex represents distant airborne spread, distant spread is probably rare, as evidenced by the very low risk of infection in patients with no plausible face-to-face exposure to a patient with symptomatic sars, by the effectiveness of simple isolation measures in hospitals that did not have sophisticated negative-pressure air-controlled rooms with separate roofline exhaust, and by the relatively few secondary cases on commercial airliners. , [ ] [ ] [ ] in an investigation of flights in which an airliner transported or more symptomatic infected passengers, laboratory-confirmed cases of secondary sars were detected on flight, with the greatest risk to other passengers close to the index case (seated within rows, relative risk . ); on flight carrying symptomatic infected persons, possible transmission occurred to only other passenger, and no secondary illness was documented on another flight that carried person with early sars. • simple control measures, most importantly the use of a high-quality filtration mask, ideally an n- -type mask but even a surgical mask, combined with full-barrier precautions in a single room were highly effective in preventing spread to other patients and health care workers where it was most carefully studied, in hong kong, singapore, and canada. , , , • patients with early sars do not pose a risk to others until they become symptomatic and start to cough, but there is considerable variability in contagiousness, probably based on the quantity of virus in the respiratory secretions and the degree of coughing. very close proximity to an infected patient who is coughing heavily poses the greatest risk. it appears that most spread of the virus can be linked to "super spreaders," and most infected persons are probably not very contagious. , , [ ] [ ] [ ] [ ] understandably, the risk of acquisition of sars-cov is far higher in the hospital than in the community. , • the mean incubation period of sars is approximately days (range, - days) and is considerably longer than that for most other human respiratory viral infections, such as the common cold or influenza a, which permits case-contact investigations and quarantine of exposed contacts before those destined to become infected and contagious can spread the disease to others. • because so few persons develop clinical sars more than days after exposure, there is no need to extend quarantine of exposed persons beyond days. • fever is so ubiquitous in sars that monitoring the body temperature of quarantined contacts and health care workers caring for patients with sars is a sensitive and specific method for detection of early infection, especially for health care workers before they become symptomatic and contagious. all health care workers caring for patients with suspected or proven sars should be monitored to times daily; fever constitutes grounds for quarantine and diagnostic studies. , for personal use. mass reproduce only with permission from mayo clinic proceedings. • to control sars, early diagnosis is essential. clinical predictors for sars, based on study of large cohorts of patients in hong kong, suggest that fever, myalgia, malaise, an abnormal chest radiograph, lymphopenia, thrombocytopenia, and, most importantly, previous contact with a patient with sars are each associated with a greatly increased likelihood of sars. in contrast, in a newly symptomatic patient older than years or younger than years without a plausible face-to-face exposure who has a cough productive of sputum, abdominal pain, sore throat, rhinorrhea, or leukocytosis, sars is unlikely. • modern-day virology has shifted rapidly during the past decade, from tissue cultures and serologic techniques to detection of the viral genome in clinical specimens by nucleic acid amplification techniques, such as polymerase chain reaction (pcr) or, for rna viruses, reverse transcriptase-pcr (rt-pcr). highly sensitive and specific rt-pcr assays were developed in most of the countries afflicted by sars, most notably in hong kong, singapore, and canada, and were invaluable in early confirmation of sars-cov infection. the sensitivity of second-generation assays has been as high as % in the first days of illness. [ ] [ ] [ ] [ ] whereas a pcr assay developed at the cdc was given immediate investigational device exemption approval by the food and drug administration, no commercial pcr assay has yet been licensed for clinical use in the united states. if sars returns and spreads in the united states, it will be essential that reliable real-time pcr assays are available in us hospitals. public health laboratories are not clinical service laboratories and are unlikely to be able to meet the need if sars reappears on a major scale. private companies should be given access to clinical strains of sars-cov and, if available, clinical specimens from infected patients in order to test and validate commercial assays that hopefully will be as accurate as, perhaps more accurate than, the current cdc assay. • ribavirin was used empirically in many patients with sars in southeast asia with the impression that it was effective therapeutically; however, in vitro studies have shown that sars-cov is not susceptible to ribavirin at concentrations achievable clinically. hence, it is unlikely that the drug is active therapeutically. uncontrolled trials suggest that interferon alfa may be of benefit. , there are a number of compounds and antiviral drugs with in vitro activity against sars-cov, including interferon alfa, interferon beta, and glycyrrhizin (licorice-root extract). theoretical rna virus targets, such as protease inhibitors and fusion inhibitors, also need to be assessed for efficacy. if sars returns on a major scale, it will be essential that the efficacy of antiviral drugs, such as commercial interferons, is tested in randomized, double-blind trials. • whereas uncontrolled studies of treated cohorts in asia have suggested that using moderate doses of corticosteroids, to mg/kg of a prednisone-equivalent daily, at the first evidence of severe sars, specifically hypoxemia, may improve survival, , , corticosteroid therapy for sars has had serious adverse effects, and a single randomized trial of preemptive pulse corticosteroid therapy did not show benefit. if sars returns, it will also be essential that efforts are made to determine the efficacy of corticosteroids in a large prospective, randomized, doubleblind trial. • advancing age (> years) and coexisting illnessespecially diabetes or heart failure-greatly increase the likelihood of severe sars (requiring mechanical ventilatory support) and the risk of death. , , inexplicably, sars is usually very mild in children, who do not appear to be very contagious. also, maternal-fetal transmission does not appear to occur. • while coronaviruses are more resistant than most other respiratory viruses, sars-cov appears to be susceptible to the commercial microbicides used for surface decontamination in hospitals. • most importantly, outbreaks in hong kong, singapore, vietnam, canada, and elsewhere in the world were successfully controlled, but only by an intensive, coordinated effort in which the national public health authorities worked very closely with the regional public health agencies and, especially, hospital infection control officers and clinicians caring for patients with sars. [ ] [ ] [ ] [ ] the measures needed for control of sars are clear , , : ( ) earliest detection of cases, having at-risk individuals isolated and queried about their face-to-face contacts during the to days before the onset of illness; ( ) expeditious contact tracing, with uncompromising home quarantine for all contacts of suspect and proven cases; and ( ) stringent isolation of symptomatic suspect and proven cases, focusing most heavily on techniques to prevent droplet and airborne spread (eg, single negative-pressure rooms, ideally with separate roofline exhaust or filtration of outlet air; fit-tested high-filtration mask respirators and a face shield or goggles or a powered air-purifying system for all health care workers and others entering the room of the case, as well as the use of nonsterile gloves and gowns to prevent contact transmission). , the value of border screening and temperature monitoring of travelers is questionable. the resources needed to control an outbreak in a city or a country are huge. in north america, the toronto outbreak consisted of documented cases in hospitals. to control sars in toronto required home quarantine of more than , contacts and an informational hotline that handled more than , calls; the economic cost of the epidemic to the city and the city and provincial govern- ments was estimated at $ . billion (canadian). the longterm psychological impact of sars on patients, families, and health care workers was also very substantial. [ ] [ ] [ ] • efforts are now under way to test candidate sars vaccines. the national tragedy of september , , was followed by the most serious instance of bioterrorism involving the us civilian population in history, the spread of anthrax through the us mail. these events coincided with growing awareness that weaponized smallpox virus almost certainly yet exists in the world, with strong suspicion that the former soviet union, as well as countries that have sponsored international terrorism, such as iran and north korea, , retained smallpox virus as a potential weapon. the unthinkable has become plausible: weaponized smallpox virus in the hands of international terrorist groups. as a consequence, the federal government has undertaken major steps to greatly improve emergency preparedness at all levels, especially the capacity to respond to the use of biologic agents such as smallpox or anthrax as weapons against the civilian population as well as our military (table ) . , billions of dollars have been appropriated to improve the capacity of public health and clinical laboratories to reliably detect infectious agents that might represent biologic weapons; to improve the likelihood that emergency department physicians and all primary care providers could recognize anthrax, smallpox, and other infectious diseases that might denote bioterrorism; to establish and coordinate surveillance programs at the regional, state, and federal levels; and to train more than a million public protection personnel and greatly improve preparedness of the us hospitals. at my center, we have spent hundreds of person-hours identifying and retrofitting a bed patient-care unit for the potential accommodation of patients with smallpox or other highly contagious infections such as sars or pandemic influenza caused by a new strain. this local effort has focused on air control and negative-pressure isolation rooms, which have the capacity for supporting mechanical ventilation, and developing comprehensive guidelines for health care workers who would staff the unit. for the first time in our generation, there has been a major injection of federal dollars into the public health sector at the state, regional, and municipal levels. the challenge will be to provide sustained support, rather than a limited bolus of supplemental funding. hopefully all this effort will never be needed to control smallpox-or an even more terrifying engineered pathogen , -that might be used as a biologic weapon. if it is not, the effort will not have been wasted because it is likely that all the planning and resource allocation will prove invaluable for controlling the spread of natural emerging pathogens, such as sars-cov or a new strain of influenza virus, which are probably far more likely to pose a serious threat to human and animal health in the united states and worldwide. the greatest and most immediate threat is the longoverdue reappearance of pandemic influenza a. the leading and most dreaded candidate for the new pandemic subtype is avian influenza a (h n ), recently reviewed in this journal, which was first recognized in a large poultry outbreak in the live-animal markets of hong kong in , where the virus had acquired the capacity to spread from infected birds to humans and killed of infected persons. to control the outbreak, authorities killed nearly million chickens to eliminate the reservoir of infection. since that time there have been contained outbreaks of different subtypes of avian influenza-h n , h n , and h n -that have caused disease in poultry, with secondary infections reported among pigs and humans, but infrequent and mild human disease, such as conjunctivitis or mild influenza-like illness. there was only human death among cases in a large h n outbreak in the netherlands in . in january , a highly pathogenic strain of avian influenza a (h n ) was identified in south korea and spread rapidly over the succeeding months to other asian countries, cambodia, china and hong kong, indonesia, japan, laos, thailand, and vietnam. [ ] [ ] [ ] to date, there have been confirmed cases in humans, nearly all in children or young adults; ( %) have proved fatal. more than million edible birds have been slaughtered by governmental authorities. all species of domestic birds appear to be susceptible to the h n strain, which is probably transmissible to all species of wild birds, some of which migrate transcontinentally. the epidemic a (h n ) strain appears to be gaining virulence and was recently shown to have acquired the capacity of infecting mammalian species, domestic cats and wild felines within zoos and pigs. most alarmingly, there is growing evidence that person-to-person spread can occur, albeit yet rarely. the epidemic strain further shows high-level resistance to amantadine and rimantadine but is thus far susceptible to neuraminidase inhibitors, such as oseltamivir or zanamivir. if the strain acquires recombinant genes that facilitate human infection and person-to-person transmission, pandemic disease could prove more catastrophic than the great h n influenza epidemic of . even more concerning has been the challenge of developing an avian influenza vaccine. current influenza vaccines are unlikely to provide any protection against the new h n avian strain. the standard method for manufacturing influenza vaccines, growing the vaccine strain in chicken embryos, does not work because the avian a (h n ) strain is so virulent that it kills the embryo before there is sufficient virus to harvest. novel genetic techniques, under way in the united kingdom, will be needed to alter the strain's phenotypic features so that it can be grown in sufficient quantities in fertilized eggs and an effective vaccine can be constructed. vaccine manufacturers are understandably reluctant to make the investment to develop and manufacture a new vaccine, particularly in large quantities, when there is uncertainty whether the avian strain will indeed spread and necessitate administration of hundreds of millions of doses. similarly, the sole manufacturer of the only oral neuraminidase inhibitor likely to be effective against avian influenza (oseltamivir) has very limited production capacity, and less than million doses are currently available in us pharmaceutical stocks; the director of the cdc has stated that it would be desirable to have at least million doses available. in summary, cases of cutaneous or inhalation anthrax traced to domestic bioterrorism and the global sars outbreak represent ill winds that have blown considerable good. the greatly expanded us federal effort to improve national preparedness for bioterrorism has strengthened public health at every level, and whereas we are far from being able to consider the united states as fully prepared, we are better prepared than only years ago. the recent us epidemic of monkeypox, traced to importation of infected exotic african rodents and the burgeoning domestic trade in us prairie dogs, could be considered a live tabletop exercise-with 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human spread of bird flu uk 'developing bird flu vaccine experts confront major obstacles in containing virulent bird flu hospital preparedness for severe acute respiratory syndrome in the united states: views from a national survey of infectious diseases consultants the detection of monkeypox in humans in the western hemisphere key: cord- - fmul c authors: vabret, nicolas; britton, graham j.; gruber, conor; hegde, samarth; kim, joel; kuksin, maria; levantovsky, rachel; malle, louise; moreira, alvaro; park, matthew d.; pia, luisanna; risson, emma; saffern, miriam; salomé, bérengère; selvan, myvizhi esai; spindler, matthew p.; tan, jessica; van der heide, verena; gregory, jill k.; alexandropoulos, konstantina; bhardwaj, nina; brown, brian d.; greenbaum, benjamin; gümüş, zeynep h.; homann, dirk; horowitz, amir; kamphorst, alice o.; curotto de lafaille, maria a.; mehandru, saurabh; merad, miriam; samstein, robert m. title: immunology of covid- : current state of the science date: - - journal: immunity doi: . /j.immuni. . . sha: doc_id: cord_uid: fmul c abstract the coronavirus disease (covid- ) pandemic, caused by severe acute respiratory syndrome coronavirus (sars-cov- ) has affected millions of people worldwide, igniting an unprecedented effort from the scientific community to understand the biological underpinning of covid pathophysiology. in this review, we summarize the current state of knowledge of innate and adaptive immune responses elicited by sars-cov- infection and the immunological pathways that likely contribute to disease severity and death. we also discuss the rationale and clinical outcome of current therapeutic strategies as well as prospective clinical trials to prevent or treat sars-cov- infection. the recent emergence and rapid global spread of severe acute respiratory syndrome coronavirus (sars-cov- ) and the resulting coronavirus disease (covid- ) poses an unprecedented health crisis that was declared a pandemic by the world health organization (who) on march , . the origin of sars-cov- was traced to the city of wuhan in hubei province, china, where a cluster of viral pneumonia cases was first detected, many in connection with the huanan seafood wholesale market. china reported this outbreak to the who on december st , and soon after identified the causative pathogen as a betacoronavirus with high sequence homology to bat coronaviruses using angiotensin-converting enzyme (ace ) receptor as the dominant mechanism of cell entry wan et al., b) . following a likely zoonotic spillover, human-to-human transmission events were confirmed with clinical presentations ranging from no symptoms; to mild fever, cough, and dyspnea; to cytokine storm, respiratory failure, and death. sars-cov- is also closely related to sars (retrospectively named sars-cov- ) and mers (middle eastern respiratory syndrome) coronaviruses, causing local outbreaks and zoonotic epidemics in , respectively (de wit et al., . while sars-cov- is not as lethal as sars-cov- or mers-cov (fauci et al., ) , the considerable spread of the current pandemic has brought tremendous pressure and disastrous consequences for public health and medical systems worldwide. the scientific response to the crisis has been extraordinary with a plethora of covid- studies posted in preprint servers, in an attempt to rapidly unravel the pathogenesis of covid- and potential therapeutic strategies. in response, trainees and faculty members of the precision immunology institute at the icahn school of medicine at mount sinai (priism) have initiated an institutional effort to critically review the preprint literature (vabret et al., ) , together with peer-reviewed articles published in traditional journals, and summarize the current state of science on the fast evolving field of covid- immunology. we thematically focus on the innate and adaptive immune responses to sars-cov- and related coronaviruses, clinical studies and prognostic laboratory correlates, current therapeutic strategies, prospective clinical trials, and vaccine approaches. (bouvet et al., ; chen et al., ; ivanov et al., ) , thereby resembling host mrna to promote translation, prevent degradation, and evade rlr sensing. finally, covs also encode an endoribonuclease, nsp , that cleaves ' polyuridines formed during viral replication, which would otherwise be detected by mda (deng et al., ; hackbart et al., ) . coronaviruses have evolved additional strategies to impede activation of prrs. sars-cov- n-protein prevents trim activation of rig-i . likewise, mers-cov ns a, which itself binds dsrna, impedes pkr activation (comar et al., ; rabouw et al., ) and inhibits pact, an activator of rlrs (niemeyer et al., ; siu et al., ) . additionally, mers-cov ns b antagonizes rnasel, another activator of rlrs (thornbrough et al., ) . the role of other prrs remains unclear. for example, sars-cov- papain-like-protease (plp) antagonizes sting, suggesting that self-dna may also represent an important trigger (sun et al., ) . the extent to which sars-cov- homologs overlap in these functions is currently unknown. following activation, rlr and tlrs induce signaling cascades leading to the phosphorylation of transcription factors, such as nf-kb and the interferon-regulatory factor family (irf), ultimately leading to transcription of ifn and proinflammatory cytokines. although no experimental studies have delineated the precise functions of sars-cov- proteins, proteomic studies have demonstrated interactions between viral proteins and prr signaling cascades. sars-cov- orf b indirectly interacts with the signaling adaptor mavs via its association with tom (gordon et al., ) , consistent with prior reports that sars-cov- orf b suppresses mavs signaling (shi et al., ) . furthermore, sars-cov- nsp interacts with signaling intermediate tbk , and nsp is associated with rnf /nrdp , an activator of tbk and irf (gordon et al., ) . similarly, sars-cov- m protein is known to inhibit the tbk signaling complex (siu et al., ) , as does mers-cov orf b (yang et al., ) . other proteins, including sars-cov- plp, n, orf b and orf , block irf phosphorylation and nuclear translocation (devaraj et al., ; . nf-kb is also inhibited by covs proteins. these include sars-cov- plp (frieman et al., ) and mers-cov orf b and orf (canton et al., ; menachery et al., ) . finally, sars-cov- nsp (huang et al., a; kamitani et al., ) and mers-cov nsp (lokugamage et al., ) initiate general inhibition of host transcription and translation, thus limiting antiviral defenses nonspecifically. to prevent signaling downstream of ifn release, cov proteins inhibit several steps of the signal transduction pathway that bridge the receptor subunits (ifnar and ifnar ) to the stat proteins that activate transcription. for sars-cov- , these mechanisms include ifnar degradation by orf a (minakshi et al., ) , decreased stat phosphorylation by nsp (wathelet et al., ) , and antagonism of stat nuclear translocation by orf kopecky-bromberg et al., ) . however, sars-cov- orf shares only % sequence homology with sars-cov- , suggesting this function may not be conserved. in support of this notion, sars-cov- infection fails to limit stat phosphorylation, unlike in sars-cov- infection (lokugamage et al., ) . mucosal immune responses to infectious agents are orchestrated and regulated by myeloid cells with specialized functions, which include conventional dcs (cdcs), monocyte-derived dcs (modcs), plasmacytoid dcs (pdcs), and macrophages (guilliams et al., ) . a growing body of evidence points to dysregulated myeloid responses that potentially drive the covid- hallmark syndromes such as acute respiratory distress syndrome (ards), cytokine release syndrome (crs) and lymphopenia (mehta et al., ) . flow cytometric analyses of pbmcs from symptomatic covid- patients have shown a significant influx of gm-csf-producing, activated cd + t cells and cd + hla-dr lo inflammatory monocytes (im) (giamarellos-bourboulis et al., ; zhou et al., b) . this matches single-cell transcriptomic (scrnaseq) data demonstrating cd + il- β + monocytic expansion wen et al., ) , interferon-mapk driven adaptive immune responses , and il- βassociated inflammasome signatures (ong et al., ) in peripheral blood of covid- patients, although systemic levels of il- β detected are conspicuously low (gnjatic et al., unpublished) . importantly, these immune signatures track with progression of clinical disease. scrnaseq studies performed on pulmonary tissues of patients with severe covid- disease have revealed an expansion of ims and ficolin- + monocyte-derived macrophages, at the expense of tissue-resident reparative alveolar macrophages (am) . the aforementioned study also observed signatures of ifn-signaling and monocyte recruitment that likely contribute to the rapid decline in alveolar patency and promote ards. although most of the clinical focus has been on pulmonary damage and mononuclear phagocyte (mnp) dysfunction therein, it is increasingly clear that covid- likely presents systemic challenges in other organ sites such as the ileum and kidneys. understanding the role of non-pulmonary myeloid cells in tissue-specific pathology associated with covid- will be important. while data on covid- patients continues to rapidly emerge, studies of myeloid cell dysfunction in sars-cov- and mers-cov can provide an important roadmap to understanding covid- pathogenesis ( figure ). sars-cov- infection in mouse models results in an aberrant am phenotype that limits dc trafficking and t cell activation (zhao et al., ) . additionally, ym + fizz + alternative macrophages can increase airway hypersensitivity, thus exacerbating sars-associated fibrosis (page et al., ) . further, as described above, murine sars-cov- studies have demonstrated that delayed ifn-i signaling and inflammatory monocytes-macrophages promote lung cytokine and chemokine levels, vascular leakage, and impaired antigenspecific t cell responses, culminating in lethal disease (channappanavar et al., ) . the role played by prominent ifn-producing pdcs in sars-cov- control or pathogenesis warrants investigation, as they have been shown to be critical in murine coronavirus (mhv) control (cervantes-barragan et al., ) . longitudinal studies in sars-cov- models are awaited, but initial phenotypic studies in humanized hace mice have shown the characteristic alveolar interstitial pneumonia, with infiltration of lymphocytes and monocytes and accumulation of macrophages in the alveolar lumen (bao et al., a) , which recapitulates patient findings . lastly, nonhuman primate (nhp) studies and patient data on sars-cov- have also shown that virus spike-specific igg responses can exacerbate acute lung injury due to repolarization of alveolar macrophages into pro-inflammatory phenotypes and enhanced recruitment of inflammatory monocyte via ccl and il- (clay et al., ; liu et al., ) . however, the extent to which the antibody response contributes to disease pathophysiology remains to be confirmed. the initial mode of viral pathogen-associated signal (pamp) recognition by innate cells has a major impact on downstream myeloid signaling and cytokine secretion (de marcken et al., ) . while macrophages are somewhat susceptible to mers-cov and sars-cov- infection (perlman and dandekar, ; zhou et al., ) , data do not suggest that they are infected by sars-cov- , although one study reported ace and sars-cov- nucleocapsid protein is expressed in lymph nodes and spleenassociated cd + macrophages of covid- patients producing il- . significantly elevated systemic levels of pro-inflammatory cytokine il- have been reported in several covid- patient cohorts and shown to correlate with disease severity (mehta et al., ) . increased il- can also be associated with higher levels of il- , il- , ifn-ɣ and gm-csf as seen in secondary hemophagocytic lymphohistiocytosis. in response to viral infections, mnps drive interleukin, and ifn-i and ifn-iii production, resulting in inflammasome activation, induction of pathogenic th and th cell responses, recruitment of effector immune cells and crs pathology (prokunina-olsson et al., ; tanaka et al., ) . independently, in vitro studies have demonstrated sars-cov- infection can induce intracellular stress pathways resulting in nlrp -dependent inflammasome activation and macrophage pyroptosis shi et al., ) . functional studies are required to implicate these myeloid inflammasome pathways in covid- lung pathology and to assess other immunogenic pathways such as ripk / -dependent necroptosis (nailwal and chan, ) . in conclusion, the strength and duration of myeloid interferon-stimulated gene (isg) signaling potentially dictate covid- disease severity, but rigorous studies are warranted to confirm this. lastly, more work is needed to ascertain the mechanistic role played by lung-resident and recruited granulocytes in sars-cov- control and pathogenesis (camp and jonsson, ; flores-torres et al., ) . in contrast to their early protective role, neutrophil netosis and macrophage crosstalk can drive later-stage inflammatory cascades (barnes et al., ) , underscoring the overall pathogenic nature of damagesensing host responses ( figure ) . collectively, the current knowledge of coronaviruses and sars-cov- infection, in particular, points to an inadvertent collusion involving myeloid cells in covid- pathogenesis, despite their critical role in early sensing and antiviral responses. innate lymphoid cells (ilcs) are innate immune effector cells that lack the expression of rearranged antigen receptors (tcr, bcr). the ilc family is divided into two main groups: the cytotoxic natural killer (nk) cells and the non-cytotoxic helper ilcs, which include ilc , ilc and ilc (vivier et al., ) . conventional nk cells include cd bright cd -nk cells and cd dim cd + cells, that are specialized in cytokine production or cytotoxicity, respectively. multiple studies have reported reduced numbers of nk cells in the peripheral blood of covid- patients, which is associated with severity of the disease wang et al., f; yu et al., ; zheng et al., b) . a recent scrnaseq analysis revealed a transcriptomic signature for nk cells that was equally represented in lungs from patients and healthy donors . the majority of lung nk cells are non-resident (gasteiger et al., ; marquardt et al., ) , and cxcr has been shown to mediate nk cell infiltration upon influenza infection (carlin et al., ) . in vitro, cxcr ligands (cxcl - ) are increased in sars-cov- -infected human lung tissue (chu et al., ) , and cxcr ligand-producing monocytes are expanded in the lungs of covid- patients . this suggests that the cxcr pathway might facilitate nk cell recruitment from the peripheral blood to the lungs in covid- patients ( figure ). nk cells express inhibitory and activating receptors that regulate their cytotoxicity. they are therefore able to induce the lysis of virus-infected cells that upregulate virus-derived proteins, as well as stress-inducible ligands, which are then recognized by nk cellactivating receptors, such as nkp (cerwenka and lanier, ; draghi et al., ; duev-cohen et al., ; glasner et al., ) . future studies should investigate the expression of nk receptor ligands on sars-cov- -infected cells, in order to better understand the mechanisms underlying nk cell activation in covid- disease. further, secretion of igg and igg antibodies during sars-cov- infection (amanat et al., ) may induce cd dim cd + nk cell activation through fc receptor recognition of antibodies, either bound to surface antigens expressed on infected cells or to extracellular virions as immune complexes ( figure ). this interaction might trigger both cytokine production by nk cells and lysis of infected cells through antibodymediated cellular cytotoxicity (adcc), as shown in influenza infection (von holle and moody, ) . emerging data highlight the capacity for nk-mediated adcc in response to naturally isolated sars-cov- anti-s igg that cross-reacts with sars-cov- s glycoprotein when transfected into chinese hamster ovary (cho) cells (pinto et al., ) . these findings suggest that triggering nk cell activation may not only contribute to the resolution of infection, but also contribute to the cytokine storm in ards. ex vivo nk cells from peripheral blood of covid- patients have reduced intracellular expression of cd a, ksp , granzyme b and granulysin, suggesting an impaired cytotoxicity, as well as an impaired production of chemokines, ifn-ɣ and tnf-α (wilk et al., ; zheng et al., b) . several pathways may contribute to the dysregulation of nk cells. while influenza virus infects nk cells and induces apoptosis (mao et al., ) , lung nk cells do not express the entry receptor for sars-cov- , ace , and are therefore unlikely to be directly infected by sars-cov- (travaglini et al., ) . the majority of nk cells found in human lung display a mature cd + kir + cd dim phenotype and are able to induce cell cytotoxicity in response to loss of hla class i or through fc receptor signaling, although to a lower extent than their peripheral blood counterpart (marquardt et al., ) . killer-immunoglobulin receptors (kirs) are acquired during nk cell development alongside cd (fcrγiiia) and are essential for nk cell licensing and subsequent capacity for cytolytic function (sivori et al., ) . frequencies of nk cells expressing cd and/or kir are decreased in the blood following sars-cov- and sars-cov- infection, respectively (xia et al., ; wang et al., d) . collectively, the data suggest either an impaired maturation of the nk compartment or migration of the mature, circulating nk cells into the lungs or other peripheral tissues of sars-cov- -infected patients. the immune checkpoint nkg a is increased on nk cells and cd t cells from covid- patients . nkg a inhibits cell cytotoxicity by binding the nonclassical hla-e molecule (braud et al., ; brooks et al., ) , and this interaction is strongly correlated with poor control of hiv- infection (ramsuran et al., ) . genes encoding the inhibitory receptors lag and tim are also upregulated in nk cells from covid- patients (wilk et al., ; hadjadj et al., ) . thus, increased immune checkpoints on nk cells might contribute to viral escape. additionally, covid- patients have higher plasma concentrations of il- , which significantly correlate with lower nk cell numbers . in vitro stimulation by il- and soluble il- receptor has previously revealed impaired cytolytic functions (perforin and granzyme b production) by healthy donor nk cells, which can be restored following addition of tocilizumab (il- r-blockade) (cifaldi et al., ) . tnf-α is also upregulated in the plasma of covid- patients , and ligand-receptor interaction analysis of peripheral blood scrnaseq data suggests that monocyte-secreted tnf-α might bind to its receptors on nk cells . tnf-α is known to contribute to nk cell differentiation (lee et al., ) , which includes downregulation of nkp (ivagnes et al., ) , though no effect of tnf-α or il- on nk cell-mediated adcc has been reported so far. collectively, these data suggest that cross-talk with monocytes might impair nk cell recognition and killing of sars-cov- infected cells, and antibodies targeting il- and tnf-signaling may benefit enhanced nk cell functions in covid- patients ( figure ). no studies, to date, have reported ilc , ilc , or ilc functions in sars-cov- infection. all three subsets are present in healthy lung (de grove et al., ; yudanin et al., ) . ilc s are essential for the improvement of lung function following influenza infection in mice, through amphiregulin-mediated restoration of the airway epithelium and oxygen saturation (monticelli et al., ) . however, ilc s also produce il- , contributing to the recruitment of macrophages to the lung and influenza-induced airway hyperreactivity (chang et al., ) . indeed, ilcs are involved in the polarization of alveolar macrophages, either towards a m -like phenotype (ilc and ilc ) or a m like phenotype (ilc ) (kim et al., ) . given the increased il- concentrations and the dysregulation of the macrophage compartment observed in covid- patients, the role played by ilcs in sars-cov- infection warrants further investigation. t cells play a fundamental role in viral infections: cd t cells provide b cell-help for antibody production and orchestrate the response of other immune cells, whereas cd t cells kill infected cells to reduce the viral burden. however, dysregulated t cell responses can result in immunopathology. to better understand the role of t cell responses in sars-cov- infection, the pursuit of two major questions is imperative: ( ) what is the contribution of t cells to initial virus control and tissue damage in the context of covid- , and ( ) how do memory t cells established thereafter contribute to protective immunity upon re-infection? some tentative answers are beginning to emerge. similar to earlier observations about sars-cov- infection , several current reports emphasize the occurrence of lymphopenia with drastically reduced numbers of both cd and cd t cells in moderate and severe covid- cases ( figure ) nie et al., b; wang et al., d; zeng et al., ; zheng et al., b) . the extent of lymphopenia -most striking for cd t cells in patients admitted to icu -seemingly correlates with covid- -associated disease severity and mortality diao et al., ; liu et al., b liu et al., , c tan et al., a; wang et al., d zeng et al., ; zhou et al., c) . patients with mild symptoms, however, typically present with normal or slightly higher t cell counts thevarajan et al., ) . the cause of peripheral t cell loss in moderate to severe covid- , though a phenomenon also observed in other viral infections, remains elusive, and direct viral infection of t cells, in contrast to mers-cov (chu et al., ) , has not been reported. several mechanisms likely contribute to the reduced number of t cells in the blood, including effects from the inflammatory cytokine milieu. indeed, lymphopenia seems to correlate with serum il- , il- , and tnf-α (diao et al., ; wan et al., a) , while convalescent patients were found to have restored bulk t cell frequencies paired with overall lower pro-inflammatory cytokine levels diao et al., ; liu et al., a liu et al., , b zheng et al., b) . cytokines such as ifn-i and tnf-α may inhibit t cell recirculation in blood by promoting retention in lymphoid organs and attachment to endothelium (kamphuis et al., ; shiow et al., ) . however, in an autopsy study examining the spleens and hilar lymph nodes of six patients who succumbed to covid- , chen et al. observed extensive cell death of lymphocytes and suggested potential roles for il- as well as fas-fasl interactions . in support of this hypothesis, the il- receptor antagonist tocilizumab was found to increase the number of circulating lymphocytes (giamarellos-bourboulis et al., ) . t cell recruitment to sites of infection may also reduce their presence in the peripheral blood compartment. scrnaseq analysis of bronchoalveolar lavage (bal) fluid of covid- patients revealed an increase in cd t cell infiltrate with clonal expansion . likewise, post-mortem examination of a patient who succumbed to ards following sars-cov- infection showed extensive lymphocyte infiltration in the lungs . however, another study that examined post-mortem biopsies from four covid- patients only found neutrophilic infiltration (tian et al., a) . further studies are therefore needed to better determine the cause and impact of the commonly observed lymphopenia in covid- patients. available information about sars-cov- -specific t cell immunity may serve as an orientation for further understanding of sars-cov- infection. immunogenic t cell epitopes are distributed across several sars-cov- proteins (s, n, m as well as orf ), although cd t cell responses were more restricted to the s protein (li et al., ) . in sars-cov- survivors, the magnitude and frequency of specific cd memory t cells exceeded that of cd memory t cells and virus-specific t cells persisted for at least - years suggesting that t cells may confer long-term immunity (ng et al., ; tang et al., ) . limited data from viremic sars patients further indicated that virusspecific cd t cell populations might be associated with a more severe disease course, since lethal outcomes correlated with elevated th cell (il- , il- , il- ) serum cytokines (li et al., ) . however, the quality of cd t cell responses needs to be further characterized to understand associations with disease severity. few studies have thus far characterized specific t cell immunity in sars-cov- infection. in patients recovering from mild covid- , robust t cell responses specific for viral n, m and s proteins were detected by ifn-γ elispot, weakly correlated with neutralizing antibody concentrations (similar to convalescent sars-cov- patients (li et al., ) , and subsequently contracted with only n-specific t cells detectable in about one third of the cases post recovery . in a second study, pbmcs from covid- patients with moderate to severe ards were analyzed by flow cytometry, approximately weeks after icu admission (weiskopf et al., ) . both virus-specific cd and cd t cells were detected in all patients at average frequencies of . % and . %, respectively, and very limited phenotyping according to cd ra and ccr expression status characterized these cells predominantly as either cd tcm (central memory) or cd tem (effector memory) and temra (effector memory ra) cells. this study is notable for the use of large complementary peptide pools comprising , sars-cov- epitopes (overlapping -mers for s protein as well as computationally predicted hla-i-and -ii-restricted epitopes for all other viral proteins) as antigenspecific stimuli that revealed a preferential specificity of both cd and cd t cells for s protein epitopes, with the former population modestly increasing over ~ - days after initial onset of symptoms. a caveat, however, pertains to the identification of specific t cells by induced cd and cd co-expression, since upregulation of cd by cd t cells, in contrast to cd , may preferentially capture regulatory t cells (treg) (bacher et al., ) . further analyses of s protein-specific t cells by elisa demonstrated robust induction of ifn-γ, tnf-α and il- concomitant with lower levels of il- , il- , il- , il- and il- . a third report focused on s-specific cd t cell responses in patients with mild, severe or critical covid- using overlapping peptide pools and induced cd and cd co-expression as a readout for antiviral cd t cells. such cells were present in % of cases and presented with enhanced cd , hla-dr and ki- expression indicative of recent in vivo activation (braun et al., ) . of note, the authors also detected low frequencies of s-reactive cd t cells in % of sars-cov- seronegative healthy control donors. however, these cd t cells lacked phenotypic markers of activation and were specific for c-terminal s protein epitopes that are highly similar to endemic human coronaviruses, suggesting that crossreactive cd memory t cells in some populations (e.g., children and younger patients that experience a higher incidence of hcov infections) may be recruited into an amplified primary sars-cov- -specific response (braun et al., ) . similarly, endemic cov-specific cd t cells were previously shown to recognize sars-cov determinants (gioia et al., ) . how previous infections with endemic coronavirus may affect immune responses to sars-cov- will need to be further investigated. finally, in general accordance with the above findings on the induction of sars-cov- specific t cells, using tcrseq, huang et al. and liao et al. reported greater tcr clonality of peripheral blood as well as bal t cells in patients with mild vs. severe covid- . moving forward, a comprehensive identification of immunogenic sars-cov- epitopes recognized by t cells (campbell et al., ) , as well as further studies on convalescent patients who recovered from mild and severe disease, will be particularly important. while the induction of robust t cell immunity is likely essential for efficient virus control, dysregulated t cell responses may cause immunopathology and contribute to disease severity in covid- patients (figure ). this is suggested in a study by zhou et al. which reported a significantly increased pbmc frequency of polyclonal gm-csf + cd t cells capable of prodigious ex vivo il- and ifn-γ production only in critically ill covid- patients . of note, gm-csf + cd t cells have been previously implicated in inflammatory autoimmune diseases such as multiple sclerosis or juvenile rheumatoid arthritis, and high levels of circulating gm-csf + cd t cells were found to be associated with poor outcomes in sepsis . additionally, two studies observed reduced frequencies of treg cells in severe covid- cases qin et al., ) . since treg cells have been shown to help resolve ards inflammation in mouse models (walter et al., ) , a loss of tregs might facilitate the development of covid- lung immunopathology. similarly, a reduction of γδ-t cells, a subset of t cells with apparent protective antiviral function in influenza pneumonia (dong et al., ; zheng et al., ) , has been reported in severely sick covid- patients lei et al., b) . currently, little is known about specific phenotypical and/or functional t cell changes associated with covid- . in the majority of preprints and peer-reviewed studies, there are reports of increased presence of activated t cells ( figure ) characterized by expression of hla-dr, cd , cd , cd , cd and ki- (braun et al., ; dong et al., ; guo et al., ; liao et al., ; thevarajan et al., ; yang et al., a; zheng et al., a) . generally, independent of covid- disease severity, cd t cells seem to be more activated than cd t cells thevarajan et al., ; yang et al., a) , a finding that echoes stronger cd , than cd , t cell responses during sars-cov- (li et al., ) . furthermore, in a case study of covid- patients, diao et al. showed that levels of pd- increased from prodromal to symptomatic stages of the disease (diao et al., ) . pd- expression is commonly associated with t cell exhaustion, but it is important to emphasize that pd- is primarily induced by tcr signaling; it is thus also expressed by activated effector t cells (ahn et al., ) . in addition, several studies reported higher expression of various co-stimulatory and inhibitory molecules such as ox- and cd , ctla- and tigit (zheng et al., a) , and nkg a . reduced numbers of cd + cd t cells as well as larger frequencies of pd- + /tim + cd t cells in icu patients were also reported . expression of most of these markers was found to be higher in cd than in cd t cells, and levels tended to increase in severe vs. non-severe cases, which may be due to differences in viral load. cellular functionality was shown to be impaired in cd and cd t cells of critically ill patients, with reduced frequencies of polyfunctional t cells (producing more than one cytokine) as well as generally lower ifn-γ and tnf-α production following re-stimulation with pma and ionomycin zheng et al., a zheng et al., , b . similarly, zheng et al. reported that cd t cells in severe covid- appear less cytotoxic and effector-like with reduced cd a degranulation and granzyme b (gzmb) production . in contrast, a different study found that both gzmb and perforin were increased in cd t cells of severely sick patients (zheng et al., a) . in accordance with the latter observation, when compared to a moderate disease group, convalescent patients with resolved severe sars-cov- infection had significantly higher frequencies of polyfunctional t cells, with cd t cells producing more ifn-γ, tnf-α and il- , and cd t cells producing more ifn-γ, tnf-α and cd a, respectively (li et al., ) . however, given the vigorous dynamics of acute t cell responses and potential differences in sample timing throughout disease course, these observations are not necessarily mutually exclusive. accordingly, rnaseq data by liao et al. showed that cd t cells in the bal fluid of severe covid- patients express cytotoxic genes such as gzma, gzmb, and gzmk at higher levels, while klrc and xcl are enriched in mild cases . in summary, t cells in severe covid- seem to be more activated and may exhibit a trend towards exhaustion, based on continuous expression of inhibitory markers such as pd- and tim- as well as overall reduced polyfunctionality and cytotoxicity. conversely, recovering patients were shown to have an increase in follicular helper cd t cells (t fh ) as well as decreasing levels of inhibitory markers along with enhanced levels of effector molecules such as gzm a, gzmb and perforin (thevarajan et al., yang et al., a; zheng et al., b) . collectively, these studies provide a first glimpse into t cell dynamics in acute sars-cov- infection, but any conclusions have to be tempered at this stage on account of significant limitations in many of the current investigations. the humoral immune response is critical for the clearance of cytopathic viruses and is a major part of the memory response that prevents reinfection. sars-cov- elicits a robust b cell response, as evidenced by the rapid and near-universal detection of virusspecific igm, igg and iga, and neutralizing igg antibodies (nabs) in the days following infection. the kinetics of the antibody response to sars-cov- are now reasonably well described . similar to sars-cov- infection (hsueh et al., ) , seroconversion occurs in most covid- patients between and days after the onset of symptoms, and antibody titers persist in the weeks following virus clearance (figure ) , (haveri et al., ; lou et al., ; okba et al., ; wölfel et al., ; wu et al., b; zhao et al., a) . antibodies binding the sars-cov- internal n protein and the external s glycoprotein are commonly detected (amanat et al., ; ju et al., ; . the receptor binding domain (rbd) of the s protein is highly immunogenic and antibodies binding this domain can be potently neutralizing, blocking virus interactions with the host entry receptor, ace (ju et al., ; wu et al., b) . anti-rbd nabs are detected in most tested patients (ju et al., ; . although cross-reactivity to sars-cov- s and n proteins and to mers-cov s protein was detected in plasma from covid- patients, no cross-reactivity was found to the rbd from sars-cov- or mers-cov. in addition, plasma from covid- patients did not neutralize sars-cov- or mers-cov (ju et al., ) . rbd-specific cd + igg + memory b cells were single-cell sorted from a cohort of covid- donors between days - after the onset of symptoms (ju et al., ) . from their antibody gene sequences, sars-cov- specific monoclonal antibodies were produced. the monoclonal antibodies had a diverse repertoire, relatively low or no somatic mutations, and variable binding reactivity, with dissociation constants reaching - to - , similar to antibodies isolated during acute infections. two potent neutralizing sars-cov- rbd-specific monoclonal antibodies were characterized that did not cross-react with the rbd of sars-cov- or mers-cov (ju et al., ) . together, these results demonstrate that antibody mediated neutralization is virusspecific and likely driven by binding of epitopes within the rbd. the b cell response to a virus serves not only to protect from the initial challenge, but also to offer extended immunity against reinfection. following resolution of an infection, plasma cells formed during the acute and convalescent phases continue to secrete antibodies, giving rise to serological memory. memory b cells that are also formed during the primary infection constitute the second arm of b cell memory. memory b cells can quickly respond to a reinfection by generating new high affinity plasma cells. longterm protection is achieved through the induction of long-lived plasma cells and memory b cells. there is great interest in understanding the life-span of b cell memory responses to sars-cov- . protection from reinfection has direct medical and social consequences as the world works to develop vaccination strategies and resume normal activities. in covid- patients, evidence of near universal seroconversion and the lack of substantial descriptions of reinfection point to a robust antibody response, which, along with the t cell memory response, would offer protection to reinfection. indeed, a case study of a single patient described induction of cd hi cd hi antibody secreting cells (ascs), concomitant with an increase in circulating follicular t helper cells (tfh) cells (thevarajan et al., ) , and a scrnaseq study of pbmc from critically ill and recently recovered individuals revealed a plasma cell population . in addition, igg memory cells specific to the rbd have been identified in the blood of covid- patients (ju et al., ) . consistent with the development of immunity after covid- infection, a recent study of sars-cov- infection in rhesus macaques found that two macaques that had resolved the primary infection were resistant to reinfection days later (bao et al., b) . due to the timing of this outbreak, it is not yet possible to know the nature and extent of long-term memory responses, but lessons may again be learned from other human coronaviruses. in the case of the human coronavirus e, specific igg and nabs are rapidly induced but wane in some individuals around a year after infection with some residual protection to reinfection (callow et al., ; reed, ) . the life span of the humoral response following sars-cov- infection is also relatively short, with the initial specific igg and nab response to sars-cov- diminishing - years after infection and nearly undetectable in up to % of individuals (cao et al., ; liu et al., ) . a long-term study following sars-cov- infected healthcare workers over a years period also found that virus-specific igg declined after several years, but the authors observed detectable virus-specific igg years after infection . in the case of mers-cov, antibodies were detected in of volunteers tested years after infection (payne et al., ) . igg specific to sars-cov- trimeric spike protein was detectable in serum up to days after symptom onset, but igg titers began decreasing by weeks post-symptom onset (adams et al., ) . long term protection from reinfection may also be mediated by reactive memory b cells. a study that analyzed sars-cov- s protein-specific igg memory cells at , , and months post-infection found that s-specific igg memory b cells decreased progressively about % from to months after infection (traggiai et al., ) . a further retrospective study of individuals found no evidence of circulating sars-cov- -specific igg + memory b cells years after infection (tang et al., ) . this is in contrast to the memory t cell response, which was robustly detected based on induced ifn-γ production (tang et al., ) . studies of common coronaviruses, sars-cov- and mers-cov indicate that virus specific antibody responses wane over time, and, in the case of common coronaviruses, result in only partial protection from reinfection. these data suggest that immunity to sars-cov- may diminish following a primary infection and further studies will be required to determine the degree of long-term protection (figure ) . several studies have demonstrated that high virus-specific antibody titers to sars-cov- are correlated with greater neutralization of virus in vitro and are inversely correlated with viral load in patients ( figure ) wölfel et al., ; zhao et al., a) . despite these indications of a successful neutralizing response in the majority of individuals, higher titers are also associated with more severe clinical cases okba et al., ; zhao et al., a; zhou et al., a) , suggesting that a robust antibody response alone is insufficient to avoid severe disease ( figure ). this was also observed in the previous sars-cov- epidemic, where neutralizing titers were found to be significantly higher in deceased patients compared to patients who had recovered . this has led to concerns that antibody responses to these viruses may contribute to pulmonary pathology, via antibody-dependent enhancement (ade) (figure ). this phenomenon is observed, when non-neutralizing virus-specific igg facilitate entry of virus particles into fc-receptor (fcr) expressing cells, particularly macrophages and monocytes, leading to inflammatory activation of these cells (taylor et al., ) . a study in sars-cov- -infected rhesus macaques found that anti-s-igg contributed to severe acute lung injury (ali) and massive accumulation of monocytes/macrophages in the lung . furthermore, serum containing anti-s ig from sars-cov- patients enhanced the infection of sars-cov- in human monocyte-derived macrophages in vitro (yip et al., ) . ade was also reported with a monoclonal antibody isolated from a patient with mers-cov . somewhat reassuringly, there was no evidence of ade mediated by sera from rats vaccinated with sars-cov- rbd in vitro (quinlan et al., ) , nor in macaque immunized with an inactivated sars-cov- vaccine candidate (gao et al. ) . as of now, there is no evidence that naturally developed antibodies towards sars-cov- contribute to the pathological features observed in covid- . however, this possibility should be considered when it comes to experimental design and development of therapeutic strategies. importantly, in all of the descriptions of ade as it relates to coronavirus, the fcr was necessary to trigger the antibody-mediated pathology. high-dose intravenous immunoglobulin (ivig), which may blunt ade, has been trialed in covid- patients shao et al., ) , but further studies are needed to determine the extent to which ivig is safe or beneficial in sars-cov- infection. vaccine trials will need to consider the possibility of antibody-driven pathology upon antigen re-challenge; strategies using f(ab) fragments or engineered fc monoclonal antibodies may prove particularly beneficial in this setting (amanat and krammer, ) . with the rapidly growing number of cases in the first few months, numerous reports on predictors of covid- severity with small cohorts were released. these offered clinicians and immunologists the first understanding of the clinical course and pathological processes that are associated with the novel sars-cov- infection. this section highlights key findings from those studies, with a major focus on the immune factors associated with disease risk or severity. there are currently limited known risk factors for susceptibility to covid- , although this has been evaluated in several studies. zhao et al. compared the abo blood group distribution in a cohort of covid- patients to that of healthy controls from the corresponding regions . they found blood group a to be associated with a higher risk for acquiring covid- , when compared to non-a blood groups; blood group o had the lowest risk for the infection. another study demonstrated an identical association (zietz and tatonetti, ) , and similar results have been previously described for other viruses (lindesmith et al., ) , including for sars-cov- (cheng et al., a) . several large collaborative efforts are currently underway to generate, share and analyze genetic data to understand the links between human genetic variation and covid- susceptibility and severity, the most prominent of which is the covid- host genetics initiative (covid hg.org). these studies are supported by previous observations on sars-cov- that followed the outbreak, which have identified significant associations between genetic variants and immune phenotypes (chan et al., ; wang et al., ; zhao et al., ) . although identifying such polymorphisms and their associated genes and pathways for sars-cov- will require large cohorts, several studies have already highlighted genetic polymorphisms that may potentially impact susceptibility, which remain to be tested in clinical trials. these studies have focused on genetic variants that may impact the expression or function of genes important in viral entry, namely ace (sars-cov- receptor) and tmprss (spike protein activator) (asselta et al., ; cao et al., c; renieri et al., ; stawiski et al., ) . cao et al. identified variants that are potentially expression quantitative trait loci (eqtl) of ace (i.e. they may potentially alter ace gene expression) and analyzed their frequencies in different populations (cao et al., c) . stawiski et al. listed variants that may be critical in ace binding and thereby its function, and compared the frequencies of these variants within different populations (stawiski et al., ) . while there are several limitations to these studies, the major question is whether the utility of these biomarkers is replicable in large populations with covid- clinical outcomes data and in targeted or large-scale genomic analyses that are currently underway. in addition, these studies will reveal the potential associations between genetic variants and susceptibility in a gene or loci agnostic fashion. several routine blood and serological parameters have been suggested to stratify patients who might be at higher risk for complications to aid in allocation of healthcare resources in the pandemic (table ). serologic markers from routine blood work were reported, by comparing patients with mild/moderate symptoms to those with severe symptoms. this includes different acute phase proteins, such as saa (serum amyloid protein) and c-reactive protein (crp) . interestingly, elevations in crp appear to be unique to covid- patients, when compared to other viral infections. other consistently reported markers in non-survivors are increased procalcitonin (pct) and il- levels , as well as increased serum urea, creatinine, cystatin c, direct bilirubin, and cholinesterase . overall, inflammatory markers are common in severe cases of covid- and appear to correlate with the severity of the symptoms and clinical outcome. moreover, the extensive damage that occurs in specific organs of severe covid- patients is possibly related to differences in the expression of ace ( figure ) . lymphopenia is the most frequently described prognostic marker in covid- (table ) , and it appears to predict morbidity and mortality even at early stages (fei et al., ) . tan et al. proposed a prognostic model based on lymphocyte counts at two time points: patients with less than % lymphocytes at days - from the onset of symptoms and less than % at days - had the worst outcomes in this study (tan et al., a) . wynants et al. compared predictors of disease severity across studies (> patients), highlighting crp, neutrophil-to-lymphocyte ratio (n/l) and lactate dehydrogenase (ldh) as the most significant predictive biomarkers . furthermore, a meta-analysis of covid- studies with a total of patients also attempted to identify early-stage patients with poor prognosis . the most consistent findings across the different studies were elevated levels of crp, ldh and d-dimer, as well as decreased blood platelet and lymphocyte counts zhou et al., d) . systemic and pulmonary thrombi have been reported with activation of the extrinsic coagulation cascade, involving dysfunctional endothelium and monocytic infiltration (poor et al., ; varga et al., ) ; thrombocytopenia and elevated d-dimer levels may be indicative of these coagulopathies in covid- patients with important therapeutic implications (fogarty et al., ; poor et al., ) . immunological biomarkers are particularly important, as immunopathology has been suggested as a primary driver of morbidity and mortality with covid- . several cytokines and other immunologic parameters have been correlated with covid- severity (table ) . most notably, elevated il- levels were detected in hospitalized patients, especially critically ill patients, in several studies, and are associated with icu admission, respiratory failure, and poor prognosis (chen et al., f; huang et al., b; liu et al., f) . increased il- r, il- , il- , and gm-csf have been associated with disease severity as well, but studies are limited and further studies with larger cohorts of patients are needed to indicate predictive power zhou et al., b) . conflicting results regarding il- β and il- have been reported gong et al., ; wen et al., ) . although elevated cytokine concentrations have been widely described in covid- patients, the vast majority (including il- , il- , il- , ctack, ifn-γ) do not seem to have prognostic value, because they do not always differentiate moderate cases from severe cases (yang et al., b) . this stratification was possible with ip- , mcp- , and il- ra. while there are reports that levels of il- at first assessment might predict respiratory failure (herold et al., ) , other publications with longitudinal analyses demonstrated that il- increases fairly late during the disease's course, consequently compromising its prognostic value at earlier stages . liu et al. developed a web-based tool using k-means clustering to predict prognosis in terms of death or hospital discharge of covid- patients using age, comorbidities (binary), and baseline log helper t cell count (th), log suppressor t cell count (ts), and log th/ts ratio . total t cell, helper t cell, and suppressor t cell counts were significantly lower, and the th/ts ratio was significantly higher in patients who died from infection, as compared to patients who were discharged. importantly, most serological and immunological changes observed in severe cases are associated with disease severity, but can not necessarily serve as predictive factors, as they may not have utility in early identification of patients at higher risk. discovery of truly predictive biomarkers and potential drivers of hyper-inflammatory processes requires comprehensive profiling of asymptomatic and mild cases and longitudinal studies which are limited to date. confounding variables including age, gender, comorbidities may dramatically affect associations observed. in addition, direct correlation with patient viral load will be important to provide a greater understanding of underlying causes of morbidity and mortality in covid- and the contribution of viral infectivity, hyperinflammation and host tolerance (medzhitov et al., ) . in summary, lymphopenia, increases in proinflammatory markers and cytokines and potential blood hypercoagulability characterize severe covid- cases with features reminiscent of cytokine release syndromes. this correlates with a diverse clinical spectrum ranging from asymptomatic to severe and critical cases. during the incubation period and early phase of the disease, leukocyte and lymphocyte counts are normal or slightly reduced. after sars-cov- binds to ace overexpressing organs, such as the gastrointestinal tracts and kidneys, increases in non-specific inflammation markers are observed. in more severe cases, a marked systemic release of inflammatory mediators and cytokines occurs, with corresponding worsening of lymphopenia and potential atrophy of lymphoid organs, impairing lymphocyte turnover (terpos et al., ). antivirals are a class of small molecules that function as inhibitors of one or more stages of a virus life cycle. because of similarities between different virus replication mechanisms, some antivirals can be repurposed against various viral infections. currently, most of the available antiviral drugs tested against sars-cov- are smallmolecules previously developed against sars-cov- , mers-cov, or other rna and dna viruses. a number of small molecules with known antiviral activity against other human rna viruses are being evaluated for efficacy in treating sars-cov- . the ribonucleoside analog β-d-n -hydroxycytidine (nhc) reduced viral titers and lung injury in mice infected with sars-cov- via introduction of mutations in viral rna (sheahan et al., ) . further, an inhibitor of host dhodh, a rate-limiting enzyme in pyrimidine synthesis, was able to inhibit sars-cov- growth in vitro with greater efficacy than remdesivir or chloroquine xiong et al., ) . merimepodib, a non-competitive inhibitor of the enzyme inosine- ′-monophosphate dehydrogenase (impdh), involved in host guanosine biosynthesis, is able to suppress sars-cov- replication in vitro . finally, n-( -hydroxypropyl)- -trimethylammonium chitosan chloride (htcc), which was previously shown to efficiently reduce infection by the less pathogenic human coronavirus hcov-nl , was also found to inhibit mers-cov and pseudotyped sars-cov- in human airway epithelial cells (milewska et al., ) . much of the antiviral computational and experimental data currently available for sars-cov- focus on targeting the cl or main protease (mpro). two prominent drug candidates targeting the sars-cov- mpro were designed and synthesized, by analyzing the substrate binding pocket of mpro (dai et al., ) . the x-ray crystal structures of the novel inhibitors in complex with sars-cov- mpro were resolved at . Å. both compounds showed good pharmacokinetic activity in vitro, and one exhibited limited toxicity in vivo (dai et al., ) . multiple studies also aimed to repurpose protease inhibitors to reduce sars-cov- titers. nine existing hiv protease inhibitors (nelfinavir, lopinavir, ritonavir, saquinavir, atazanavir, tipranavir, amprenavir, darunavir, and indinavir) were evaluated for their antiviral activity in vero cells infected with sars-cov- (yamamoto et al., ) and nelfinavir was the most potent at inhibiting viral replication. the coronavirus rna-dependent rna polymerase (rdrp) catalyzes the synthesis of viral rna making it essential for viral replication and a prime target for antiviral inhibitors. remdesivir, an adenosine triphosphate analog, inhibits rdrp by binding to rna strands and preventing additional nucleotides from being added, thereby terminating viral rna transcription ( figure a ) (agostini et al., ) . remdesivir has been previously shown to be effective against mers-cov and sars-cov- infections in animal models (sheahan et al., ; de wit et al., ) . similarly, a study investigated the efficacy of remdesivir treatments on rhesus macaques with sars-cov- infections (williamson et al., ) . macaques treated with remdesivir showed a reduction in lung viral loads and pneumonia symptoms, but no reduction in virus shedding. this study does provide evidence that if administered early enough, remdesivir may be effective at treating sars-cov- infections. a large number of clinical trials using experimental antiviral drugs are currently underway. a small proportion of them are aimed at repurposing existing antivirals including: arbidol (umifenovir), a broad-spectrum antiviral that blocks viral fusion; lopinavir/ritonavir (lpv/r), a combination of anti-hiv protease inhibitors; favipiravir, an rdrp inhibitor used to treat severe influenza infections (hayden and shindo, ) ; and remdesivir ( figure a ). chen et al. conducted a multicenter, randomized priority trial on patients with confirmed covid- infection to test favipiravir or arbidol . favipiravir was suggested to significantly improve symptom relief. however, the interpretation of this study is limited by a short clinical recovery window of days, only of patients with confirmed covid- , and the lack of a control group. lpv/r has previously shown efficacy in treating sars-cov- (chu et al., ) , prompting an early sars-cov- clinical trial . patients were enrolled in a trial investigating the efficacy and safety of lpv/r (n= patients), arbidol (n= ), or control (n= ) as treatment for mild-to-moderate covid- . at day of treatment, . %, . % and . % of patients had a positive to negative conversion in the lpv/r, arbidol, and control groups, respectively, with no statistical significance between groups. a randomized controlled trial (rct) with severe covid- patients did not observe a significant benefit of lpv/r either (cao et al., a) . however, a study that looked at the impact of earlier administration of (lpv/r) treatment showed that when treatment of lpv/r was started within days of symptom onset, a shorter duration of virus shedding was observed. thus, timing of lpv/r administration may be critical to its efficacy . in a multicenter clinical study assessing the compassionate use of remdesivir in severe covid- patients, patients across several countries were treated with remdesivir for days (grein et al., ) . % of the patients who received remdesivir showed clinical improvement assessed through improved oxygen-support/extubations. without a proper control group, limited conclusions can be drawn with regards to the efficacy of remdesivir from this study. the measured % clinical improvement may be in line with average clinical improvement across patients treated with standard of care . a small rct in china with severe covid- patients randomized : to remdesivir vs. placebo demonstrated no significant benefit in time to clinical improvement . almost simultaneously, preliminary results from a larger niaid rct with more than patients were announced with remdesevir to be associated with quicker time to recovery: days compared with days (ledford, ) . a non-significant benefit in mortality was also noted and the trial was stopped early to allow access to remdesivir in the placebo arm. complete safety data and full publication are awaited but this study offers encouraging results and have resulted in an fda emergency use authorization for remdesivir in hospitalized covid- patients. chloroquine (cq) and its derivative hydroxychloroquine (hcq) have gained traction as possible therapeutics for covid- . both drugs are used as antimalarial agents and as immunomodulatory therapies for rheumatologic diseases. however, the application of cq and hcq to covid- stems for their past use as antivirals (savarino et al., ) , including for sars-cov- (keyaerts et al., ; vincent et al., ) . cq and hcq interfere with lysosomal activity and have been reported to have immuno-modulatory effects. cq augments antigen processing for mhc class i and ii presentation, directly inhibits endosomal tlr and tlr , and enhances the activity of regulatory t cells (garulli et al., ; lo et al., ; schrezenmeier and dörner, ; thomé et al., a thomé et al., , b . early studies involving in vitro infection of host cells with sars-cov- demonstrated that both cq and hcq significantly impact endosomal maturation, resulting in increased sequestration of virion particles within endolysosomes. however, there has been conflicting evidence whether cq is more potent than hcq in reducing viral load yao et al., a) . notably, one group reported that treatment of infected cells with hcq before and during infection significantly reduced viral load, suggesting that combined prophylactic and therapeutic hcq use yields maximum efficacy (clementi et al., ) . to better understand host immune responses to treatment, one group compared bulk transcriptomic changes in primary pbmcs treated with hcq for hours to pbmcs from confirmed sars-cov- positive patients and controls, followed by a comparison of hcq treated primary macrophages to bal and postmortem lung biopsies from covid- patients (corley et al., ) . across all comparisons, there was minimal overlap between host differential gene expression and genes altered by in vitro hcq treatment. thus, the potential mechanistic action of hcq in the context of sars-cov- remains poorly defined. despite the apparent widespread use of hcq and cq to treat covid- ( figure b ), few controlled clinical trials have been performed so far and thus the potential benefits of these drugs for covid- remains controversial. one of the earliest trials ( - - ) was a single-arm, open label trial of mg daily hcq in covid- patients. they reported that hcq alone, or in combination with the antibiotic azithromycin (az), reduced viral load by day (gautret et al., a) . a follow up trial in patients treated with hcq + az reported that % of patients had a negative pcr result on day of treatment, and . % were discharged within days of treatment. however, it is important to note that both trials had no control arms (gautret et al., b) . rigorous statistical analyses by others that accounted for the patients excluded from the original analysis found limited evidence for hcq monotherapy (hulme et al., ; lover, ) . a double blind rrct assessed hcq monotherapy in the treatment of mild covid- (chictr ) . a total of patients were enrolled; the treatment arm received mg hcq daily over days. by day , patients who received hcq had clinical resolution on average one day earlier than controls; no patients progressed to severe disease compared to patients in the control arm. in a smaller rct treated patients with mild covid- (nct ) with mg hcq for days, there were no significant differences in the number of patients with negative pcr results on day (all but one positive), median duration of hospitalization, time to fever resolution, or progression of disease on chest ct . the largest rct to date enrolled patients with mild covid- across centers in an open label trial of hcq + standard of care (chictr ). there were no significant differences between groups in conversion to negative sars-cov- rt-pcr result on day or rate of symptom resolution; there were significantly more adverse events in the hcq arm, though largely non-serious; they reported some evidence for faster normalization of c-reactive protein in the patients who received hcq plus standard of care, but this finding was not significant (tang et al., b) . a metaanalysis including most of the studies described here found no clinical benefits to patients receiving standard of care plus an hcq regimen (shamshirian et al., ) . two studies have assessed hcq efficacy in severe covid- . in a prospective study of patients who had received mg hcq over days with az on days - , there were several patients with worsening clinical status and one death; / patients had a positive pcr result on day ( molina et al., ) . an ongoing double blind rct of patients with severe covid- (nct ) randomized patients into high dose hcq ( mg x/d for days) or low dose ( mg/day for days) treatment groups (borba et al., ) . recruitment into the high dose arm was halted prematurely due to poor safety outcomes. there was no significant difference in negative pcr results on day or need for mechanical ventilation on day . taken together, the clinical trials performed thus far to evaluate the efficacy of hcq ± az for covid- have not demonstrated clear evidence of clinical benefit in patients with severe disease. a search of clinicaltrials.gov on april , found clinical trials investigating hcq. this number is rapidly growing, indicating the heightened interest in this therapeutic and pressing need for evidence-based recommendations. because of their anti-inflammatory activity, corticosteroids (cs) are an adjuvant therapy for ards and cytokine storm. however, the broad immunosuppression mediated by cs does raise the possibility that treatment could interfere with the development of a proper immune response against the virus. a meta-analysis of , patients with mers-cov, sars-cov- , or sars-cov- infection found that cs treatment was associated with higher mortality . a more recent meta-analysis of only sars-cov- infection assessed , patients and found no mortality difference associated with cs treatment, including in a subset of patients with ards (gangopadhyay et al., ) . other studies have reported associations with delayed viral clearance and increased complications in sars and mers patients (sanders et al., ) . in fact, the interim guidelines updated by the who on march , advise against giving systemic corticosteroids for covid- (world health organization, a) . yet, new data from covid- are conflicting. one group reported no significant difference in time to viral clearance between patients who received methylprednisolone orally (mild disease) or iv (severe) and those who did not . retrospective studies from groups in china report that patients who were transferred to the icu were less likely to have received cs and that patients with ards who received methylprednisolone had reduced mortality risk . in contrast, another retrospective analysis found that patients who received cs were more likely to have either been admitted to the icu or perished, although the cs treated group also had significantly more comorbidities . a smaller observational study of patients found no association between corticosteroid treatment and time to viral clearance, length of hospital stay, or symptom duration (zha et al., ) . a larger study of adjuvant cs in patients with critical covid- found no association with -day mortality; subgroup analysis of patients with ards found no association between treatment with cs and clinical outcomes . they also found that increased dosage was significantly associated with increased mortality risk. a retrospective review of patients, of whom received iv methylprednisolone, found that early, low-dose administration significantly improved spo and chest ct, time to fever resolution, and time on supplemental oxygen therapy (wang et al., h) . others have published perspectives in support of early and short-term, low dose administration based on anecdotal evidence, but not clinical trials. most of the current data on cs use in covid- are from observational studies, and support either modest clinical benefit or no meaningful effects. larger rcts are necessary to understand the risks and benefits of cs for these patients; there are trials evaluating various corticosteroids registered on clinicaltrials.gov as of april , . one of the first defenses of the human body against rna viruses like sars-cov- is the release of type i and iii ifns. it is important to note that type i ifn (ifnα/β) receptors are ubiquitously expressed, so ifnα/β signaling can result in not only antiviral effects, but also in the activation of immune cells that potentially exacerbate pathogenesis. in contrast, type iii ifn (also known as ifnλ) signals mainly in epithelial cells, as well as in a restricted pool of immune cells. because type iii ifns have immunomodulatory functions, subsequent signaling could induce a potent antiviral effect without enhancing pathogenic inflammation (andreakos et al., ; prokunina-olsson et al., ) . recently, there has been a growing interest in the potential therapeutic impact of modulating the ifn response to disable covid- pathogenesis. before the current pandemic, groups have studied the role of ifns in other betacoronavirus infections. one study of patients with sars-cov- infection described unresolved elevated type i ifns and ifn-stimulated genes (isgs) in those with poor outcomes (cameron et al., ) . others report that exogenous type i ifn does not improve outcomes when given with ribavirin in patients with mers-cov infection (arabi et al., ) , suggesting that the role of ifn as a therapeutic or prophylactic option may be strain-or species-specific (sheahan et al., ) . interestingly, a recent study by mount sinai virology groups revealed that type i ifn signaling is impaired in the early response to sars-cov- ; in vitro, sars-cov- may be more susceptible to type i ifn than sars-cov- (blanco-melo et al., ) . based on additional evidence that ifn responses to betacoronaviruses are altered as compared to other respiratory viruses (blanco-melo et al., ; channappanavar et al., ; okabayashi et al., ) , trials of ifn-i/iii administration have been initiated (nct , nct ). hyperinflammatory responses and elevated levels of inflammatory cytokines, including interleukins (il)- , , and , have been shown to correlate with covid- severity diao et al., ; gong et al., ; moore and june, ; wan et al., a; xu et al., b) . the drivers of this cytokine storm remain to be established, but they are likely triggered initially by a combination of viral pamps and host danger signals. the heterogeneous response between patients suggests other factors are involved, possibly including the sars-cov- receptor, ace (hirano and murakami, ) . several studies have begun to report the cellular programs that may contribute to the cytokine storm detected in covid- patients. one group reported that in the context of generalized lymphopenia, certain subsets of cd t cells that express gm-csf and il- are more abundant in severe covid- patients than in covid- patients who do not require intensive care . reports that other major proinflammatory cytokines (tnf-α, ifn-ɣ, il- ) and chemokines (ccl , ccl , ccl ) are elevated underscore a potentially pathogenic t h / program in covid- (diao et al., ; giamarellos-bourboulis et al., ) . histological and single-cell analyses identified monocytes/macrophages as other potent sources of inflammatory cytokines in covid- cytokine storm giamarellos-bourboulis et al., ; law et al., ; moore and june, ; zhou et al., b) . studies of other betacoronavirus infections, including sars-cov- and mers-cov, have also identified similar hyperactivation of monocytes, macrophages, and dendritic cells as a driver of cytokinemediated immunopathology in humans chien et al., ; huang et al., c; konig et al., ; wang et al., ; wong et al., ; xu et al., b; zhou et al., b) . following preliminary reports of il- as a critical cytokine in covid- -associated cytokine release syndrome (crs), monoclonal antibodies that target the il- signaling pathway have been proposed as therapeutic candidates (moore and june, ) ( figure c ). the commercial anti-il- r antibodies tocilizumab (actemra) and sarilumab (kevzara), and the anti-il- antibody siltuximab (sylvant), are now being tested for efficacy in managing covid- crs and pneumonia in ongoing clinical trials (table ). to date, only one group has reported preliminary results from a cohort of covid- patients treated with a single administration of tocilizumab ( mg, iv), along with lopinavir, methylprednisolone, and oxygen therapy (chictr ) . the single observation study found recuperated lymphocyte counts in / patients and resolution of lung opacities in / patients on chest ct; / patients were discharged. all patients experienced an improvement in symptoms, and no subsequent pulmonary infections were reported. a second report described an association between use of tocilizumab and reduced likelihood of icu admission and mechanical ventilation. still, in declining patients with severe covid- pneumonia, this retrospective study did not report significant improvement in mortality on weighted analysis (roumier et al., ) . nevertheless, these studies are encouraging but like other treatment approaches, larger rcts are needed. in addition to the il- signaling pathway, other cytokine-/chemokine-associated elements, including il- r, gm-csf and the chemokine receptor ccr , have been proposed as potential targets for blockade to manage covid- crs ( figure c ). finally, complement activation was shown to be over-activated in lungs of covid- patients. although results from the randomized trial are not yet published, anti-c a monoclonal antibody therapy showed benefits in two critically-ill covid- patients . while vaccines are being developed to educate a person's immune system to make their own nab against sars-cov- , there is interest in using adoptive transfer of nab as a therapeutic approach ( figure d ). this strategy has already proven to be effective against sars-cov- (cao et al., ; ho et al., ; ter meulen et al., ; sui et al., ; zhu et al., ) and mers-cov (forni et al., ; jia et al., ; ying et al., ) . in the case of sars-cov- , these efforts are primarily centered on identifying nab made during natural infections or generating nab through animal vaccination approaches. patients who have recovered from sars-cov- infection are one potential source of nabs (ju et al., ; walls et al., ; wölfel et al., ; ye et al., ; yuan et al., ) . in an effort to obtain these nabs, scientists sorted rbd specific memory b cells and cloned their heavy and light variable region to express recombinant forms of the corresponding antibodies (ju et al., ; ye et al., ) . four of the antibodies produced in these studies ( b , d , p c- f p c- f ) showed high neutralizing potential in vitro, and all inhibited ace /rbd binding. successful antibody-mediated neutralization of sars-cov- seems to be dependent on the inhibition of ace /rbd binding. however, ye et al. showed that nearly all antibodies derived from serum of recovered patients bound to s and rbd, with only actually inhibiting ace /rbd binding . of note, a sars-cov- derived neutralizing antibody ( d ) and a single chain antibody against sars-cov- (n ) have also been shown to neutralize sars-cov- without inhibiting ace /rbd binding. thus, blocking this interaction may not be a prerequisite for an effective sars-cov- nab. the generation of a hybridoma producing a monoclonal nab against sars-cov- provides the potential for a therapeutic ab that can be directly administered to patients to block ongoing infection and potentially even as a prophylactic ( figure d ). sars-cov- and sars-cov- consensus sequences share about % identity (tai et al., ) . thus, a wide range of sars-cov- nabs have been tested for crossreactivity with sars-cov- , as they could help speed up the development of potential covid- treatments. in a recent study, antibodies were isolated from the memory b cells of an individual who recovered from sars-cov- infection. while out of isolated antibodies could bind sars-cov- s protein, one of them (s , see table ) also neutralizes sars-cov- (pinto et al., ) . the combination of s with a weakly neutralizing antibody that could bind another rbd epitope led to enhanced neutralization potency. in addition, cr (table ) was found to bind sars-cov- rbd , but this antibody did not neutralize sars-cov- . computational simulations identified amino-acids that could be modified on cr to enhance its binding affinity with sars-cov- rbd (corrêa giron et al., ) , potentially augmenting its neutralization potential. animal models represent another tool to generate nabs against sars-cov- (table ) . in one study, the authors developed a protocol to synthetize human nanobodies, smaller antibodies that only contain a heavy variable (vh) chain as first described in camelids (figure d ). another antibody isolated from camelids immunized with sars-cov- and mers-cov s proteins then fused to a human fc fragment showed neutralization potential against sars-cov- (vhh- -fc) (wrapp et al., ) . genetically modified mice with humanized antibody genes can also be used to generate therapeutic monoclonal antibodies, as successfully experimented against ebola virus (levine, ) . similar studies are now focused on the use of sars-cov- or derivatives to generate highly effective nab in animal models, which can be directly given to infected patients, and efforts are already underway with estimates of clinical trials of pooled antibody cocktails beginning in early summer by regeneron. finally, another approach to nab development is to fuse ace protein and the fc part of antibodies as they would bind rbd and potentially be cross reactive among other coronaviruses ( figure d ). indeed, an ace -fc as well as an rbd-fc have been shown to neutralize both sars-cov- and sars-cov- in vitro. although recombinant nabs could provide an effective treatment, they will require a significant time investment to develop, test, and bring production to scale before becoming widely available to patients. a faster strategy consists of transferring convalescent plasma (cp) from previously infected individuals that have developed high titer nabs that target sars-cov- ( figure d ). despite the current lack of appropriately controlled trials, cp therapy has been previously used and shown to be beneficial in several infectious diseases such as the influenza pandemic (luke et al., ) , h n influenza (hung et al., ) , and sars-cov- (arabi et al., ) . thanks to the development of serological tests (amanat et al., ; cai et al., ; xiang et al., b; zhang et al., d) , recovered covid- patients can be screened to select plasma with high antibody titers. some studies and case reports on cp therapy for covid- have evaluated the safety and the potential effectiveness of cp therapy in patients with severe disease (ahn et al., ; duan et al., ; pei et al., ; shen et al., ; zhang et al., b) (table ). these studies were neither controlled nor randomized, but they suggest that cp therapy is safe and can have a beneficial effect on the clinical course of disease. further controlled trials are needed to determine the optimal timing and indication for cp therapy. cp therapy has also been proposed for prophylactic use in at-risk individuals, such as those with underlying health conditions or health care workers exposed to covid- patients. the fda has approved the use of cp to treat critically ill patients (tanne, ) . determining when to administer the cp is also of great importance, as a study in sars-cov- patients showed that cp was much more efficient when given to patients before day day of illness (cheng et al., b) , as previously shown in influenza (luke et al., ) . this study also showed that cp therapy was more efficient in pcr positive, seronegative patients. the amount of plasma and number of transfusions needed requires further investigation (table ) . overall, cp therapy seems to be associated with improved outcomes, and appears to be safe, but randomized clinical trials are needed to confirm this. several clinical trials are currently in progress worldwide (belhadi et al., ) the devastating effects of the pandemic spread of sars-cov- in a globally naïve population has resulted in unprecedented efforts to rapidly develop, test, and disseminate a vaccine to protect against covid- or to mitigate the effects of sars-cov- infection. although vaccination has a long and successful history as an effective global health strategy, there are currently no approved vaccines to protect humans against coronaviruses (andré, ) . previous work after the sars-cov- and mers-cov epidemics has provided a foundation on which many current efforts are currently building upon, including the importance of the s protein as a potential vaccine. diverse vaccine platforms and preclinical animal models have been adapted to sars-cov- , facilitating fast-moving and robust progress in creating and testing sars-cov- vaccine candidates. a number of vaccine candidates are already being tested in clinical trials and more are continuing to progress towards clinical testing. since sars-cov- first emerged, the s protein has been favored as the most promising target for vaccine development to protect against coronavirus infection. this particular viral protein has important roles in viral entry and in stimulating the immune response during natural infection and in vaccination studies of both sars-cov- and mers-cov (du et al., ; song et al., ; zhou et al., ) , which has also been confirmed for sars-cov- . the s protein has been found to induce robust and protective humoral and cellular immunity, including the development of nabs and t cell-mediated immunity (du et al., ) . in animal models, correlates of protection against sars-cov- infection appear to be induction of nabs against the s protein, although antibodies to other proteins have been detected such as those against nucleoprotein (n) and orf a (qiu et al., ; sui et al., ) . nabs are also believed to protect against infection by blocking receptor binding and viral entry, which has been shown with pseudovirus-based neutralization assays nie et al., a) . studies of sars-cov- indicate that t cell responses, which were targeted to the s protein after natural infection, included cd + t cell responses against the membrane (m) and n proteins, may also be a correlate of protection and that memory t cell responses can persist even years after infection (li et al., ; ng et al., ) . rbd-specific antiviral t cell responses have also been detected in people who have recovered from covid- , further validating its promise as a vaccine target (braun et al., ; dong et al., ) . although the antibodies targeting the rbd of the s protein have greater potential for providing cross-protective immunity, other fragments of the s protein and additional viral proteins have been investigated as target epitopes, especially for t cells. researchers have taken advantage of the genetic similarity between sars-cov- and sars-cov- and mers-cov and bioinformatics approaches to rapidly identify b and t cell potential epitopes in the s and other proteins, with many studies providing data regarding antigen presentation and antibody binding properties and one study looking into the predicted evolution of epitopes (ahmed et al., ; baruah and bose, ; bhattacharya et al., ; fast et al., ; grifoni et al., ; lon et al., ; zheng and song, ) . while the s protein has been found to be the most immunodominant protein in sars-cov- , the m and n proteins also contain b and t cell epitopes, including some with high conservation with sars-cov- epitopes . for sars-cov- and mers-cov, animal studies and phase i clinical trials of potential vaccines targeting the s protein had encouraging results, with evidence of nab induction and induction of cellular immunity martin et al., ; modjarrad et al., ) . these findings are being translated into sars-cov- vaccine development efforts, hastening the progress drastically. the who provided a report earlier in april that reported sixty-three vaccine candidates in preclinical testing and three in clinical testing (world health organization, b) . a recent search on may , , on clinicaltrials.gov revealed ten registered vaccine candidates (table ). the university of pittsburgh is also looking to move their microneedle array vaccine candidate containing a codon-optimized s subunit protein into clinical trials . sanofi and glaxosmithkline (gsk) have recently reported their intent to collaborate and bring together sanofi's baculovirus expression system, which is used to produce the influenza virus vaccine, flublok, to create an s protein vaccine adjuvanted with gsk's as . the purified inactivated sars-cov- virus vaccine candidate (picovacc) of sinovac biotech ltd. will also be starting a clinical trial in china after finding that their candidate protected rhesus macaques from viral challenge without signs of detectable immunopathology . although some of these vaccine candidates are based on platforms that have been used or tested for other purposes, there remain questions regarding their safety and immunogenicity, including the longevity of any induced responses, that will require continual evaluation. although the development of a vaccine to protect against sars-cov- infection has progressed at an unprecedented rate and produced an impressive volume of candidates for testing, many challenges lie ahead. the prior knowledge gained after sars-cov- was first discovered in and the subsequent emergence of mers-cov in provided a significant jumpstart, but the progress of sars-cov- vaccine development has already far outstripped the point of the blueprint created before covid- became a pandemic. while a variety of platforms are simultaneously being innovated or adapted, they each have strengths and limitations, many of which relate to the delicate balance between safety and immunogenicity. many shortcuts have been taken and will continue to be taken due to the urgency of the ongoing covid- pandemic, but significant concerns need to be addressed. one such concern involves the accumulating data supporting the initial assessment that covid- is disproportionately severe in older adults. in conjunction with the large body of work related to immune-senescence, these findings indicate that vaccine design should take into consideration the impact of aging on vaccine efficacy (nikolich-Žugich, ). furthermore, questions remain regarding the possibility of antibody-dependent enhancement of covid- , with in vitro experiments, animal studies, and two studies of covid- patients supporting this possibility (cao, ; tetro, ; zhao et al., a) . assuming vaccine candidates are identified that can safely induce protective immune responses, additional major hurdles will be the production and dissemination of a vaccine. for some types of vaccines, large-scale production will not be as much of an issue and infrastructure already in place to produce current good manufacturing practice (cgmp)-quality biologics can be repurposed, but this will only be applicable to a subset of the candidates (thanh le et al., ) . in order to address the urgent need and stem the covid- pandemic, regulatory agencies need to continue to support rapid testing and progression of vaccine candidates, companies need to disseminate important findings directly and openly, and researchers need to investigate correlates of protection using in-depth immune monitoring of patients with a broad range of clinical presentations and clinical trial participants. the newly announced accelerating covid- therapeutic interventions and vaccines (activ) is designed to bring together numerous governmental and industry entities to help address this need. the rapid spread of sars-cov- and the unprecedented nature of covid- has demanded an urgency in both basic science and clinical research, and the scientific community has met that call with remarkable productivity. within months, there has been a significant generation of scientific knowledge that has shed some light on the immunology of sars-cov- infections. studies of past coronavirus outbreaks, involving sars-cov- and mers-cov, have provided a foundation for our understanding. the pathology of severe cases of covid- do indeed resemble certain immunopathologies seen in sars-cov- and mers-cov infections, like crs. however, in many other ways, immune responses to sars-cov- are distinct from those seen with other coronavirus infections. the emerging epidemiological observation that significant proportions of individuals are asymptomatic despite infection, not only reflects our current understanding that sars-cov- has a longer incubation period and higher rate of transmission than other coronaviruses, but also speaks to significant differences in the host immune response. therefore, it is imperative that immune responses against sars-cov- and mechanisms of hyperinflammation-driven pathology are further elucidated to better define therapeutic strategies for covid- . here, we reviewed the recent literature and highlighted hypotheses that interrogate mechanisms for viral escape from innate sensing; for hyper-inflammation associated with crs and inflammatory myeloid subpopulations; for lymphopenia marked by t cell and nk cell dysfunction; and for correlates of protection and their duration, among others. still, additional studies are needed to address how these immune differences across patients or between different types of coronavirus infections dictate who succumbs to disease and who remains asymptomatic. existing studies of sars-cov- and mers-cov and ongoing studies of sars-cov- will likely provide a robust framework to fulfill that unmet need. overview of innate immune sensing (left) and interferon signaling (right), annotated with the known mechanisms by which sars-cov- and mers-cov antagonize the pathways (red). [based on data from preliminary covid- studies and earlier studies in related coronaviruses] il- , il- β and ifn-i/iii from infected pulmonary epithelia can induce inflammatory programs in resident (alternate) macrophages while recruiting inflammatory monocytes as well as granulocytes and lymphocytes from circulation. sustained il- , and tnf-ɑ by incoming monocytes can drive several hyperinflammation cascades. inflammatory monocyte-derived macrophages can amplify dysfunctional responses in various ways (listed in top left corner) . the systemic crs-and shlh-like inflammatory response can induce neutrophilic netosis and microthrombosis, aggravating covid- severity. other myeloid cells such as pdcs are purported to have an ifn-dependent role in viral control. monocyte-derived cxcl / / might recruit nk cells from blood. preliminary data suggest that the antiviral function of these nk cells might be regulated through cross-talk with sars-infected cells and inflammatory monocytes. a decrease in peripheral blood t cells associated with disease severity and inflammation is now well documented in covid- . several studies report increased numbers of activated cd and cd t cells which display a trend towards an exhausted phenotype in persistent covid- , based on continuous and upregulated expression of inhibitory markers as well as potential reduced polyfunctionality and cytotoxicity. in severe disease, production of specific inflammatory cytokines by cd t cells has also been reported. this working model needs to be confirmed and expanded on in future studies to assess virus-specific t cell responses both in peripheral blood and in tissues. in addition, larger and more defined patient cohorts with longitudinal data are required to define the relationship between disease severity and t cell phenotype. abbreviations: il, interleukin; ifn, interferon; tnf, tumor necrosis factor; gm-csf, granulocyte-macrophage colony-stimulating factor; gzma/b, granzyme a/granzyme b; prf , perforin. virus-specific igm and igg are detectable in serum between and days after the onset of symptoms. viral rna is inversely correlated with neutralizing antibody titers. higher titers have been observed in critically ill patients, but it is unknown whether antibody responses somehow contribute to pulmonary pathology. the sars-cov- humoral response is relatively short lived, and memory b cells may disappear altogether, suggesting that immunity with sars-cov- may wane - years after primary infection. the gastrointestinal tract, kidneys and testis have the highest ace expressions. in some organs, different cell types have remarkably distinct expressions, e.g. in the lungs, alveolar epithelial cells have higher ace expression levels than bronchial epithelial cells; in the liver, ace is not expressed in hepatocytes, kupffer cells nor endothelial cells, but is detected in cholangiocytes, which can explain liver injury to some extent. furthermore, ace expression is enriched on enterocytes of the small intestine compared to the colon. ace , angiotensin-converting enzyme ; bnp, b-type natriuretic peptide; crp, creactive protein; il, interleukin; n/l, neutrophil-to-lymphocyte ratio; pt, prothrombin time; aptt, activated partial thromboplastin time. a. rdrp inhibitors (remdesivir, favipiravir), protease inhibitors (lopinavir/ritonavir), and anti-fusion inhibitors (arbidol) are currently being investigated in their efficacy in controlling sars-cov- infections. b. cq and hcq increase the ph within lysosomes, impairing viral transit through the endolysosomal pathway. reduced proteolytic function within lysosomes augments antigen processing for presentation on mhc complexes and increases ctla expression on tregs. c. antagonism of il- signaling pathway and of other cytokine-/chemokine-associated targets has been proposed to control covid- crs. these include secreted factors like gm-csf that contribute to the recruitment of inflammatory monocytes and macrophages. d. several potential sources of sars-cov- neutralizing antibodies are currently under investigation, including monoclonal antibodies, polyclonal antibodies, and convalescent plasma from recovered covid- patients. abbreviations: gm-csf, granulocyte-macrophage colony-stimulating factor; cq, chloroquine; hcq, hydroxychloroquine; rdrp, rna-dependent rna polymerase. tai, w., he, l., zhang, x., pu, j., voronin, d., jiang, s., zhou, y., and du, l. 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immunogenicity of a recombinant adenovirus type- vector-based ebola vaccine in healthy adults in sierra leone: a single-centre, randomised, double-blind potent cross-reactive neutralization of sars coronavirus isolates by human monoclonal antibodies sars-cov- receptor ace is an interferon-stimulated gene in human airway epithelial cells and is enriched in specific cell subsets across tissues testing the association between blood type and covid- infection, intubation, and death. medrxiv. lymphocyte count predicted the disease severity and the outcomes of hospitalized patients prognostic value was confirmed in numerous studies decreased continuously in non-surviving patients were reported to be more likely to develop severe illness and to require intensive care unit (icu) admission on admission a risk factor for short-term progression of patients with moderate pneumonia to severe pneumonia confirmed to be of prognostic value in covid- in several studies even in early stages, crp levels were positively correlated with lung lesions and reflected disease severity confirmed in numerous studies predicted the risk of acute myocardial injury ldh (lactate dehydrogenase) higher in severe cases than in mild cases we apologize to all authors whose work we could not cite due to space limitations. trainee (phd and md/phd or postdocs) and faculty contributing authors are listed in alphabetical order. illustrations by jill k gregory and used with permission of ©mount sinai health system. we would like to acknowledge funding sources including fastgrant (mm), nci cancer center support grant supplement (mm, rms) burroughs wellcome fund (rms), nih director's early independence award (rms) and nih r ai (nv, nb, bdg) purpose ro uti ne bl oo dw or k predicted severity independently of other variables . elevated levels and disseminated intravascular coagulation are found in non-survivors . identified patients at risk for acute cardiac injury . other coagulation parameters such as fibrin degradation product levels, longer prothrombin time and activated partial thromboplastin time, were also associated with poor prognosis (tang et al., a) . saa (serum amyloid protein) saa was proposed to be used as an auxiliary index for diagnosis as it was elevated in % of the patients in a small cohort . nt-probnp (n terminal pro b type natriuretic peptide) nt-probnp was an independent risk factor of in-hospital death in patients with severe covid- . high platelet-to-lymphocyte ratio is associated with worse outcome (qu et al., ) . thrombocytopenia is associated with poor outcome and with incidence of myocardial injury in covid- (liu et al., h; shi et al., ) . im mu nol ogi cal cd +, cd + and nk cell counts lower cd +, cd + and nk cells in pbmc correlated with severity of covid- (nie et al., b) . validated by several studies zheng et al., b) . pd- and tim- expression on t cells increasing pd- and tim- expression on t cells could be detected as patients progressed from prodromal to overtly symptomatic stages (diao et al., ) .expression was higher in infected patients versus healthy controls and in icu versus non-icu patients in both cd and cd t cells . phenotypic changes in peripheral blood monocytes the presence of a distinct population of monocytes with high forward scatter (cd b+, cd +, cd +, cd +, cd +, cd +, cd + which secrete il- , il- and tnf-alpha) was identified in patients requiring prolonged hospitalization and icu admission . cd +cd +il- + monocytes are increased in icu patients . ip- , mcp- , and il- ra ip- , mcp- , and il- ra were, among examined cytokines, the only ones that closely associated with disease severity and outcome of covid- in a study by yang et al. (yang et al., b) . associated with disease severity (hospitalization and icu admission) and poor prognosis (chen et al., f; huang et al., b; liu et al., b . increase levels were associated with higher risk of respiratory failure (yao et al., b) . il- positively correlated with disease severity (chen et al., d; gong et al., ) , with severe cases showing the highest il- levels. increased in severe or critical patients as compared to mild patients zhou et al., d ) without a statistically significant difference between severe and critical cases . associated with disease severity in a study that, amongst other cytokines, also associated ferroprotein levels, pct levels, and eosinophil counts with covid- severity . il- β cd +il- β+ monocytes are abundant in early recovery patients as shown in a single-cell rna-seq analysis and thought to be associated with cytokine storm . il- β did not correlate with disease severity in a cross-sectional study with mild, severe and critical patients . il- il- was associated with impaired lung lesions , but some reports point to a potential mediator effect . in modeling immune cell interaction between dc and b cells in late recovery covid- patients, il- was found to be important in b cell production of antibodies, which suggests its importance in recovery . gm-csf (granulocytemacrophage colony-stimulating factor) gm-csf+ifn-γ+ t cells are higher in icu than non-icu patients, cd +cd +gm-csf+ monocytes are higher in covid- patients as compared to healthy controls .il- and ifn-γ il- and ifn-γ levels were shown to be increased in severe cases . anti-sars-cov- antibody levels prolonged sars-cov- igm positivity could be utilized as a predictive factor for poor recovery . higher anti-sars-cov- igg levels and higher n/l were more commonly found in severe cases . between and days after hospital admission.-body temperature normalized within days in of patients -clinical improvement -viral loads became negative within d after the transfusion -neutralizing antibody titers increased severe patients ( - yo)median . dpo.-disappearance of clinical symptoms after d -chest ct improved -elevation of lymphocyte counts in patients with lymphocytopenia.-increase in sao in all patients -resolution of sars-cov- viremia in patients increase in neutralizing antibody titers in patients (ahn et al., ) key: cord- - innqoip authors: huang, y.; lyu, x.; li, d.; wang, y.; wang, l.; zou, w.; wei, y.; wu, x. title: a cohort study of patients explores the clinical risk factors for the severity diagnosis of covid- date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: innqoip background: coronavirus disease (covid- ) has recently become a public emergency and a worldwide pandemic. the clinical symptoms of severe and non-severe patients vary, and the case-fatality rate (cfr) in severe covid- patients is very high. however, the information on the risk factors associated with the severity of covid- and of their prognostic potential is limited. methods: in this retrospective study, the clinical characteristics, laboratory findings, treatment and outcome data were collected and analyzed from covid- patients stratified into non-severe patients and severe patients. in addition, a pooled large-scale meta-analysis of cases was performed. results: we found that the age, gender and comorbidities are the common risk factors associated with the severity of covid- . for the diagnosis markers, we found that the levels of d-dimer, c-reactive protein (crp), lactate dehydrogenase (ldh), procalcitonin (pct) were significantly higher in severe group compared with the non-severe group on admission (d-dimer: . % vs. . %, p< . ; crp, . % vs. . %, p< . ; ldh: . % vs. . %, p< . ; pct: . % vs. . %, p< . ), while the levels of aspartate aminotransferase (asp) and creatinine kinase (ck) were only mildly increased. we also made a large scale meta-analysis of cases combined with related literatures, and further confirmed the relationship between the covid- severity and these risk factors. moreover, we tracked dynamic changes during the process of covid- , and found crp, d-dimer, ldh, pct kept in high levels in severe patient. among all these markers, d-dimer increased remarkably in severe patients and mostly related with the case-fatality rate (cfr). we found adjuvant antithrombotic treatment in some severe patients achieved good therapeutic effect in the cohort. conclusions: the diagnosis markers crp, d-dimer, ldh and pct are associated with severity of covid- . among these markers, d-dimer is sensitive for both severity and cfr of covid- . treatment with heparin or other anticoagulants may be beneficial for covid- patients. in december , a novel coronavirus disease (covid- ) caused by severe acute respiratory syndrome coronavirus (sars-cov- ) infection broke out in wuhan and spread rapidly throughout china. as of april th, , world health organization (who) reported , , covid- cases have been confirmed in most countries or areas, and , people died from covid- ( ). the overall covid- case-fatality rate (cfr) has approached about %. human to human transmission has accounted for most if not all of the covid- infections, including the medical personnel in hospital ( , ) . the clinical manifestations of covid- included fever, cough, diarrhea, dyspnea, fatigue and pneumonia ( ) ( ) ( ) . most patients with covid- were considered as non-severe patients and the symptoms generally mild. however, the symptoms in about % of covid- patients are severe and some progress rapidly to critical conditions, including organ dysfunction, such as acute respiratory distress syndrome (ards), acute cardiac injury, acute kidney injury and even death ( , ) . so far, there is no specific medicine or effective vaccines for covid- , and the diagnosis methods for the severity of covid- patients were limited. thus, our objective of this cohort is to explore the risk factors which related with the severity of covid- and put it into practice. in addition, we used meta-analysis to verify these risk factors. our study would be helpful for setting up different treatment and personal therapy route for covid- patients. the general clinical characteristics of covid- cases in our study are shown in table . patients were categorized into non-severe and severe subgroups on admission respectively. in our cohort, the median age in the non-severe population was . years, interquartile range (iqr) was . to . , in severe population was years (iqr: . to . ). the age differed significantly between the two subgroups, and . % of severe patients were aged above years. men are more likely to develop severe covid- than women ( . % vs. . %). fever ( . %), cough ( . %) and fatigue ( . ) were the most common symptoms, whereas diarrhea ( . %) was rare. notably, fever occurred more frequently in the patients in the severe group than the non-severe group ( . % vs. . %). we also found the comorbidities were risk factors associated withe the severity of covid- in our cohort. more severe patients had common chronic diseases than non-severe patients, such as hypertension ( . % vs. . %) and diabetes ( . % vs. . %). the representative radiologic findings of one patient with non-severe and another three patients with severe covid- were demonstrated in figure . severe cases yielded more prominent radiologic abnormalities on ct than non-severe cases. the laboratory findings on admission were shown in table table is the summary of case studies ( , , , ) examining the association between clinical characters and covid- in the meta-analysis, dividing into two subtypes: severe and non-severe patients. for each study, we investigated the association between the risk factors and covid- , assuming different calculation models. overall, when all the eligible studies were pooled into the meta-analysis, significant associations were found for elevated levels of crp [ figure ). we also evaluated the dynamics of clinical risk factors during hospital stay between non-severe and severe covid- patients. of the risk factors evaluated, we found the levels of d-dimer, crp, ldh, and pct are significantly higher in severe patients than in non-severe patients (figure ) . moreover, we found that, while levels of crp, ldh, and pct were consistently high in severe patient, d-dimer level still increased in some severe patients, which means it was most likely related to severity (figure ). strikingly, when further stratified the severe patient into two subgroups: recovered (r-severe, cases) and death (d-severe, cases), we found d-dimer kept increasing in d-severe group but dropped in the r-severe group after days from the admission onset, and the levels of d-dimer in these two subgroups were significantly different ( . vs. . , p< . ) after days from the admission onset. these results suggested d-dimer might be more related with the cfr compared with other markers. in addition, we checked the case reports of covid- patients and found that d-dimer was also related with the status of patients, and preventive and adjuvant antithrombotic treatment in some severe patients achieved good therapeutic effect (the representative chest x ray images were shown as figure ). recently, a number of the epidemiological and clinical features of covid- have been reported. phylogenetic studies indicated that it is closely related to a bat-derived sars-like coronaviruses ( ) , and belongs to a large coronavirus family including severe acute respiratory syndrome coronavirus (sars) ( ) and middle east respiratory syndrome (mers) ( ) , which often cause the clinical symptoms ranging from the common fever to severe pneumonia ( ) . it is reported that covid- has higher infection rates compared with sars and mers ( ) . the median daily all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted april , . . reproduction number (rt) of covid- in wuhan declined from . ( % ci, . - . ) ( ) , and the cfr in the severe patient is very high. however, our knowledge about the difference between severe and non-severe covid- patients has not been well established, and the information on prognosis of covid- severity is still lacking. the aim of this study is to elaborate the different clinical features between severe and non-severe covid- patients, and find the risk factors for predicting the severity of covip- . in our cohort, we have stratified patients based on the severity on admission according to international guidelines ( ) . our study has shown covid- are more severe in elder people, male and the patients with comorbidities. fever and cough were the dominant symptoms whereas gastrointestinal symptoms were rare. for the diagnosis markers, we found the d-dimer, crp, ldh, pct were increased quickly in severe group compared with the non-severe group (crp, . % vs. , while asp, alp, ck and creatinine were mildly increased. this means more inflammation, thrombus, myocardial injury or liver and kidney damage existed in severe patients. in addition, we made a meta-analysis of cases with additional related studies to further confirm the risk factors. moreover, we studied the dynamic changes during the process of covid- by timeline tracking. we found crp, d-dimer, ldh, pct kept in higher levels in severe patients. notably, d-dimer still increased quickly in some severe patients and most likely related with the cfr than other markers. it is noticed that another recent cohort study also showed that d-dimer is a critical risk factors for the cfr of covid- ( ) .we checked the case reports of severe patients in our cohort retrospectively, and found that prophylactic antithrombotic therapy effectively improved the prognosis in some patients. thus, we propose that severe patients of covid- can be given preventive anticoagulants or physical therapy. indeed, when there is serious hypoxemia and hypotension, the possibility of embolism should be considered, and thrombolysis should be timely in the absence of thrombolysis contraindication. our study still has several limitations. firstly, it is reported that some patients with positive chest ct findings may have negative rt-pcr results. in that case, there may be selection bias since only the positive cases confirmed by the rt-pcr were included in our study. secondly, although we want to track the timeline of each patient's blood test and other clinical tests, but we did not collect all the data of some patients at each time point for various reasons. this may result in a large standard error of the time axis. in summary, our study showed crp, d-dimer, ldh, pct are the high risk factors related with covid- . especially, d-dimer is a critical indicator for both severity and cfr of covid- . as high cfr in the severe covid- patients need to be resolved immediately, the risk factors explored in our study are important of taking into account the disease severity in practice. though the prognostic factors in our study still need to be further validated by future studies, they should be helpful to provide warning model for predicting severity and mortality in covid- , which is very useful for clinical application. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted april , . we performed a retrospective study on laboratory-confirmed cases with covid- based on clinical indications from hospitals in the hubei provinces, china, from january through march , , were included wuhan tongji hospital and central hospital of wuhan. initially, suspected cases were diagnosed as having fever, respiratory symptoms or pneumonia. confirmed case with covid- was defined as a positive result to real-time reverse-transcriptase polymerase-chain-reaction (rt-pcr) assay using nasal and pharyngeal swab specimens ( ) . the clinical characteristics and laboratory assessments were extracted from electronic medical records. laboratory assessments consisted of complete blood count, blood chemistry, coagulation test, liver and renal function, c-reactive protein, lactate dehydrogenase and creatine kinase. radiologic assessments included chest x-ray and computed tomography (ct). the severity of covid- was defined based on the international guidelines for community-acquired pneumonia ( ) . the ethics committee of wuhan tongji hospital and central hospital of wuhan appoved this study and granted a waiver of informed consent in light of the urgent need to collect clinical data. continuous variables were expressed as the means and standard deviations or medians and interquartile ranges (iqr) as appropriate. categorical variables were summarized as the counts and percentages in each category. we divided covid- patients into severe and non-severe subgroups according to the american thoracic society guideline on admission ( ) . mann-whitney test and two-tailed t-test and were applied to continuous variables, the frequencies of categorical variables were compared using the chi-square test and fisher's exact test as appropriate. the candidate risk factors included age, sex, abnormal laboratory findings. the tests with p value less than . was considered statistically significant. statistical analyses were done using the statistical package for the social sciences (spss) software (version . ). in meta-analysis, we first carried out a computerized literature search of the pubmed, web of science, ebsco and cnki database (prior to march th, ) using the following words and terms: ''covid- '', ''sars-cov- ''. references of the retrieved publications were also screened. we only selected the literatures which distinguished the clinical characteristics between non-severe and severe covid- patients available with the specific numbers of cases. then we combined our own cohort study with the data extracted from the selected literatures, and calculated the odd ratio (or) to assess the association between different risk factors and covid- ( table ) . chi-square-based q-statistic test was performed to evaluate the between-study heterogeneity of the studies. if p < . , the between-study heterogeneity was considered to be significant, we chose the random-effects model (dersimonian and laird model) to calculate the or, otherwise we chose the fixed-effects model (mantel-haenszel model). a pooled or obtained by meta-analysis was used to give a more reasonable evaluation of the association. a z test was performed to determine the significance of the pooled or (p ≤ . suggests a significant or). analyses were performed by stata . software. author contributions: drs wu and wei had full access to all of the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis. drs huang, y., lyu, x., and li, d. contributed equally and share first authorship. study concept and design: wu, x., wei, x., huang, y. acquisition, analysis, or interpretation of data: wu, x., huang, y., wei, x., lyu, x., li, d., wang, y., zou, w. drafting of the manuscript: huang, y., wu, x., wei, x., wang, l., critical revision of the manuscript for important intellectual content: huang, y., lyu, x., and li, d. statistical analysis: huang, y., li, d. administrative, technical, or material support: wu, x., wei, x., huang, y., lyu, x., and li, d. study supervision: wu, x., wei, x. a. ct images of a non-severe patient. after days of treatment, the foci of infection were observed to be significantly absorbed in a -year-old female patient. a and a were ct images before and after treatment. b. ct images of a severe patient. after three weeks of treatment, significant improvement was observed in large area flaps infection of double lower lung of a -year-old female patient. b and b were ct images before and after treatment, respectively. figure shows the association between elevated risk factors and severity of covid- in the meta-analysis: crp (a), ldh (b), pct(c) and d-dimer (d). the squares and horizontal lines correspond to the study-specific odd ratios (or) and % confident incidence (ci). the area of the squares reflects the weight. the diamond represents the summary or and % ci. abbreviations: crp, c-reactive protein; ldh, lactate dehydrogenase; pct, procalcitonin. figure shows temporal changes of crp (a), ldh (b), pct(c) and d-dimer (d) in patients within days from admission onset. we stratified the patients into subgroups: severe, non-severe r-severe (the severe patients recovered) and d-severe (the patient died). data represent mean ± sem. abbreviations: crp, c-reactive protein; ldh, lactate dehydrogenase; pct, procalcitonin. a. the chest x-ray suggested that diffuse large area infection of both lungs accompanied, d-dimer (d-d) reached . mg/l. b. after antiviral therapy and adjuvant antithrombotic treatment with heparin, the bilateral lung infection foci were significantly reduced compared with before, and the level of d-d was found to have subsequently declined to . mg/l. novel coronavirus (covid- ) situation a novel coronavirus from patients with pneumonia in china genomic characterisation and epidemiology of novel coronavirus: implications for virus origins and receptor binding clinical characteristics of coronavirus disease in china clinical features of patients infected with novel coronavirus in wuhan hospitalized patients with novel coronavirus-infected pneumonia in wuhan, china epidemiological and clinical characteristics of cases of novel coronavirus pneumonia in wuhan, china: a descriptive study epidemiologic and clinical characteristics of novel coronavirus infections involving patients outside wuhan clinical characteristics of patients infected with sars-cov- in wuhan the clinical features of the patients with covid- in north-east of chongqing identification of a novel coronavirus in patients with severe acute respiratory syndrome isolation of a novel coronavirus from a man with pneumonia in saudi arabia early dynamics of transmission and control of covid- : a mathematical modelling study diagnosis and treatment of adults with community-acquired pneumonia. an official clinical practice guideline of the clinical course and risk factors for mortality of adult inpatients with covid- in wuhan, china: a retrospective cohort study median (iqr) abbreviations: wbc, white blood cell we thank qi wang, md (tongji hospital), xin long, md (tongji hospital) for collecting the data, and zhongshi zhou,md (hubei university of chinese medicine ) for preparing the table. none of these individuals received compensation for their contributions. key: cord- - a gmzz authors: huh, kyungmin; lee, rugyeom; ji, wonjun; kang, minsun; cheol hwang, in; ho lee, dae; jung, jaehun title: impact of obesity, fasting plasma glucose level, blood pressure, and renal function on the severity of covid- : a matter of sexual dimorphism? date: - - journal: diabetes res clin pract doi: . /j.diabres. . sha: doc_id: cord_uid: a gmzz aims this study aimed to assess whether body mass index (bmi), fasting plasma glucose (fpg) levels, blood pressure (bp), and kidney function were associated with the risk of severe disease or death in patients with covid- . methods data on candidate risk factors were extracted from patients’ last checkup records. propensity score-matched cohorts were constructed, and logistic regression models were used to adjust for age, sex, and comorbidities. the primary outcome was death or severe covid- , defined as requiring supplementary oxygen or higher ventilatory support. results among , patients with confirmed covid- , , ( . %) received checkups and severe covid- occurred in patients ( . %). a bmi of . – . was associated with the outcome among women (aor, . ; % ci,: . – . ) and patients aged – years (aor, . ; % ci, . – . ). an fpg ≥ mg/dl was associated with poor outcomes in women (aor, . ; % ci, . – . ) but not in men. similarly, estimated glomerular filtration rate (egfr) < ml/min/ . m was a risk factor in women (aor, . ; % ci, . – . ) and patients aged < years. conclusions the effects of bmi, fpg, and egfr on outcomes associated with covid- were prominent in women but not in men. coronavirus disease caused by a novel severe acute respiratory syndrome coronavirus (sars-cov- ) has spread worldwide and resulted in more than . million confirmed cases and million deaths as of early october . most patients with covid- develop mild and self-limiting disease; however, a considerable proportion of patients suffer from severe disease characterized by respiratory or multiorgan failure, or hyperinflammatory syndromes. [ ] multiple factors have been associated with the risk of severe or fatal covid- , including hypertension, diabetes mellitus, and chronic cardiopulmonary or renal diseases. [ , ] furthermore, obesity has been reported to increase the risk of poor outcome. [ , ] although these conditions are frequently associated with disease severity and fatality in other infectious diseases, their degree of association with severe covid- remains unclear. the impact of baseline health condition on the disease course is difficult to evaluate using medical records, as the information on premorbid status recorded in admission records and epidemiological reports are often inaccurate or incomplete unless they are collected prospectively on purpose. in addition, plasma glucose levels, body weight, and blood pressure (bp) at admission might already deviate from true baseline due to the effects of acute infection with sars-cov- . as a result, it is plausible that baseline health assessment performed prior to the onset of covid- would provide a more accurate representation of risk factors associated with disease severity. to examine the association between baseline health status and the risk of severe disease in patients with covid- , we performed a case-control study, using data from the nationwide registry of covid- cases and from the biennial health checkup database in south korea. this retrospective study compared bp, and metabolic and kidney function parameters between covid- patients with severe disease (including fatalities) and those with mild-tomoderate disease. the study protocol was approved by the institutional review board of the gil medical center, gachon university college of medicine and science (gfirb - ) with a waiver of consent. health checkup data were extracted from the national health insurance service (nhis) database, which was linked to the korea centers for disease control and prevention (kcdc) covid- patient registry, dedicated to collecting information on all confirmed cases in korea. all korean residents are covered by nhis, which provides universal access to healthcare. in addition, the nhis provides a biennial health checkup to all beneficiaries aged ≥ years; this database was interrogated for data on baseline health status. the participation rate of nhis-provided checkups among the eligible population was . % in . [ ] the korean health checkup data have been previously validated and used in studies that assessed the risk of mortality, cardiovascular events, and diabetes. [ ] [ ] [ ] information on comorbidities (identified using the international statistical classification of diseases and related health problems, th revision) was extracted from the nhis reimbursement database. the last date of data entry was may , . patients aged ≥ years who were diagnosed with covid- and who received the nhis health checkup in or later were included in this study. all included patients were diagnosed with covid- based on nasopharyngeal swab or sputum samples examined with a reverse transcriptase polymerase chain reaction (rt-pcr) for the presence of sars-cov- , as per national guidelines. [ ] the primary outcome was death or severe covid- , defined as requiring any of the following during hospitalization: supplementary oxygen, high-flow nasal cannula, noninvasive ventilation, mechanical ventilation, or extracorporeal membrane oxygenation. data on body mass index (bmi), fasting plasma glucose (fpg), bp, serum lipid levels, and the estimated glomerular filtration rate (egfr) were extracted from last checkup records. bp and fpg were categorized using guidelines from the american college of cardiologists/american heart association and the american diabetes association, respectively (supplementary table ). [ , ] bmi was categorized according to the korean society for the study of obesity guidelines. [ ] total cholesterol (tc), low-density lipoprotein cholesterol (ldl-c), high-density lipoprotein cholesterol (hdl-c), and triglyceride (tg) levels were classified with commonly used cutoffs. finally, egfr was categorized using the kidney disease improving global outcomes guidelines. [ ] study variables were compared between patients with severe and non-severe covid- . baseline demographic and clinical characteristics were compared between patient groups using the χ test for categorical variables and student's t-test for continuous variables. the charlson comorbidity index (cci) was calculated using standard methods and compared using the wilcoxon rank-sum test, as its distribution was non-normal. [ ] adjusted logistic regression models were constructed, based on the following covariates: age, sex, nhis expanded coverage for low household income, and comorbidities. comorbidities were categorized into disease groups (supplementary table ), identified from diagnostic codes included at least twice in reimbursement records covering the past years and dated before covid- onset. to mitigate the impact of confounders, we constructed propensity score-matched cohorts. propensity scores (ps) of severe covid- risk were calculated using logistic regression with the following covariates: age, sex, coverage for low income, and cci. each patient with severe covid- was matched with up to five patients with non-severe disease by greedy neighbor nearest matching. the model was further adjusted in logistic regression analysis, using age, sex, and low-income coverage as covariates. subgroup analyses planned a priori were performed for sex and age group. all statistical tests were two-tailed and the threshold for significance was set at p-values < . . all analyses were performed using sas software (version . ; sas institute inc., cary, nc, usa). among , patients with a confirmed diagnosis of covid- , a total of , ( . %) patients had received an nhis health checkup in or later and were included in the analysis. patients who had received checkups tended to be older and have more comorbidities than those who did not receive checkups (supplementary table ). patients with severe covid- or death comprised . % (n= ) of the total cohort, including deaths ( . %). patients who were male, older in age, with low household income, higher cci, or underlying conditions (except rheumatologic disease) were more likely to develop severe or fatal covid- than patients without these characteristics ( table ). the ps-matched cohort showed smaller differences in matching variables (age, sex, income, and cci), but these differences remained significant. high bmi, classified as class obesity ( . - . ), was associated across subgroups with severe and fatal covid- ( figure ) p= . ). in contrast, patients aged ≥ years with higher-than-normal fpg levels showed a lower risk of severe disease. bp was not associated with the risk of severe or fatal covid- in the total and ps-matched cohorts ( figure ). however, there was a trend toward an association among men (aor, . per mmhg of systolic bp; % ci, . - . ; p= . ); in contrast, hypertension classes did not show any significant association. neither hypertension classes nor systolic bp was associated with disease course among women; a similar lack of association was observed across age groups. a total of , patients had egfr calculated in their health checkup results; of these patients ( . %) had severe disease (supplementary table ). an egfr < ml/min/ . m was associated with a higher risk of severe covid (including fatal disease) (aor, . ; % ci, . - . ; p< . ; figure ). this association was observed among women (aor, . ; % ci, . - . ; p= . ), but not among men (aor, . ; % ci, . - . ; p= . ). moreover, egfr < ml/min/ . m and covid- severity were significantly associated among patients aged < years but not among those aged ≥ years (aor, . ; % ci, . - . ; p= . ). our findings were consistent in the overall and ps-matched cohorts and in linear regression analyses. serum lipid profile was not consistently associated with covid- severity (supplementary table and supplementary figure ). in fact, levels of ldl ≥ mg/dl, hdl < mg/dl, and tg ≥ mg/dl did not affect the risk of severe or fatal disease in the present study. however, there was a trend toward increased risk of severe disease associated with low hdl and high tg levels. in the present study based on a nationwide covid- registry combined with an independent regular health checkup data, the effect of fpg levels and egfr on the risk of severe or fatal covid- varied between sex and age groups. high fpg levels and low egfr were associated with severe covid- among women and patients aged < years; however, this association was not observed among men or older patients. previous studies have reported that case fatality and severity rates were higher among men than among women with covid- . [ ] [ ] [ ] moreover, official sex disaggregated data from > countries and territories have shown that the case fatality rate was lower among women than among men in most countries. [ ] the gene coding angiotensin converting enzyme (ace ) is located on the x chromosome, and estrogen is known to increase the level of ace expression. [ , ] although ace is a receptor used for cell entry of sars-cov- , downregulation of ace expression is associated with lung injury caused by respiratory viruses, including sars-cov. [ , ] this apparent paradox could be explained by the anti-inflammatory function of ace through its role in the renin-angiotensin system (ras). male sex, older age, and sars-cov- binding all lead to lower levels of ace expression, which may cause acute lung injury and organ damage through exaggerated angiotensin ii signaling. [ ] as estrogen is involved in the regulation of ace and its expression level is higher in women, it has been suggested that estrogen levels may have a protective role against respiratory failure and mortality in women with covid- . [ , , ] nevertheless, metabolic abnormalities associated with altered ace expression might counteract these benefits in women with covid- in a manner analogous to that observed in cardiovascular and renal disease, where the risk in women with and without diabetes is lower and higher than in male counterparts, respectively. [ ] [ ] [ ] [ ] it remains unknown whether these documented interactions between sex and metabolic diseaserelated complications is augmented in patients with covid- . further studies are required on the interaction between sex and covid- -related organ damage in patients with specific metabolic characteristics. in the present study, bmi - . was associated with severe or fatal covid- across subgroups, except among males and patients aged < years. obesity has been reported as a significant risk factor for respiratory failure and mortality in patients with covid- . [ ] [ ] [ ] high levels of ace expression by adipocytes, overactive ras (characterized by higher level of angiotensin ii), impaired baseline pulmonary function, endothelial dysfunction, and higher risk of thromboembolism have been proposed as plausible mechanisms linking obesity with the risk of severe or fatal covid- . [ ] our findings are consistent with those from previous reports and meta-analyses. furthermore, the present findings suggest that a bmi - , categorized as "class obesity" in korean guidelines but "overweight" in western guidelines, is a significant risk factor for severe disease among women and patients aged - years. the impact of ethnic differences in metabolic parameters on disease severity should be considered in future research. interestingly, high bp was not associated with the risk of severe disease in any of the present study subgroups. hypertension has been associated with mortality risk since early reports on covid- cases confirmed this in china; this association was replicated in subsequent studies. [ , ] hypertension alters ace expression, increases the risk of cardiovascular and renal failure, and may cause organ damage; thus, the detrimental effect of hypertension on covid- outcomes seems biologically plausible. [ ] however, it remains unclear whether hypertension is an independent rather than a confounding factor for severe covid- . [ ] hypertension is common among older adults and it might reflect their general health status. our findings suggest that further studies are required to elucidate this uncertainty. similarly, serum lipid profiles showed no clear association with the risk of severe or fatal covid- . while levels of tg ≥ mg/dl were marginally associated with increased risk of severe disease in men, this effect was not statistically significant. dyslipidemia considered separately from other components of metabolic syndrome did not affect the risk of severe covid- in the present study. a small number of previous studies investigated cholesterol levels in patients with covid- , reporting lower cholesterol levels in patients with severe disease than in those with mild disease. [ ] however, the study authors measured serum cholesterol levels after the diagnosis of covid- ; thus, it remains unclear whether low cholesterol levels were risk factors for covid- or a consequence of the disease. in the present study, serum lipid levels measured before the diagnosis of covid- suggest a lack of association. our study has several strengths. first, we used a large, nationwide registry of confirmed covid- cases. south korea has successfully controlled the covid- epidemic because of an aggressive trace-and-isolate strategy, made possible owing to a large testing capacity, experience from the previous mers-cov outbreak, and public cooperation. the dataset used in the present study is unlikely to miss a substantial number of the country's cases, including those of mild disease, allowing the present study to account for patients who did not require hospitalization, a task often difficult if not impossible in other countries. second, baseline health status data were extracted from a health checkup database, created before the covid- pandemic. as a result, our data were likely an accurate and objective representation of patients' health status before they acquired the sars-cov- infection. however, this study has some limitations. first, less than a half of patients diagnosed with covid- had data available from a health checkup performed in or later. patients who were older or had more comorbidities were more likely to have received checkups; thus the baseline characteristics of study participants differed from those of patients excluded from the study due to the lack of checkup data. nevertheless, we compared variables of interest in a robust design, adjusting for underlying conditions and using a ps-matched analysis to mitigate confounding. second, there was a time gap between the last checkup and sars-cov- infection. some of our study variables are prone to temporal change and the measurements recorded during checkups might not fully reflect patient status immediately before infection. finally, we could not account for the effect of treatment in our analysis. in our retrospective study using a nationwide health checkup database, high fpg levels and low egfr were significantly associated with the risk of severe covid- (including fatal illness among women. no consistent increase in risk was observed in association with high bp or dyslipidemia. these findings suggest that the baseline metabolic characteristics exert differential sex-and age-related effects on disease severity among patients diagnosed with covid- . characteristics of and important lessons from the coronavirus disease (covid- ) outbreak in china: summary of a report of cases from the chinese center for disease control and prevention case-fatality rate and characteristics of patients dying in relation to covid- in italy presenting characteristics, comorbidities, and outcomes among patients hospitalized with covid- in the new york city area obesity and covid- : a call for action from people living with obesity national health screening statistical yearbook predictors of all-cause mortality among , participants from the korean national health screening cohort incremental value of repeated risk factor measurements for cardiovascular disease prediction in middle-aged korean adults: results from the nhis-heals (national health insurance system-national health screening cohort) development and validation of the korean diabetes risk score: a -year national cohort study guideline for the prevention, detection, evaluation, and management of high blood heart association task force on clinical practice guidelines american diabetes association. . classification and diagnosis of diabetes: standards of medical care in diabetes- korean society for the study of obesity guideline for the management of obesity in korea summary of recommendation statements a new method of classifying prognostic comorbidity in longitudinal studies: development and validation risk factors for mortality and respiratory support in elderly patients hospitalized with covid- in korea host susceptibility to severe covid- and establishment of a host risk score: findings of cases outside wuhan a comparison of mortality-related risk factors of covid- , sars, and mers: a systematic review and meta-analysis covid- sex-disaggregated data tracker the "his and hers" of the reninangiotensin system landscape of x chromosome inactivation across human tissues a crucial role of angiotensin converting enzyme (ace ) in sars coronavirus-induced lung injury angiotensin-converting enzyme inhibits lung injury induced by respiratory syncytial virus the dilemma of coronavirus disease , aging, and cardiovascular disease: insights from cardiovascular aging science estrogen regulates the expression of sars-cov- receptor ace in differentiated airway epithelial cells sex-related differences in covid sex and gender differences in risk, pathophysiology and complications of type diabetes mellitus incidences, treatments, outcomes, and sex effect on survival in patients with end-stage renal disease by diabetes status in australia and new zealand excess mortality among persons with type diabetes gender differences in the progression of target organ damage in patients with increased insulin resistance: the lod-diabetes study obesity is a risk factor for developing critical condition in covid- patients: a systematic review and meta-analysis obesity and mortality of covid- . meta-analysis obesity aggravates covid- : a systematic review and metaanalysis obesity and outcomes in covid- : when an epidemic and pandemic collide arterial hypertension and risk of death in patients with covid- infection: systematic review and meta-analysis patients with arterial hypertension and covid- are at higher risk of icu admission association of hypertension, diabetes, stroke, cancer, kidney disease, and high-cholesterol with covid- disease severity and fatality: a systematic review covid- and arterial hypertension: hypothesis or evidence? key: cord- - d o w z authors: omori, ryosuke; mizumoto, kenji; nishiura, hiroshi title: ascertainment rate of novel coronavirus disease (covid- ) in japan date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: d o w z we analyzed the epidemiological dataset of confirmed cases with covid- in japan as of february and estimated the number of severe and non-severe cases, accounting for under-ascertainment. the ascertainment rate of non-severe cases was estimated at . ( % confidence interval: . , . ), indicating that unbiased number of non-cases would be more than twice the reported count. severe cases are twice more likely diagnosed and reported than other cases. would be more than twice the reported count. conclusions: severe cases are twice more likely diagnosed and reported than other cases. considering that reported cases are usually dominated by non-severe cases, the adjusted total number of cases is also about a double of observed count. our finding is critical in interpreting the reported data, and it is advised to interpret mild case data of covid- as always under-ascertained. keywords: coronavirus cc-by-nc-nd . international license it is made available under a author/funder, who has granted medrxiv a license to display the preprint in perpetuity. is the (which was not peer-reviewed) the copyright holder for this preprint . the majority of covid- cases exhibit limited severity; % of reported cases in china has been mild and only % are severe (guan et al., ) . it is natural that the ascertainment rate would be different between severe and non-severe cases. the present study aims to estimate the ascertainment rate of non-severe cases, employing a statistical model. we analyzed the epidemiological dataset of confirmed cases with covid- in japan as of february . the confirmatory diagnosis was made by means of reverse transcriptase polymerase chain reaction (rt-pcr). the present study specifically analyzed cases by (i) prefecture, (ii) age, and (iii) severity. severe case was defined as (i) severe dyspnea that required oxygen support plus pneumonia or intubation or (ii) case that required management in intensive care unit. we estimated the number of severe and non-severe cases using the ratio of non-severe to severe reported cases (guan et al., , novel, . we estimated the ascertainment rate among non-severe cases by /k, describing data generating process of both severe and non-severe generated from poisson process with probabilities p x,a for severe cases and kf a p x,a for non-severe cases in age . cc-by-nc-nd . international license it is made available under a author/funder, who has granted medrxiv a license to display the preprint in perpetuity. is the (which was not peer-reviewed) the copyright holder for this preprint . https://doi.org/ . / . . . doi: medrxiv preprint group a and prefecture x, respectively. here f a denotes the ratio of non-severe to severe reported case of age group a, as estimated from age-specific severity and incidence rate ratio in china (guan et al., , novel, . we estimate k and p x,a using the loglikelihood function: where n x,a , d ns,x,a and d s,x,a represent the population size, the observed counts of non-severe and severe cases of age group a in prefecture x, respectively. maximum likelihood estimates were obtained by maximizing the equation ( ) and the profile likelihood-based confidence intervals were computed. the ascertainment rate of non-severe cases, k, was estimated at . ( % confidence interval (ci): . , . ). resulting estimate of non-severe cases is shown in figure a , showing along with reasonably good fit to severe case data in figure b . age-specific pattern of estimated non-severe cases was similar to that among severe cases. the largest estimated number of non-severe cases was cases ( % ci: , ) among those aged - years and ( % ci: , ) among cases aged - years, respectively. such adjustment gives adjusted estimate of the total cases by age group. the present study estimated the ascertainment-adjusted number of cases in japan, using age-specific severe fraction of cases. we assumed that the ratio of severe to non-severe cases in a given age group is a constant and that the age- independent gap is explained by the under-diagnosis and under-reporting, estimating the ascertainment rate among non-severe cases to be . . . cc-by-nc-nd . international license it is made available under a author/funder, who has granted medrxiv a license to display the preprint in perpetuity. is the (which was not peer-reviewed) the copyright holder for this preprint . https://doi.org/ . / . . . doi: medrxiv preprint as a take home, it must be remembered that severe cases are twice more likely diagnosed and reported than other cases. reported cases are usually dominated by non-severe cases, and the adjusted total number of cases is about a double of observed count. our finding is critical in interpreting the reported data, and it is advised to regard the mild case data as always under-ascertained. in addition to the proposed adjustment, it should be noted that the ascertainment rate of severe cases needs to be additionally estimated, and such estimation requires direct measurement of the total number of cases or infected individuals by means of seroepidemiological study or other testing methods of all samples (nishiura et al., ) . that is, the actual total number of cases is greater than what it was adjusted in the present study. using seroepidemiological datasets, we plan to address relevant issues in the future. other limitations include that (i) we did not explore detailed natural history, e.g. dynamically changing symptoms over the course of infection, and underlying comorbidities, (ii) we ignored right-censored data, e.g. the time delay from illness onset to severe manifestations, for simplicity. the latter led us to underestimate the ascertainment rate. (iii) it is worth noting that the data of age dependent severity employed in our analysis is only based on the observed data in china. considering the possibility of underreporting or biased age distribution, the nature of this age distribution may lead to underestimation. despite multiple future tasks, we believe that the present study successfully demonstrated that the ascertainment rate can be partly adjusted by examining age-dependent number of cases including severe cases. the proposed adjustment should be practiced in other country settings and also for other diseases. . cc-by-nc-nd . international license it is made available under a author/funder, who has granted medrxiv a license to display the preprint in perpetuity. top: non severe cases, middle: severe cases, and bottom: total cases. x-marks represent observed counts, while unfilled circles show estimated cases. whiskers extend to lower and upper % confidence intervals, derived from profile likelihood. . cc-by-nc-nd . international license it is made available under a author/funder, who has granted medrxiv a license to display the preprint in perpetuity. is the (which was not peer-reviewed) the copyright holder for this preprint . https://doi.org/ . / . . . doi: medrxiv preprint clinical characteristics of coronavirus disease in china the rate of underascertainment of novel coronavirus ( -ncov) infection: estimation using japanese passengers data on evacuation flights. multidisciplinary digital publishing institute the epidemiological characteristics of an outbreak of novel coronavirus diseases (covid- ) in china. zhonghua liu xing bing xue za zhi= covid- ) situation report - key: cord- -gmcugd h authors: song, jian-xin; zhu, lin; zhu, chuan-long; hu, jin-hua; sun, zi-jian; xu, xiang; xin, min-you; zhang, qiong-fang; zhang, da-zhi; shang, jia; huang, jia-quan; xu, dong title: main complications of aechb and severe hepatitis b (liver failure) date: - - journal: acute exacerbation of chronic hepatitis b doi: . / - - - - _ sha: doc_id: cord_uid: gmcugd h this chapter describes the clinical features, and diagnosis of complications in aechb including secondary bacterial infections, coagulation disorder, water electrolyte disorder, hepatorenal syndrome, hepatic encephalopathy, hepatopulmonary syndrome and endotoxemia: . patients with severe hepatitis have impaired immunity and are therefore vulnerable to all kinds of infections. after infection, these patients may experience shock, dic and multiple organ failure, all of which seriously affect their prognosis and are major causes of death. concurrent infections consist primarily of infections of the lungs, intestines, biliary tract, and urinary tract, as well as spontaneous bacterial peritonitis and sepsis. . severe hepatitis may reduce the synthesis of coagulation factors and enhance their dysfunction and increase anticoagulants and platelet abnormalities, leading to coagulopathy. infection, hepatorenal syndrome and complications can further aggravate coagulopathy, resulting in dic and seriously affecting patient prognosis. . hepatorenal syndrome, which is characterized by renal failure, hemodynamic changes in arterial circulation and abnormalities in the endogenous vascular system, is a common clinical complication of end-stage liver disease, and one of the important indicators for the prognosis of patients with severe hepatitis. . water electrolyte disorder (water retention, hyponatremia, hypokalemia, hyperkalaemia) and acid-base imbalance are common in patients with severe hepatitis. these internal environment disorders can lead to exacerbation and complication of the illness. . hepatic encephalopathy is a neurological and psychiatric anomaly syndrome based on metabolic disorder, and an important prognostic indicator for patients with severe hepatitis. . the hepatopulmonary syndrome is an important vascular complication in lungs due to systemic hypoxemia in patients with cirrhosis and portal hypertension. the majority of patients with hps are asymptomatic. long-term oxygen therapy remains the most frequently recommended therapy for symptoms in patients with severe hypoxemia. . endotoxemia, an important complication of severe hepatitis, is not only a second hit to the liver, but also leads to other complications including sirs and mods. (nacseld) for survival analysis of prospectively enrolled cirrhosis patients hospitalized with infections, bajaj found that there is . % of nosocomial infection in all patients [ ] . domestic data showed significant difference of infection incidence in different times. it is related to raising diagnosis and treatment level. according to the data from shanghai public health clinical center, in patients who were treated as severe hepatitis from january to december , there were patients obtained nosocomial infection with a rate of . % [ ] . the incidences of infection complicated with acute liver failure, acute on chronic liver failure or chronic liver failure are similar. patients with old age or long hospitalization are more easily to get infection. there was rare report about infection in children patients with severe hepatitis. in , godbole et al. from uk reported that % in children patients with severe hepatitis were complicated with infection, and mostly are sepsis, respiratory tract infection and urinary infection. most of the infections occurred within weeks of admission, while patients with infection had longer hospitalization [ ] . reduced innate immunity the innate immune system is the first line of defense against premier environmental challenges and injury. in liver, it is a complex system that includes nk cells, nkt cells, kcs, neutrophils, eosinophils and complement components. the innate immune response acts much more rapidly compared with adaptive immunity. monocytes and macrophages: the liver plays an important role in defense and immune function. the main cellular components of the innate immune system within the liver are the kupffer cells. kupffer cells represent - % of the tissue macrophages in the human body [ ] . in normal condition, kupffer cells in liver help to clear the macromolecular substance such as pathogen, endotoxin, heteroantigen and immune complex to defense infection. in severe hepatitis, due to massive hepatocytes necrosis, the number and function of monocytes/macrophages are impaired, thus the activity of fibronectin, which is opsonin of macrophages, is decreased. therefore, bacterium, endotoxin and other poison from gut directly access into circulation. subrat kumar acharya from india reported that the plasma fibronectin (fn) level in severe hepatitis patients was significantly lower than that in healthy controls ( . μg/ml ± . μg/ml, vs. . μg/ ml ± . μg/ml). the fn level was remarkably correlated with incidence of infection and mortality [ ] . complement: liver is the organ where complements are mainly produced, such as c , c , c , c . the complements help to expand phagocytosis of phagocytes by chemotaxis, opsonization or adhesion, as well as help antibody to kill or solute some gram negative bacilli [ ] . report from wyke showed that the defect of complement closely correlated with impaired opsonization [ ] . defect of c or c can result to weakened movement of polymorphonuclear leucocyte. in severe hepatitis, the ability of liver to produce complement has been weakened due to massive injury of parenchymal hepatic cells, which leads to decreased activity of complement to % of normal condition. meanwhile, serum opsonization to e. coli or saccharomycetes are also diminished. besides, high ammonia level in severe hepatitis also restrains complement activity to impact germicidal effect. the most direct and also the most important result for decreased complement production and reduced opsonization is the susceptibility to infection. it was also reported that complement and the alternative pathway play an crucial role in lps/d-galn-induced fulminant hepatic failure [ ] . neutrophils: a majority of patients with severe liver disease have altered function of neutrophil granulocytes [ ] . the most common manifestation include abnormal ultrastructure or function of neutrophil granulocytes, as cytoplasm degeneration, organelle reduction, mitochondrial swelling, pyknotic nuclei, etc. decrease filtration and phagocytosis of reticuloendothelial system, as well as impaired chemotaxis of blood cells, making immunity weakened, lead to invasion of bacterium. therefore, patients with severe hepatitis are vulnerable to be infected with bacterium or fungi due to decreased phagocytosis and germicidal effect of neutrophil granulocytes, and impaired adhesion of macrophages and white cells [ ] . data from liu h demonstrated pretreatment neutrophil-lymphocyte ratio was associated with the prognosis of patients with hbv-aclf, and elevated nlr predicted poor outcome within weeks [ ] . natural killer and natural killer t cells: natural killer (nk) and natural killer t cells (nkt) are important components of the innate immune response. natural killer cells have potent cytolytic activities that are exerted through the death receptor and perforin/granzyme pathways. activated nkt cells have both perforindependent and fas-ligand dependent cytotoxic function that are triggered upon tcr recognition of an antigen [ ] . nk cells and nkt cells play an important role in many experimental models of liver injury, such as viral hepatitis, alcoholic liver disease, and autoimmune liver disease [ ] . however, their role in aclf has not yet been clearly elucidated, it was reported the median percentage of nk cells in the lymphocytes of patients with acute and fulminant liver failure were significantly lower compared to healthy controls. meanwhile, patients with acute and fulminant liver failure had significantly high and comparable nkt cells compared to control group [ ] . the important pathophysiological role of innate immune dysfunction in patients with acute-on-chronic liver failure (aclf) has been investigated in recent years. however, dysregulation of adaptive immunity remains poorly elucidated [ ] . patients with severe hepatitis has varying degrees of impaired cellular immunity, manifested as decreased cd + cell number and declined cd +/cd + ratio, which is pathogenesis of opportunistic infection. it was reported that there exists a reduction in cd (+) t lymphocytes in hbv-aclf patients. these cd (+) t cells predominantly are cd (+) tconv, and the development of suppressive cd (+) tregs greatly surpass tconv, which constitutes important characteristics of adaptive immune dysfunction of hbv-aclf [ ] . a report from china showed total amount of lymphocytes, cd (+) t cells, cd (+) t cells and nk cells in circulation were lower in the hbv-aclf patients compared to the chb patients [ ] . hbv-specific cd (+)t-cell responses are considered to be of great importance in viral control and immune-mediated liver damage [ ] . however, cd (+) t cell has seldom been studied in aclf. ye [ ] reported decreased activated cd (+) t cells may be related to poor outcomes in patients with sh. the frequency of circulating th cells increased with disease progression from chb to aclf patients compared to healthy control. th cells were also found largely accumulated in the livers of chb patients. the increases in circulating and intrahepatic th cells were positively correlated with plasma viral load, serum alanine aminotransferase levels and histological activity index. in addition, the serum concentration of th -associated cytokines was also augmented in both chb and aclf patients [ ] . in process of diagnosis and treatment for severe hepatitis, repeatedly abdominal paracentesis, retention catheterization, venous cannula, hemofiltration, or trachea cannula are usually necessary. unthoroughly sterilization or nonstandard sterile operation will lead to pathogen invading to develop an infection. in addition, artificial liver treatment is also an important cause of infection. it was reported that the incidence of fungal infection in severe hepatitis correlated with the number of artificial liver treatment. application of broad spectrum antibiotics is also a major cause of infection in severe hepatitis. antibiotics inhibit or kill sensitive normal bacterium as well as pathogens, especially normal bacterium colonized in natural orifice, leading to flora disproportionality. this time, nonpathogen could cause infection, or mass produced pathogen become dominant colony to develop infection. it was proved that dosage, exposure time, or varieties of antibiotics used in patients was closely correlated with severity of dysbacteriosis and incidence of sbp [ ] . nosocomial origin of infection, longterm of norfloxacin prophylaxis, history of recent infection by multiresistant bacteria and recent use of β-lactams were independent inducements associated with the development of multiresistant infections. bacterial translocation is defined as the migration of viable microorganisms or bacterial products (i.e., bacterial lps, peptidoglycan, and lipopeptides) from theintestinal lumen to the mesenteric lymph nodes and other extraintestinal sites [ ] . there are multiple mechanisms which are involved in defective gut functions and altered microbiota in patients with cirrhosis or liver failure. these include small intestine bacterial overgrowth (sibo), increased intestinal permeability, and impaired antimicrobial defense. additionally, decreased bile acids, due to decreased synthesis and defective enterohepatic circulation, contribute to altered gut microbiota [ ] . small intestinal bacterial overgrowth (sibo) was defined as ≥ total colonyforming units per milliliter of proximal jejunal aspirations, which presents in % of cirrhotic patients and is associated with systemic endotoxemia [ ] . in the condition of hepatic failure, due to impaired immunity, bacterium overgrowth and translocation happened, which increased incidence of infection [ ] . in vivo study from wang showed phagocytosis of monocytes were prominently and immediately diminished after resection of liver tissue up to %, accompanied by decreased oxygen ingestion [ ] . mass propagation of e. coli in the lower part of the small intestine and bacterium translocation were all happened h after resection. in rats with ascites in severe liver disease induced by carbon tetrachloride, the bacteria easily transferred from the intestine to extra-intestine, including local lymph nodes and blood circulation. these evidences proved that liver failure prompted the proliferation of intestinal bacterial overgrowth and translocation, which may be the main reason for the endogenous infections in patients with liver failure. another reason for bacterial translocation is that patients with cirrhosis or liver failure display increased intestinal permeability. in patients with severe hepatitis, gastrointestinal mucosa membrane has impaired ability for regeneration and repair, leading to dysfunction of intestinal barrier and declined anti-infection ability. patients with severe hepatitis have weakened regeneration and repair capacity of gastrointestinal mucosa, as well as decreased gastrointestinal barrier function and resistance to infection. the intestinal mucosa often manifested as congestion, edema or erosion. especially when intestine get infection, bacteria can invade to abdominal cavity directly or through slight mucosal defect [ ] . besides, ascites often present in severe hepatitis patients, which provide a good medium for bacteria to multiply. patients with severe hepatitis often have ascites combined with portal hypertension. due to lobular structural damage, abnormal construct of liver sinusoids or blood vessels, bacteria directly access into the systemic circulation without filtration and phagocytosis by liver, leading to formation of bacteremia. meanwhile, bacteria in blood circulation may access into the abdominal cavity and cause abdominal infections. hence, patients with severe hepatitis are not only prone to get endogenous infection but also prone to have intestinal endotoxemia. patients with severe hepatitis have decreased bile secretion, changes of bile composition, so that infection are prone to occur in epithelium of bile duct and gallbladder. in patients with severe hepatitis, intestinal edema and cellulitis are obvious, some may develop acute serositis. for patients with portal hypertensive gastrointestinal disease, regeneration and repair capacity of gastrointestinal mucosa are decreased, accordingly, the natural immune barrier function is reduced. once esophageal and gastric venous bleeding occur, gastrointestinal ischemia aggravate, which result in decreased resistance to infection. invasive procedures, complications, prophylactic use of broad-spectrum antibiotics, underlying diseases, long hospital stay, and old age are risk factors for infection in patients with severe hepatitis. elderly patients with other underlying diseases, decreased immune function, more severe primary disease, have a high risk of infection and may predispose to severe infections. in addition, albumin level is closely related to ascites production and sbp occurrence. irrational uses of immunosuppressive agents such as corticosteroids suppress immune function, lead to flora, and promote the formation of drug-resistant strains and pathogenic bacteria. mortality of severe hepatitis is closely related to infection, which directly affects the prognosis of severe hepatitis. to reduce the incidence of infection in patients with severe hepatitis, we should aim at a variety of risk factors, improve patients' immunity, rectify hypoproteinemia, treat the primary disease, prevent complications, strengthen disinfection and isolation, operate with strict aseptic technique and strict indications for invasive procedures, and use antibiotics rationally. once patients with severe hepatitis are infected, the consequences are often serious, as infection easily diffuse, which is difficult to control. after infection, the bacteria produce more toxins, which aggravate liver disease and cause a series of adverse effects, finally lead to severe complications (hepatic encephalopathy, hepatorenal syndrome, and etc.), which is the major cause for death in patients with severe hepatitis. bacterial endotoxin plays a major role in development and prognosis of severe hepatitis when infection happened [ ] . endotoxemia may directly or indirectly worsen liver injury. intrahepatic cholestasis positively correlated with endotoxin levels. endotoxin can trigger hepatorenal syndrome. for kidney, endotoxin is a potent renal vasoconstrictor substance, which can make a strong contraction of the renal artery and renal blood flow reduction. in addition, endotoxin can cause kidney capillary thrombosis and acute renal tubular necrosis, or even renal cortex necrosis. endotoxin can activate coagulation factor vii and factor vi, making the intrinsic coagulation system startup, as well as directly damage hepatocytes which release tissue thromboplastin to start the extrinsic coagulation system. meanwhile, endotoxin can also damage the vascular endothelial cells to cause bleeding, especially upper gastrointestinal bleeding. in the event of gastrointestinal bleeding, infections are more prone to happen or primary infection aggravates. in recent years, due to extensive use of broad-spectrum antibiotics, the mortality of severe hepatitis has been significantly reduced. however, infection is still an important cause of death. nosocomial infection control directly affects the prognosis of patients with severe hepatitis, which is an important part for the treatment of severe hepatitis. once the etiology and other evidence of infection appear, patients should be treated with antibiotics. effective prevention and treatment for nosocomial infections are positive to promote recovery and reduce mortality. bacterial translocation (bt) is the key mechanism in the pathogenesis of sbp, which is because of the concurrent failure of defensive mechanisms in liver failure [ ] . investigators have demonstrated pronounced impairment of gastrointestinal tract motility in cirrhosis [ ] . the disturbance of gut flora microecology was associated with changes in the (ultra)structure of the gastrointestinal tract [ ] . meanwhile, reduced cellular and humoral immunity make it easier for the free flow of microorganisms and endotoxins to the mesenteric lymph nodes [ ] . besides, it was reported that hypoalbuminemia was related to sbp. a prospective study from runyon reported that the incidence of sbp in patients with ascites protein less than . g/l was %, while the number in patients with ascites protein more than . g/l was only %. after years of follow-up, sbp incidence in patients with ascites protein higher than . g/l were negligible [ ] . the level of ascites protein correlates with opsonin activity, which means ascites protein < . g/l was an independent risk factor for sbp. the clinical manifestations of sbp are diverse. most patients have subtle and insidious onset, which usually manifested as abdominal pain and fever. acute onset may burst chills, fever and abdominal pain. generally the body temperature of patient is around °c, sometimes as high as °c. fever type usually presents as remittent fever. abdominal pain is always around the navel or lower abdomen, paroxysmal or persistent. nausea and vomiting are obvious, sometimes with diarrhea. ascites can occur in patients without ascites, or increased significantly. patients may also have significant bloating, abdominal muscle tension, tenderness, rebound tenderness, diminished or disappeared bowel sounds, and so on, or even shock in severe cases. according to clinical manifestations, sbp can be divided into five types: . common type: acute onset, protruding abdominal pain, followed by fever. or irregular fever take place followed by abdominal pain, abdominal tenderness and rebound tenderness, mild to moderate abdominal tension and growing ascites. increased wbc count and nuclear left shift. routine examination of ascites showed acute inflammatory changes. . shock: septic shock break out in a few hours to one day after abdominal pain or fever. the clinical manifestations include low temperature, lip cyanosis, abdominal tenderness, and hardly relieved shock. wbc count increases, blood culture is positive. . encephalopathy: fever and abdominal pain are often obscure, the early emergence of trance and other neuropsychiatric symptoms rapidly develop into a coma. careful examination of the abdomen in patients with light coma may still find in patients with pain expression. jaundice is deep, liver damage is serious. . refractory ascites: ascites progressively increase, which is difficult to subside with invalid diuretic therapy. abdominal distension is obvious, often without pain. carefully check the abdomen may still find slight peritoneal irritation. . asymptomatic: symptoms are mild, exhibited as slight bloating, occasional fever. mild tenderness can be found by deep palpation. in addition, a considerable part of the patients showed non-specific symptoms and signs, such as deepened coma, deepening jaundice, oliguria, azotemia or dramatically increased ascites. diagnosis of sbp should be considered if following conditions exist: ( ) fever, which can't be explained by other reasons or other parts of infection; ( ) abdominal pain, abdominal tenderness or rebound tenderness, but not serious; ( ) sudden increased ascites or poor diuretic effect manifested as refractory ascites; ( ) sudden onset of septic shock; ( ) rapid deterioration of general condition, or deteriorated liver and kidney function in a short term, deepening jaundice, or hepatic encephalopathy. the diagnoses of sbp mainly rely on ascites puncture. easl clinical practice guidelines recommend that a diagnostic paracentesis should be performed in all patients with new onset grade or ascites, and in all patients hospitalized for worsening of ascites or any complication of cirrhosis (level a ) [ ] . for patients with severe hepatitis, diagnostic paracentesis indications are: . in liver cirrhosis patients with ascites that paracentesis should be performed after admission to determine whether sbp exist. . if the following conditions happened during hospitalization, diagnostic paracentesis should be performed: ( ) abdominal sighs suggest abdominal infections, such as abdominal pain, rebound tenderness and gastrointestinal symptoms (such as vomiting, diarrhea, intestinal paralysis); ( ) systemic infection signs such as fever, leukocytosis, or septic shock; ( ) no clear incentive for hepatic encephalopathy, or rapidly emerged renal dysfunction. . in patients with ascites and gastrointestinal bleeding, paracentesis should be performed before prophylactic antibiotics. once ascites was acquired, polymorphonuclear cells (polymorphonuclears, pmn) count and ascites culture should be performed. diagnosis of sbp is made according to international guidelines [ , ] in patients with liver cirrhosis if the ascites polymorphonuclear (pmn) cell count exceeds cells/μl and other forms of peritonitis have been excluded. an ascites fluid polymorphonuclear (pmn) leukocyte count ≥ /mm should be considered as sbp, if pmn > /mm can be confirmed as sbp. if there is bloody ascites (erythrocytes more than , /mm ), pmn count are as / of red blood cells. the leukocyte esterase reagent strips (lers) test is based on the esterase activity of the leucocytes. since , studies have examined the validity of using leukocyte esterase reagent strips (lers) in sbp diagnosis. lers appeared to have low sensitivity for sbp. on the other hand, a high negative predictive value (> % in the majority of the studies)supported the use of lers as a preliminary screening tool for sbp diagnosis [ ] . there is bacteria colonization in ascites but no inflammation. the diagnosis is based on positive ascites culture, ascites pmn countless than /mm , without evidence of systemic or local infection. bacterascites have two outcomes: a shortterm or transient bacterial ascites (mostly asymptomatic), or the development of sbp (mostly with symptoms). once the diagnosis was established, paracentesis examination should be conducted again after - days. take appropriate action under the circumstances. if the second sample has a pmnl count > /mm , treat as for sbp. if the pmnl count is< /mm and a second set of cultures is positive, treat as for sbp. if the pmnl count is < /mm and the second set of cultures is negative, no further action is recommended [ ] . if ascites culture is positive but ascites pmn < /mm and there were signs of abdominal infection, it usually progress to sbp within a few days. these patients should be given appropriate antibiotic therapy. brolin first proposed the concept in . the mechanism: in early stage of severe hepatitis, ascites has not been exist yet, however because of necrosis of hepatocytes, dysfunction of kupffer cell, impairment of liver immune barrier, intestinal bacteria easily invasive into the systemic circulation through liver, causing spontaneous bacteremia, and then sbp. diagnostic criteria: ( ) primary disease was severe hepatitis, no ascites was detected by strict examination and ultrasound. ( ) clinical manifestation include fever, varying degrees of abdominal pain, diarrhea, diffuse abdominal tenderness and rebound tenderness, increased peripheral blood leukocyte, positive rivalta's test, ascites fluid leukocyte count> /mm , or pmn > /mm . ( ) exclude abdominal organ perforation or primary foci. it was used to describe the clinical situation when the ascitic pmnl count is > /mm but cultures fail to grow any bacteria. however, the term is now considered obsolete. in severe hepatitis with secondary infection, pneumonia is most common. patients with hepatic encephalopathy are susceptible to pulmonary infections as pneumonia since bed-ridden, impaired cough reflex and inadequate ventilation, especially comatose patients with intubation and tracheotomy. inpatients underwent thoracentesis, paracentesis and other invasive procedures, non-specific damage to immune barrier provide conditions for the bacterial invasion. reported in patients with invasive procedures, lung infection risk increased significantly, which demonstrated blood infection is an important way for pulmonary infection. furthermore, there was a significant increase of pulmonary infection in patients with intestinal infections or abdominal infection. pathogen: in recent years, pneumonia was classified as "community acquired pneumonia" (community acquired pneumonia, cap) "and" nosocomial pneumonia (hospital acquired pneumonia, hap). cap is the pneumonia acquired outside hospital [ ] . although cap can be caused by a wide variety of micro-organisms, the pneumococcus, mycoplasma pneumoniae and chlamydophila pneumoniae, staphylococcus aureus and certain gram-negative rods are more usual pathogens encountered [ ] . hap is the pneumonia that develops h or more after hospital admission and that was not incubating at hospital admission. hap infected with gram-negative bacilli accounts for more than %, of which pseudomonas aeruginosa, klebsiella pneumoniae, escherichia coli, acinetobacter baumannii, other aeruginosa are common bacteria. the primacy is pseudomonas aeruginosa, while gram-positive cocci accounted for about %, mainly are staphylococcus aureus, coagulase-negative staphylococci, viridans and streptococcus pneumoniae. anaerobic is rare. clinical manifestations and diagnosis: clinical manifestations are characterized as fever, cough, expectoration, dyspnea, and cyanosis. diagnostic criteria: . chest percussion of dullness, auscultation of rales, along with one of the following conditions: ( ) purulent sputum or change of phlegm; ( ) positive pathogens in sputum culture. . the chest x-ray and/or ct examination revealed new or progressive exudative lesions, and the emergence of one conditions above. urinary tract infection is divided into upper urinary tract infection and lower urinary tract infection. upper urinary tract infections are mainly pyelonephritis, which can be manifested as acute pyelonephritis and chronic pyelonephritis. lower urinary tract infections include urethritis and cystitis. the most common pathogen of urinary tract infection is escherichia coli, followed by enterococcus faecalis, klebsiella pneumoniae and streptococcus agalactiae. urinary tract infections often occur in patients with indwelling catheter, therefore, aseptic urethral catheterization and management and timely replacement of catheterization are effective to prevent such infections [ ] . . acute pyelonephritis: ( ) acute onset; ( ) chills, chills; ( ) fever; ( ) malaise, headache, fatigue; ( ) loss of appetite, nausea, vomiting; ( ) urinary frequency, urgency, dysuria; ( ) low back pain, kidney area discomfort; the ( ) tenderness of upper ureter point; ( ) tenderness of rib waist point; ( ) percussion pain in kidney area or bladder. . chronic pyelonephritis: ( ) acute onset of acute pyelonephritis with the same, but usually much lighter, even without fever, malaise, headache and other systemic manifestations, urinary frequency, urgency, dysuria and other symptoms are not obvious; ( ) edema; ( ) hypertension. . urocystitis and urethritis: frequent urination, urgency, dysuria, urinary bladder pain. urethral secretions. laboratory tests and diagnosis: diagnosis elements include: ( ) tenderness in rib waist point, percussion pain in kidney area. ( ) urine leukocytosis, pyuria. ( ) urinary sediment smear find bacteria. ( ) positive in urine culture. ( ) urinary colony counts> /ml; in patients with urinary frequency and other symptoms, colony counts> /ml are meaningful; counts - /ml also helpful in diagnosis; ( ) one hour urine wbc count> , . ( ) blood test showed leukocytosis, neutrophil nucleus left. ( ) increased esr. biliary tract infection is a common complication in severe hepatitis, which is often in company with cholelithiasis to reinforce each other. hepatitis b virus can directly violate bile duct cells and cause cholecystitis. on this basis, cholelithiasis and secondary bacterial infections are easy to happen and become a major focus, which can cause other parts of infection in severe hepatitis patients. furthermore, severe hepatitis patients often have reduced gastric acid secretion, so that e. coli in duodenum easily reproduce and cause ascending infection. biliary tract infection is usually a mixed infection of aerobic and anaerobic infections. enteric gram-negative bacteria include escherichia coli, klebsiella, enterobacter, proteus and enterococcus. anaerobic bacteroides include clostridium and fusobacterium, in which bacteroides is common, about - %, particularly bacteroides fragilis. common symptoms of biliary tract infection include chills, fever, nausea, vomiting, right hypochondrium pain and tenderness in gallbladder area. clinical performance of concurrent biliary tract infection in severe hepatitis is not always very clear, often unconfirmed by pathogen test. symptoms were epigastric or right upper quadrant pain, radiating to the right shoulder, hours after a heavy meal or a high fat meal. the patient may have severe colic, often accompanied by nausea and vomiting. patients with chronic biliary tract infection have indigestion symptoms as heartburn, belching, acid reflux and bloating, or sometimes fever and right upper quadrant pain. severe hepatitis patients have weakened intestinal resistance, especially for reduced immunoglobulin a secretions, which creates good invasion opportunities for bacterium. in addition, as mentioned above, intestinal flora are prone to happen in severe hepatitis patients, which makes intestinal infection very common [ ] . few patients exhibit cellulitis like colitis, which can further result in peritonitis and septicemia, and finally lead to death. pathogens in intestinal infections could be shigella spp., salmonella, campylobacter jejuni, clostridium difficile, and salmonella typhi etc. clinical manifestations are nausea, vomiting, abdominal pain, diarrhea, watery stool or bloody mucopurulent stool. some patients may have fever and tenesmus. depending on the pathogenesis and clinical manifestations, intestinal infections are classified as enterotoxigenic bacterial enteritis and invasive bacterial enteritis. pathogenesis of enterotoxigenic bacterial enteritis is that bacteria adhere but not invade intestinal mucosa. intestinal toxins are secreted in the process of bacteria growth and reproduction, which stimulate small intestinal epithelial cells secreting large amount of water and electrolytes. overuptake of water and electrolyte and retention in intestine makes watery stool, which is called "secretory diarrhea." secretory diarrhea is exhibited as frequent, large amount stool with no pus, usually without pain or tenesmus. it is often accompanied by vomiting, which is prone to bring out dehydration, electrolyte imbalance and acidosis, but less severe systemic toxic symptoms. stool examinations show less erythrocytes or leukocytes. invasive bacterial enteritis refers to pathogenic bacteria adhere and invade the intestinal mucosa and submucosa, causing significant inflammation. different pathogens violate different parts of intestine, small intestine, or colon, and sometimes cause inflammation both of small intestine and colon. the basic clinical manifestations include obvious systemic sepsis, high fever, and even septic shock in severe patients. stool can be mucus bloody or purulent, less amount and more frequency. abdominal pain are often severe, paroxysmal colic. if the lesion invades the rectum and distal colon in particular, there may be tenesmus. sigmoidoscopy examination showed diffuse inflammation and ulceration. if only the small intestine or upper part of colon are invaded, the stool is more moisture, and without tenesmus. stool examination show large amount of leukocytes. although diagnosis of intestinal infection is not difficult, it should be careful to distinguish the site of infection, make sure of pathogens, and pay particular attention to water, electrolyte imbalance and acid-base imbalance. therefore, in addition to routine examinations and stool culture, keeping abreast of the general condition is also important to avoid disturbance of the internal environment which aggravate liver damages. in severe hepatitis, the more severe hepatic damage and immune dysfunction, the higher of incidence of sepsis. bacterium most commonly enters through intestine into the portal vein and then into the systemic circulation, followed by skin, respiratory tract, urinary tract or other intrusion. pathogens are often opportunistic bacteria, in which gram-negative bacteria are more than gram-positive bacteria, especially escherichia coli. clinical manifestations of nosocomial infection concurrent with severe sepsis are not specific, easily masked by the primary disease and complications. in some cases primary focus is not obvious, therefore the diagnosis mainly rely on blood culture. clinical manifestations include: ( ) unexplained sudden chills, fever, shock, increased peripheral blood leukocytes or neutrophils; ( ) deepening jaundice, an increase in ascites, or appearance of hepatic coma, hepatorenal syndrome in a short term. when complications appear in severe hepatitis patients, sepsis should be alert. the mortality of sepsis in severe hepatitis is high. nosocomial infection not only aggravates liver damages, but also induces hepatic coma, hepatorenal syndrome, upper gastrointestinal bleeding and other serious complications, leading to multiple organ failure. nancy rolando reported mortality of septicemia in patients with liver failure was as high as %, in which % with septic shock [ ] . fungal infections can be classified into endogenous and exogenous infection. according to the source of the fungus, the former belongs to opportunistic pathogens, while the latter is in the environment, being infected through various routes of exposure. fungal infections in severe hepatitis are invasive fungal infection in most occasions, and the majority are nosocomial infection and endogenous pathogenic fungi infection [ ] . candida infection is most common, followed by aspergillus, again neoformans and histoplasma monocytogenes. candida albicans is widely present in normal human digestive tract. other fungi such as cryptococcus neoformans, aspergillus are widely found in nature, which can colonize in body surface, or non-enclosed cavity. they also exist in hospital work environments, increasing the chance of nosocomial infection in hospitalized patients. clinically, severe hepatitis with fungal infections are mostly superinfection. the rate of fungal infection in severe hepatitis is increasing in recent years. nancy rolando reported fungal infection was present in of acute severe hepatitis patients ( candida, aspergillus) and in seven cases was considered the major cause of death [ ] . domestic data reported that in a group of patients with liver failure, fungous infection was found in cases in which the rate of nosocomial infections was . %. in separated fungous strains, strains ( . %) were candida albicans, strains ( . %) were aspergillus fumigatus and ( . %) strains were non-candida albicans. the main sites of fungus infection were lungs ( cases) and oral cavity ( cases) [ ] . for severe hepatitis complicated by fungal infection, the mechanism is complex. there are many influencing factors such as immune dysfunction and reduced defensing ability, besides, application and abuse of broad-spectrum antimicrobial drugs are also related [ ] . because of broad-spectrum antibiotics destroy the imbalance between bacteria and fungi in digestive system, which inhibit the growth some gram-negative bacteria that had an anti-fungal effect and some bacteria that are able to synthesize vitamin b family. lack of vitamin b can lead to inhibition of oxidation coenzyme, enabling weakened immunity, which is conducive to fungal growth. research and experience have demonstrated that repeatedly use of glucocorticoid is also an important factor to induce fungal infection in patients with severe hepatitis [ ] . and therefore, the use of corticosteroids also need special care of. currently the use of glucocorticoids in patients with severe hepatitis is still controversial. the majority do not advocate glucocorticoids, or consider a short-term use in the early stages of the disease and be removed as soon as possible. otherwise it will cause a large increase of possibility of fungal infection. fungal infections are among the leading causes of death in patients with severe hepatitis. according to an analysis from rolando, among cases of severe hepatitis, seven cases of deaths directly related to fungal infections [ ] . a recent domestic research analyzed outcomes of patients with severe hepatitis, it was found that the mortality of patients with fungal infection was significantly higher than those without fungal infections ( . % compared to . %) [ ] . according to statistics, fungal infections often occur eight days after admission ( - days) or . days after broad-spectrum antibiotics usage ( - days). symptoms of fungal infection are often covered by severe liver injury related symptoms, while clinical systemic manifestation such as fever is also difficult to identify with the bacterial infection. the site of infection often occurs in mouth, respiratory, digestive or urinary tract. severe immunocompromised persons may appear disseminated infection. oropharyngeal is the site that fungal infections mostly occur, and candida albicans is the most common pathogen, followed by non-candida albicans and aspergillus infection. bedridden patients with severe hepatitis can not properly maintain oral hygiene, which results in change of oral local environmental ph and blood flow. candida albicans easily retains in the mouth, and multiplies, causing oral flora and opportunistic infections. usually general symptoms are mild. patients often have abnormal taste or loss of taste, followed by xerostomia, mucosal burning and other symptoms. candida stomatitis may have pseudomembrane formation which is not easy to peel, accompanied by angular cheilitis, or sometimes manifests as mucosal congestion, erosion or clumps shrink of tongue papillae, thickening of coating on the tongue. there are clear lines between oral pseudomembranous damages. if remove the pseudomembrane it will leave a bright red base, sometimes thick film is like a layer of cheese. take the film directly under microscopic examination shows hyphae and spores. oral fungal infection is often a prelude of deep fungal infection, which should be on the alert. simple oral candida albicans infection is not always accompanied with fever. oral fungal infection is easy to be found, therefore, if oral fungal infection exists with fever and increased leukocytes, it should be paid attention to merger of deep fungal infections such as the lungs or other organs [ ] . aspergillus is ubiquitous in nature which can be spread through air flow. the most common pathogen that causes invasive aspergillosis is aspergillus fumigatus, while aspergillus flavus, aspergillus niger and aspergillus soil are less common. inhalated aspergillus spores can reproduce in healthy or immunity weakened people. respiratory tract is an infective route of invasive aspergillosis, accounted for %. once tissue infection exists, blood vessels violation and bloodstream invasion are common. invasive aspergillosis infection has three characteristics: tissue necrosis, hemorrhage, dissemination [ ] . the mortality of invasive pulmonary aspergillosis accompanied by severe hepatitis is up to % [ ] . it is lack of specific clinical manifestations. in most patients fever is the first arisen symptom, mostly with middle or low heat, sometimes with sudden high fever. hemoptysis occurs with fever simultaneously or - days later, often accompanied with purulent tan sputum or bloody sputum, sometimes with a pinhead size of gray-green particles in it. shortness of breath and chest tightness often exist in late stage of infection, as well as dyspnea, cyanosis, hypoxemia, and expectoration of a lot of bright red bloody sputum or blood clots. pulmonary signs appear later, manifests as pulmonary wet or dry rales, occasional pleural friction sound. in some cases there are no pulmonary signs. x-ray examination of invasive pulmonary aspergillosis showed patchy infiltrates and (or) nodular lesions. typical nodules were like cotton, which can occur in unilateral or bilateral lobes. the lesion progresses rapidly to cause expanded infiltrates, and segmental or lobar consolidation. ct examination showed mass shadow, nodules, or exudative lesions. there are two typical imaging performance: ( ) a characteristic finding on chest ct is the halo sign: a solid nodule surrounded by a halo of ground-glass attenuation [ ] . ( ) the formation of voids, which appear in late infection. signs are hollow cavity lesions, the balloon "crescent" sign was seen around necrosis tissue [ , ] . in order to prevent hepatic encephalopathy, patients can be given high doses of laxatives such as lactulose, which can reduce the intestinal ph value, creating an environment for growth of fungi, thereby increasing the chances of fungal infection of the intestine. in particular, some antibiotics combination increases the incidence of intestinal candidiasis. patients usually have watery or jerry-like diarrhea, with foam or blood. diarrhea is accompanied by bloating, sometimes with vomiting and fever, however, abdominal pain is not obvious. in patients with severe hepatitis, due to decreased intestinal mucosal defense capability, candida may invade the muscle and cause intestinal bleeding, or even perforation. in part of patients, it may even progress to fungal peritonitis, which is similar to clinical manifestations of bacterial peritonitis. in patients with oral candidiasis, if there is dysphagia or pain, especially retrosternal burning, it should be considered that esophagus is invaded. incoordination motor of the upper and lower esophageal can be found by esophageal barium enema. gastroscopy helps to confirm the diagnosis. urinary fungal infection usually involves bladder and kidney, among which candida infection is the most common. however, all pathogenic fungi (such as cryptococcus neoformans, aspergillus species, mucor species, histoplasma, blastomycete, coccidioides) can spread to the urinary system as a systemic or part of disseminated fungal infection, which is more related to usage of broad-spectrum antibiotics and indwelling catheters. clinical manifestations include fever and urinary tract irritation, while some patients were asymptomatic with candidiasis urine. candida and bacterial infections often occur simultaneously. candida infections of the kidney, mostly secondary, are caused by the spread of blood candida. renal cortex and medulla abscesses can occur, which affect renal function in severe cases. patients may have low back pain, abdominal pain, fever and chills, accompanied by urgency, urinary frequency, proteinuria and hematuria. urine tests may find hyphae and fungal spores, culture for candida is positive. candida albicans is common, but now there is a growing trend of candida glabrata [ ] . the main pathogen of fungal sepsis is saccharomyces. most patients have high fever, often above °c. the thermal type varies, of which intermittent fever or remittent fever is more common. wbc count and neutrophil are usually increased. disease in patients with fungal sepsis followed by severe hepatitis is deteriorating rapidly, even progressing to shock. fungal septicemia may invade all the tissues and organs. involved organs have corresponding performance, such as fungal pneumonia, oral fungal infections, intestinal and urinary tract infections. disseminated fungal infection often occurs in severe immunocompromised patients who have long-term use of antimicrobial agents. candida, cryptococcus, aspergillus can disseminate along with blood to all of the organs, such as kidneys, lungs, heart, and liver. the condition is dangerous, which often lead to death in a short term. in addition to common bacterial and fungal infections, severe hepatitis can also be complicated by other pathogens, such as viruses, tuberculosis, protozoa and others. cmv (cytomegalovirus, cmv), herpes simplex virus (herpes simplex virus, hsv), or varicella -zoster virus (varicella-herpes and zoster virus, vzv) infections are three of common herpes viruses infections in severe hepatitis. their common characteristics rely on that once the host is infected, the virus can persist for long periods in the host. when the host immunity is weakened, the virus can re-proliferative, which leads to disease resurgence [ ] . hsv infections manifest as perioral or external genital herpes, oral and esophageal mucosa inflammation and ulcers, also viremia which leads to pneumonia and encephalitis. common clinical symptoms of herpes simplex are mild, only a few people show fatigue, fever and other symptoms. local manifestations are single or multiple blisters on skin or mucous membrane, with tingling. due to reduced immunity, skin rashes in patients with severe hepatitis perform as varicella-like rash, vaccination herpes, herpes keratoconjunctivitis and disseminated herpes simplex. severe herpes usually manifests as herpes simplex virus encephalitis with high mortality. there are fever, headache, mental disorders, coma and other clinical symptoms, often without skin herpes lesions. the sites of infection are commonly in the frontal and temporal lobes. elevated serum antibodies help confirm the diagnosis. cytomegalovirus infections are common in cirrhosis of the liver [ ] . interstitial pneumonia is the most common clinical manifestation in severe hepatitis concurrent with cmv infection. chest ct examinations mainly perform as diffuse interstitial or alveolar infiltrations, very few case show as nodular shadows, occasionally as pleural effusion [ ] . pulmonary consolidation reminds complicated bacterial or fungal infections. pathology manifestations show alveolar interstitial edema, with varying degrees of fibrosis, lymphocyte infiltration and epithelial cell proliferation. blood examination shows leukopenia. because viral pneumonia shares a certain similarity in clinical manifestations, diagnosis mainly depends on pathologic examination. pathogenic examinations for cmv often use methods below: ( ) detect of cmv inclusion body cells and viral particles: eosinophilic nuclear inclusions giant cells are found in respiratory secretions and bronchoscopy lung biopsy specimens. respiratory secretions, saliva, urine, cervical secretions, liver or lung biopsy specimens were inoculated to human embryonic fibroblast cell culture medium, where cytomegalovirus was separated. ( ) immunological methods: cmv antigen from secretions was detected by fluorescent antibody assay, or elisa, which is conducive to the early diagnosis. serum antibodies can also be detected by complement combined experiment, in which acute and convalescent serum antibody titer more than times are positive. ( ) molecular biology methods: pcr technology and nucleic acid hybridization, which helps to make distinction between a variety of different subtypes of the virus. most of vzv infections in patients with severe hepatitis are due to latent virus reactivated. in patients who have had chickenpox, there is a small amount of virus lurking in the spinal cord dorsal root ganglia or cranial nerve sensory ganglia. when severe hepatitis happened, latent virus is reactivated in ganglia due to decreased immunity. the activated viruses spread along with sensory nerve axons to downstream disposal areas, proliferate and cause shingles. in early time, there is paresthesia, itching, and pain in local skin. and then rashes and herpes break out, chaining into a strip, which distribute in denomination or trunk, unilateral, with duration of about three weeks or several months. part of patients with severe hepatitis show severe disseminated herpes zoster. varicella-like rashes appear in a few days, often accompanied by fever, which may be complicated by lung, brain damage with a high mortality rate. latent tuberculosis can burst to tuberculosis and extrapulmonary tuberculosis when cellular immune function gets weakened. under normal host immune function, lymphocytes, macrophages and common langerhans cell may promote granuloma formation and make infection localized. when host immunity is dysfunctional, tissue reaction is very small or even disappeared, leading to mycobacterium growing rather than formation of granulomas, nor any effective defense against infection. severe hepatitis patients with m. tuberculosis infection may develop acute miliary tuberculosis that manifest as fever, cough, expectoration, bloody sputum, chest pain, and shortness of breath in addition to deteriorated liver function. moreover, tuberculosis easily spread in patients with severe hepatitis, with poor anti-tuberculosis efficacy. common extrapulmonary tuberculosis can be lymphatic tuberculosis, intestinal tuberculosis, bone tuberculosis, renal tuberculosis, epididymal tuberculosis, genitourinary tuberculosis, nervous system tuberculosis, tuberculous meningitis [ ] . in patients with severe hepatitis concurrent with tuberculous mycobacterial infections, tuberculin reaction is negative in about half of patients, especially in those applied with glucocorticoid. diagnosis relies on sputum acid-fast staining or pcr, but the diagnostic yield is not high. interferon gamma release assays (igras), including quantiferon ® -tb gold in-tube, and the t-spot tb, have been extensively used for the auxiliary diagnosis of tuberculosis infection in adults. igras detect circulating t-cells responsive to specific mycobacterium tuberculosis antigens, which are absent in bcg and other non-tuberculosis mycobacteria, and exhibited similar sensitivity and higher specificity than tst in adults [ , ] . however, these igra tests are also affected by host immune status [ ] . in addition, the decision regarding whether to treat ltbi should be dependent not only on igras results but also on clinical histories. ntm generally causes local wound infections. however, in severe hepatitis patients with impaired immune function, non-tuberculous mycobacteria can invade the lungs, causing tuberculosis-like diseases, but rarely causes hematogenous dissemination. histological examination of the lesion is mainly characterized by epithelioid cell granulomas and foam cell-like balls of tissue proliferation, detection of non-tuberculous mycobacteria [ ] . protozoa and worms, such as toxoplasma gondii, giardia lamblia, cryptosporidium and stercoralis, can also infect patients with severe hepatitis, especially those with immunosuppression drugs or combined with cancer. main lesions of toxoplasmosis manifest as lymphadenopathy, hepatosplenomegaly, encephalitis and pneumonia [ ] . clinical manifestations of giardia lamblia infection are chronic diarrhea and malabsorption, also fever and cholecystitis [ ] . pathological changes are deformation of small intestine villi and lymphoid hyperplasia. parasites present in small intestinal surface and gallbladder, and the detection of parasites from the stool and duodenal drainage fluid that is eligible confirmed. strongyloides stercoralis is a weak virulent worm, there is few clinical stercoralis infection [ ] . but this worm infection in patients with severe hepatitis is a serious threat, even causing death. clinical manifestations include long-term nausea, vomiting, diarrhea, bloating, intestinal paralysis, dehydration, electrolyte imbalance, edema, weight loss and difficulty breathing in cases with extensive lung lesions. all patients have hypoproteinemia and anemia. patients with increased eosinophils have good prognosis, whereas eosinophils reduction often is a dangerous signal. varieties of complications, such as system or local infections, coagulopathy, hepatic encephalopathy, hepatorenal syndrome, hepatopulmonary syndrome, acid-base imbalance and water-electrolyte imbalance, are main causes of hbv-associated mortality during deterioration of liver function of aechb. reducing and effective treatments of these complications are still nodules in therapy of severe phase of aechb. infection is one of usual complications of aechb, which shows - % morbidity in civil china. the most common localities are respiratory system, especially lungs, and abdomen, which shows the incidence of spontaneous bacterial peritonitis (sbp) as much as %. others include urinary tract and bloodstream. gram-negative bacteria are the predominant causes including aerobic gram-negative bacilli, escherichia coli, klebsiella, enterococcus and anaerobic bacteroides fragilis, etc. however in recent years, gram-positive bacterial infections are found increasing in patients with aechb, including pneumococcus and other streptococcus, but staphylococcus aureus infection is relatively infrequent. fungal infections are usually secondary to bacterial infection. candida, aspergillus, sporotrichosis, histoplasmosis and coccidioidomycosis infection, especially candida albicans, are most frequent in occurrence. but, infections of cryptococcus and mucormycosis are relatively rare. beyond of bacterial and fungal infection, other infectious pathogens in aechb are virus, mycobacterium tuberculosis, mycoplasma, chlamydia, parasites and protozoa. bacterial infection is the most frequent complication in aechb. infections are more likely occurred in abdomen, including abdominal cavity, biliary tract, gastrointestinal tract, or other parts like respiratory tract, and urinary tract. keeping oral and perineal asepsis, unobstructed respiratory tract and urine tract, anti-bedsore care for patients with limited mobility, rational use of antibiotics, avoiding long-term use of broad-spectrum antibiotics, strict controlling usage of glucocorticoid, aseptic practices in invasive operation, parenteral nutrition and protecting the intestinal mucosa are all necessities to prevention of bacterial infection in aechb. empirical use of antimicrobial agents could be determined by the localities of infection without antibiotic susceptibility testing results. gram-negative infections are more frequent in peritoneal or biliary tract, in which cephalosporin or quinolone could be the first choice. penicillin and vancomycin could be considered in pulmonary infection. azithromycin and quinolone could be adopted in urinary tract infection. metronidazole or tinidazole could be used in anaerobic infection. broad-spectrum antibiotics can be used in serious infection, such as ceftriaxone, cefoperazone, cefepime, and carbapenems, but the secondary infections need to be highly concerned. initial antibiotics should be adjusted according to antibiotic susceptibility testing results as soon as possible. non-specific immune enhancer agents, such as thymosin, are well used in treatment of infection during aechb, but it currently still lack of evidence-based support. the incidence of fungal infection ranks secondly in aechb associated infection. respiratory tract, gastrointestinal tracts and genitourinary system are the major sites. keeping wards dry and ventilated can help to reduce environmental fungal growth. oral and perineal cleaning and regularly dental check are necessary. if oral fungal spots are found, alkaline mouthwash and nystatin with glycerol can be used in treatment. for patients with limited mobility, anti-bedsore care is terribly necessary. rational use of antibiotics, especially avoiding long-term usage of broadspectrum antibiotics, can prevent secondary fungal infection. glucocorticoid should be used strictly according to indication. during invasive procedures, aseptic operations are obligated. regular check of sputum, lungs, urine and feces will facilitate early diagnosis. empiric antifungal treatment is generally not recommended, but to patients with aechb with aids, especially with count of peripheral blood cd + t cell-less than /μl or oropharyngeal candidiasis found, the sulfamethoxazole (smz-tmp) should be chosen to prevent pneumocystis pneumonia. fluconazole should be considered when moderate amount of candida albicans has been indicated by sputum culture. empirical treatment. if candida albicans infection is highly susceptible, the initial treatment choice is fluconazole ( - mg/days). but if aspergillus infection is preferred, amphotericin b liposome, itraconazole, caspofungin or voriconazole could be considered as the initial treatment, in which process liver and kidney function should be intensively monitored. to patients with cryptococcal encephalopathy combining severe liver damage, fluconazole or flucytosine combined with intrathecal injection of amphotericin b treatment could be implemented. evidence based treatment. initial antifungal strategy should be adjusted according to antibiotic susceptibility testing results. for certain invasive pulmonary aspergillosis, combination treatment of several different types of antifungal agents should be considered under monitoring of liver and kidney function. however, for pulmonary mucormycosis infection, the combination of amphotericin b and flucytosine is the only effective strategy. amphotericin b is a type of polyene antifungals, mainly for aspergillus, candida, cryptococcus, histoplasma infection, but is invalid in aspergillus terreus or ringworm fungus infection. intravenous or intrathecal injection of amphotericin b should be administrated because of non-digestive absorbance. the recommended initial dose of intravenous administration is - mg/days, and gradually increases to . - mg/kg.days. the infusion track needs to be dark and process needs not less than h. the initial dose of intrathecal injection is . mg/days and gradually increases to . - mg/days. amphotericin b has an extra toxicity to liver and kidney, and also causing hypokalemia. intensive monitoring of liver and kidney function and serum potassium levels is necessary during treatment, and be sure largely avoiding combination with other agents with liver and kidney toxicity. itraconazole is one of triazole antifungal agents, mainly for aspergillus, candida, cryptococcus and histoplasma infection, but it is invalid in fungi fusarium or zygomycetes infection. the intravenous dose for the initial days is mg/days, administrated by twice, and then followed by mg/days for weeks. the oral doses of mg bid could be subsequent. the total curative course could be determined by the improvement of symptoms and largely absorption of radiographic lesions of infection. the monitoring of liver function is necessary and recommended, especially in long duration treatment. furthermore, combination treatment with other hepatotoxic drugs should be avoided. fluorocytosine is one of bacteriostatics, mainly for cryptococcosis and candida infection, frequently based on the combination with amphotericin b. the daily dose for adults is . g with intravenous dripping speed of - ml/min. a half dose should be conducted in renal insufficiency. the inhibited administration includes severe liver or kidney dysfunction and allergy to fluorocytosine, and also cautious treatment for pregnant or breastfeeding women. no combination treatment of fluorocytosine with cytarabine or bone marrow suppression agents is recommended. fluconazole is a triazole antifungal, mainly for candida albicans or cryptococcus infection, but not as good in candida glabrata infection, totally invalid in aspergillus or candida krusei infection. the recommended daily dose for adult is - mg, and the initial dose should be doubled. voriconazole belongs to triazole antifungals, mainly for candida, cryptococcus, aspergillus, fusarium and histoplasma capsulatum infection, especially used for invasive aspergillosis and invasive fluconazole-resistant candida infection, but is invalid for mucor or rhizopus infection. the recommended intravenous initial dose in adult is mg/kg, q h, with infusion rate of mg/kg within - h. the maintenance dose from the second day is mg/kg, q h. to patients without tolerance to normal dose, it could be reduced to mg/kg, q h. the reduction of daily dose could not be necessary in mild to moderate liver dysfunction. but intravenous administration should be avoided in patients with severe renal dysfunction. the side effects of voriconazole include transient visual disturbances, mental disorders, thrombocytopenia and so on. caspofungin belongs to echinocandin antifungals with antibiotic spectrum of pathogenic aspergillus, candida and pneumocystis, without in cryptococcus neoformans, fusarium and mucor. it is mainly used for invasive aspergillosis. the initial dose for adults is mgqd, and with subsequent mg qd. the infusion time is no less than h. no fixation curative course is suggested. caspofungin should be avoided for patients with severe liver function, because of highly hepatic distribution and metabolic pathways during treatment. liver is the largest solid organ in the body, and it plays a central role in the clotting process, except for general function such as metabolism, detoxification and choleresis [ ] . a majority of the coagulation factors are synthesized almost exclusively in the liver. in the pathogenesis of severe hepatitis, massive hepatic necrosis leading to reduced production and dysfunction of coagulation factor. in addition, the increased levels of anticoagulation and platelet abnormalities also contribute to occurrence of coagulopathy, which were further exacerbated by severe complications such as spontaneous bacterial peritonitis (sbp) and hepatorenal syndrome (hrs). coagulation abnormalities, even disseminated intravascular coagulation (dic) often occur in those patients with severe hepatitis. therefore, the changes in coagulation factors were usually used to evaluate prognosis of severe hepatitis [ ] . the normal procedure of coagulation includes two independent coagulation process, namely intrinsic and extrinsic pathway, which initiated by coagulation factor xii, and factor viia/tissue factor, respectively. the two pathways reach a confluence at the points of factor xa and va. wherein factor xa activates prothrombin to thrombin in the presence of ca + and factor va bound to membrane surfaces, and then thrombin converts fibrinogen to fibrin. thus, blood becomes clotted and the "y" shaped pathway was established [ ] . simultaneously, there are some anticoagulation systems existing in our body, which prevents the formation of thrombus, then keeps normal circulating of blood flow. it also participates in maintaining of normal permeability of the blood vessel [ ] . among anticoagulation systems, the most important one is the fibrinolytic system, whose basic process can divided into two stages, i.e. plasminogen activation and fibrin degradation [ ] . plasminogen activators include tissue-type plasminogen activator (tpa) and urokinase-type plasminogen activator (upa), and the principal function of these two plasminogen activators is to convert plasminogen to plasmin, then plasmin cleaves fibrin into soluble small peptides, namely fibrin degradation products [ ] . moreover, the process of fibrinolysis was affected by fibrinolysis inhibitors, such as plasminogen activator inhibitor (pai) and alpha antiplasmin (α -ap). those fibrinolysis inhibitors can either inhibit plasminogen activation, or reduce the function of plasmin [ ] . thus, there is a dynamic balance between coagulation and anti-coagulation systems under physiological conditions ( fig. . ) [ ] . many factors involved in coagulation process, i.e. clotting factors, thrombin inhibitor, fibrinolytic system and so on, are synthesized almost exclusively in the liver. thus, liver plays a pivotal role in remaining the balance between hemorrhage and coagulation under physiological conditions (table. . ) [ ] . it is the important basis for pathogenesis of coagulopathy that massive hepatocyte necrosis occurs in severe hepatitis patients, resulting in reduced production and dysfunction of those blood clotting factors [ ] . coagulation abnormalities, such as decreased production and dysfunction of coagulation factors, are commonly found in severe hepatitis, there are at least types of factors participated in the coagulation process, including classical coagulation factors, namely factor i/fibrinogen, factor ii/prothrombin, factor iii/tissue factor, factor iv/calcium ions, as well as factor v, vii, viii, ix, x, xi, xii, and factor xiii. some bradykinin factors, such as rk (prekallikrein) and hmwk (high molecular weight kininogen) are also associated with coagulation. among above factors, the others belong to proteins excepting factor iv. plasma coagulation factors are synthesized exclusively in the liver, excepting factor iii/tissue factor, factor iv, factor vi/activated factor v, factor viii and factor viiia [ ] . massive hepatic necrosis leads to the decreased production of clotting factor in patients with severe chronic hepatitis. moreover, the coagulation factors are sensitive indicators for clinical evaluation of severe hepatitis. it is common that the serum levels of factor v, vii, ix, x, xi and prothrombin decreased in the patients with severe hepatitis. on the contrary, clotting factor viii is synthesized, and then excreted increasingly by the mononuclear phagocytic cells with stimulation of various inflammatory cytokines in the patients with severe hepatitis [ ] . anorexia and antibiotics overuse lead to obstacles in assimilation, absorption, and application of vitamin k in these patients. vitamin k dependent coagulation factors includes factor ii, vii, ix, and x. the function of γ-hydroxylation was weaken by the deficiency of vitamin k and damage of hydroxylase, then the abnormal clotting factors without γ-hydroxy glutamate were synthesized. finally, these abnormal clotting factors lead to dysfunction of the vitamin k dependent coagulation factors [ ] . coagulation factor vii eliminated significantly in the initial stage of liver injury was usually used as an early diagnosis index [ ] . in addition, coagulation factors ii and x, then factor ix decreased markedly, along with exacerbation of liver injury. the deficiency of vitamin k induced by obstructive jaundice, malabsorption syndrome, can be rescued after intravenous injection of vitamin k, which is different from coagulopathy caused by liver injury. the plasma level of fibrinogen is within normal range in patients with compensatory cirrhosis. however, patients with severe hepatitis or liver failure have a significant reduction in the level of fibrinogen, and then develop dysfibrinogenemia [ ] . fibrinogen, as the main hydrolysis substrate of thrombin, is the crucial factor in the coagulation process. thus, decreased levels or abnormal structure of fibrinogen leads to coagulation abnormalities in patients with severe hepatitis [ ] . there are two clotting factors associated with thrombin except for fibrinogen, i.e. factor v and factor xiii. factor xiii induce a crosslink of sfm (soluble fibrin monomer), resulting in the formation of insoluble fibrin [ ] . it reports that the levels of circulating coagulation factor xiii were decreased in % of patients with liver disease, resulting in fibrin decline or abnormality. these patients with liver disease will receive a bad prognosis if the plasma level of fibrin was at level below % of normal concentration [ ] . it is uncommon that the plasma level of plasma factor v decreased markedly in patients with severe hepatitis, except for those complicated with dic or hyperfibrinolysis. the low level of factor v cannot induce the formation of the enzyme-substrate complex, which delay the activation of prothrombin [ ] . contact factors include classical coagulation factor xii and xi, as well as some bradykinin factors, i.e. pk and hmwk. these factors contact with vascular intima damage or abnormal surface in blood vessel walls, then active the intrinsic coagulation pathway [ ] . moreover, the contact factors can connect with bradykinin, fibrinolysis and complement system. in addition, coagulation factor xiii also activates fibrinolysis system as the initiating factor. factor xii deficiency does not cause excess bleeding, but induce thrombus due to its decreasing activation of fibrinolytic system [ ] . there is a dynamic balance between pro-and anti-coagulation systems in physiological conditions, which keep normal blood circulation in the body. the anticoagulation system includes physiological anti-coagulation factors (e.g. antiprothrombin-iii, protein c, and so on) and fibrinolysis factors (e.g. plasminogen, α- plasmin inhibitor, and so on) [ ] . at-iii, with a half-life period of . days, synthesized mainly in hepatocytes and partly in endotheliocyte, is responsible for about % of anticoagulation in plasma. it is the main reason for low plasma level of at-iii that decreased production and increased consumption of at-iii caused by hepatocytes necrosis in patients with severe hepatitis. the activity of at-iii obviously decreased in severe hepatitis [ ] . therefore, heparin, thrombin, activated coagulation factors (e.g. factor x, ix, xi, xii) cannot be inhibited, due to rare at-iii combining with them [ ] . there is a negative correlation between at-iii: a (the activity of at-iii) and pt (prothrombin time), which indicate that the level of at-iii: a decreased obviously along with exacerbation of liver injury [ ] . the plasma levels of pro-and anti-coagulant factors are low in patients with liver injury. when the necrotic tissue and cytolysis are released in the blood, the balance of pro-and anti-coagulant is destroyed. finally the depletion of at-iii by massive activated coagulation factors will lead to dic [ ] . massive hepatocytes necrosis, vitamin k deficiency, and protein c without γ-hydroxyglutamic acid result in blocking activation of protein c. thus, va, viiia and pai cannot be degraded, and the plasma level of them will rise up, due to reduction of activated protein c (apc) in the patients with severe hepatitis [ ] . plasminogen synthesis will decreased by about % in the liver when sever hepatitis occurs. as the activators of plasminogen, tpa and upa are synthesized by vascular endothelial cells, and their production will increase after vascular endothelial cells initiated with virus, immunocomplex or endotoxin in the patients with severe hepatitis b [ ] . with exacerbation of liver injury, pai synthesis decreased significantly in the liver. the main physiological activity of pai is to inhibit tpa induced plasminogen activation. therefore, the activity of tpa will increase with reduction of pai synthesis, resulting in promoting the conversion of plasminogen into plasmin [ ] . plasmin, as a kind of powerful proteolytic enzyme, can hydrolyse fibrinogen into fdps, degrade coagulation factors, and inhibit platelet aggregation, resulting in the aggravation of bleeding [ ] . hyperfibrinolysis can be caused by congenital or acquired reason, and it commonly leads to a rapid depletion of coagulation and anticoagulation factors, especially in those patients with dic [ ] . synthesis and secretion of tpa and upa markedly increase, while pai, namely tpa or upa inhibitor, has been a decrease in the plasma of severe hepatitis patients, resulting in hyperfibrinolysis [ ] . at the same time, mononuclear phagocyte system cannot degrade plasminogen activator, also leading to hyperfibrinolysis. it is not necessarily accompanied by hemorrhagic tendency, although there are many risk factors that can cause hyperfibrinolysis, even hyperfibrinolysis occurred in severe hepatitis [ ] . in addition, fdps inhibit fibrin monomer polymerize function, and also block platelet aggregation, then further worse the deficiency of hemostasis and coagulation, finally leading to the aggravation of bleeding tendency [ ] . studies have shown that low-level endogenous small molecule heparin in patients with chronic hepatitis may be associated with many factors, such as increased mastocyte, decreased production of heparinase in the liver and reduced activity of ph (platelet factor ) [ ] . when the disease progress to cirrhosis or chronic severe hepatitis, pt was significantly prolonged, while endogenous heparin wasn't increased markedly, indicating that low-level endogenous heparin has little effect on pt elongation and hemorrhagic tendency [ ] . however, if the patient is undergoing the following condition together, the endogenous small molecule heparin will increase significantly. ( ) esophageal variceal bleeding with serious infections (such as abdominal infection, biliary tract infection and pulmonary infection) or portal hypertension. ( ) combining with the significant increased white blood cell count. ( ) percentage of neutrophils > %. the level of endogenous small molecule heparin in plasmin will decrease after those infections were cured. taken together, these data indicate that increased level of endogenous small molecule heparin in blood circulation was closely related to severe hemorrhagic tendency when combining with infection [ , ] . platelet significantly promotes blood coagulation through connecting with many coagulation factors (such as fibrinogen, factor v, factor xi, factor xiii and so on). α-granule includes fibrinogen, factor xiii, and some platelet factor such as platelet factor (pf ) and platelet factor (pf ), which promote coagulation activation process of factor xii and factor xi can be accelerated by activated platelets [ ] . it is estimated that pf provided by platelets could accelerate activating of thrombin by twenty thousand times. xa and factor v could not be inhibited by at-iii and heparin if they were linked with pf . when platelets aggregated and formed platelet plugs, the process of coagulation was initiated at this site, and then platelets reveal a large number of phospholipid surfaces, which were helpful for activating of factor x and fibrinogen [ ] . various platelet factors were released from α-granule after platelet aggregating, and then hemostyptic fibre was produced increasingly. those hemostyptic fibre finally produced blood clot formation after capturing the other hemocytes. thus, platelet plugs would progress independently of platelet disintegrating gradually [ ] . two aspects of platelet abnormalities consist of depletion and dysfunction. in patients with chronic hepatitis and cirrhosis, the occurrence of decreased platelet level is usual through hypersplenism accompanied with other hemocyte decreasing [ ] . the ratio of which reaches - %. its incidence rate in severe hepatitis and explosive hepatic failure also reaches approximately by %. when severe hepatitis occur, myelosuppression, decreased megacaryocyte replication leading short-life platelet, lacking of hemopoietic material such as vitamin b, folate and so on, all of these can initiate thrombocytopenia [ ] . other reasons leading to thrombocytopenia contain low expression and metabolic disturbance of thrombopoietin (tpo), as well as platelet antibody production. researches show that the serum level of tpo related positively to platelet expression [ ] . we next studied platelet associated immunoglobulin (paig) and its sorts such as paigg, paiga, paigm etc., with chronic hepatitis. in line with previous reports, we found that serum levels of paig and paigg negatively correlated with blood platelet count, corroborating the crucial role of the paig-mediated autoimmunization in controlling thrombocytopenia in viral hepatitis [ ] . various factors impact platelet function forwardly or passively. increased expression of oxide and prostacyclin, two kinds of platelet inhibitor derived by endothelium, may inhibit platelet activation in vivo [ ] . on the other hand, increased serum level of von willebrand factor (vwf) can promote platelet adhering and aggregation in patients with cirrhosis. when severe hepatitis occur, . % and . % of patients have decreased levels of platelet adhesion rate and platelet aggregation rate (par), respectively. in addition, reduced effectiveness of pf and abnormal clot contraction occur in patients with severe hepatitis by an incidence of . % and . %, severally [ ] . patients with terminal liver disease significantly exert hemorrhagic tendency, especially in the digestive tract [ ] . the latest report indicated that basic laboratory examinations for coagulation function testing in common use at present, such as pt, aptt, international normalized ratio (inr) etc., have little correlation with occurrence of gastrointestinal bleeding in these patients, thereby revealing the importance to search and pay close attention to those complicating disease upregulating bleeding risk, such as bacterial infection, renal failure, hemodynamic change after portal hypertension, dysfunction of endotheliocyte as well as macrophagocyte and so on [ ] . it is common to see renal failure occurrence in advanced hepatopathy. when it happened, acquired platelet dysfunction, abnormal activation of platelet and vascular wall, anemia and so on, all of them significantly promote hemorrhage [ ] . as another severe complication, bacterial infection is also very important and common [ ] . when tumor necrosis factor (tnf) were injected into healthy individuals, we found that endotoxin have an important role to exert its function directly in clotting cascade reaction. early researches indicated that the body can express endogenous heparin-like substance through stimulation by endotoxin in patients with cirrhosis [ ] . furthermore, some studies revealed the relevance of endotoxin with prothrombin fragment, indicating that infection promoted occurrence of dic-like status in laboratory examination in cirrhosis [ ] . coagulation system became weaker in patients with chronic hepatitis. it can hardly mediate factors due to the relative deficiency of pro-and anticoagulation factors. when the balance was destroyed, it may tend to hemorrhage or thrombosis depending on related risk factors which were in the ascendant. to assess abnormality of blood coagulation in patients with liver disease, we should be in consideration of hypercoagulability, one state may easily be overlooked. otherwise, it will be unfair. current literatures indicated that, unlike previous concept that the body of patients initiated anticoagulation state automatically when infected by the hepatitis virus. various clinical evidences revealed that hepatitis virus infecting cannot inhibit thrombus forming. furthermore, it may increase dangerous to thrombus forming especially in the portal system, for existence of individuals having hereditary mutation to promote thrombus forming [ ] . slower bloodstream, abnormal fibrinolysis initiated by blood stasis, and decreased activity of anticoagulant accelerate formation of venous thrombosis. moreover, the changes of platelet phospholipids membrane activity were also helpful to the formation of thrombosis in patients with chronic liver disease [ ] . it is reported that the incidence of periphery deep venous thrombosis and pulmonary embolism was . and . % in patients with cirrhosis, respectively [ ] . the rate of portal vein thrombosis is about % in patients with compensated cirrhosis, but it reaches - % in the candidates waiting for liver transplantation [ ] . for mutations (such as leiden mutation of factor v, g a mutation of prothrombin, c mutation of methylenetetrahydrofolate reductase) or the existence of antiphospholipid antibody syndrome, the hypercoagulable state will be represented in patients with liver disease [ ] . the hypercoagulable state represents diverse modes in the body of patients with chronic hepatitis. among them, thrombosis is more traditional and common. the mechanism of dic complicated with severe hepatitis may contain several aspects as follows. . with attacks from endotoxin, virus and immune complex etc., endotheliocyte was injured, and then it activated plasmakinin system and complement system, leading to the aggravation and participation of dic progress. damaged endotheliocyte can simultaneously activate factor vii, intrinsic coagulation pathway and platelets, and participate in micro-thrombosis with platelets adhesion and aggregation beneath endothelium [ , ] . . in patients with severe hepatitis, massive necrotic hepatocytes activated extrinsic coagulation system with the releasing of various tissue thromboplastin-like substances [ ] . . hepatocellular necrosis or dysfunction decreased expression of anti-coagulation factors such as at-iii, pc, protein s and so on, and then it enhanced activation of thrombin and plasmin [ , ] . . impaired function of the mononuclear phagocyte system. mononuclear phagocytes can express activated tissue factor with the releasing of tnf, il- and platelet activating factor (paf) on its surface after stimulation by endotoxin, inflammatory cytokines and complement activation. tnf and il- can decrease the activation of protein c through its function to increase the expression of plasminogen activator and plasminogen activator inhibitor- and to inhibit the production of thrombomodulin (tm) [ ] . activated clotting factor and other factors with promoting coagulation can lead to the occurrence and aggravation of dic, since it cannot be promptly removed [ ] . . the onset and enhancement of fibrinolysis. massive clotting factors and platelets were depleted in the extensive formation in vivo microthrombus. thrombin promoted the conversion of fibrinogen into fibrous protein [ ] . simultaneously, it activated fragments which dropped in the formation of activated factor xiii, factor xa and factor xiia. all of them can activate plasmin, and it enhanced fibrinolysis with tpa, one factor released by damaged blood vessel endothelium [ ] . fibrinogen and fibrous protein were degraded after plasminogen activation to generate the corresponding fdps, one factor inhibit blood coagulation and also block platelet aggregation [ ] . taken together, the above process exacerbates bleeding initiated by coagulation factors depletion and platelet deficiency. coagulopathy, characterized by prolongation of blood clotting time, tendency of hemorrhage, or even dic, were commonly found in severe hepatitis patients. it may cause uncontrolled external or internal hemorrhage. as common clinical symptoms, the external hemorrhages include gingivale or nasal mucosal bleeding, skin petechiae, the punctures or injection-site ecchymosis, and so on. the internal hemorrhages include esophagogastric varices, intracranial, subcutaneous, and muscle interval bleeding. except for esophagogastric varices hemorrhage, the other internal hemorrhages rarely occur in those patients. in addition, massive hemorrhage from esophagogastric varices is a serious medical emergency, can potentially cause death and cardiac arrest if proper medical treatment is not received quickly [ ] . when dic occur, a wide range of thrombogenesis in microvasculature can cause circulatory collapse, characterized by low blood pressure and shock. microcirculatory dysfunction occur after microthrombosis producing, resulting in hypofunction of multiple organs (e.g. kidney, liver, lung and pancreas), which perform from dysfunction to failure, with illness progressing [ ] . crushed by the fibrous protein in the vessels, red blood cell destruction can lead to intravascular hemolysis. at the early stage of disease, it shows hypercoagulability. the blood in the needle is easy to coagulate, when getting blood sampling from the vein. after that, it comes with the stage of consumed hypocoagulation. the consuming of a great number of blood coagulation factors leads to significant tendency of hemorrhage [ ] . it is difficult to stop bleeding in many parts of the body, including visceral organs, operative sites, injection sites, puncture sites, and mini-invasiva sites [ ] . when the third stage, namely secondary fibrinolytic stage comes, a great number of the blood coagulation factors have already been used up, resulting in severe bleeding under the condition of low coagulation. shock, acidosis and mods make the patient's condition continue deteriorating, and are also the main reasons for death [ ] . the latest study indicates that thrombosis is also a noticeable state in cirrhosis and end-stage liver disease. portal thrombosis and peripheral vein thrombosis (e.g. deep vein thrombosis and lung embolism) are commonly seen. the deep vein thrombosis is more danger than lung embolism. anyway, the incidence rate of thrombosis in cirrhosis and end-stage liver disease is still very low. the clinical symptoms depend on the embolizing position after the thrombosis occurs [ ] . presently blood clotting factors test is the most maturely and frequently used test in the term of blood coagulation function in the world. the indicators including prothrombin time (pt), international normalized ratio (inr) and prothrombin time activity (pta) have always been chosen in the patients with severe hepatitis. prothrombin time (pt) reflects whether there are anticoagulant substances in the extrinsic blood coagulation system and blood circulation or not. the elongation of prothrombin time (pt) presents the declined activity of several blood clotting factors including factorii (fii), factorv (fv), factor vii (fvii), factorx (fx) or the existence of anticoagulant substances. in severe hepatitis patients, the incidence rate of prothrombin time (pt) elongated can reaches to %, thus it is regarded as a sensitive and frequently used indicator in the term of liver function [ ] . another index international normalized ratio (inr), a reckoned ratio calculated from prothrombin time (pt) and international sensitivity index (isi), making prothrombin time (pt) between different laboratories and different reagents comparable, is an international general indicator. the guides about acute-on-chronic liver failure and acute liver failure take the index, inr ≥ . , as one of the most significant diagnostic criteria in the american association for the study of liver diseases (aasld) and the asian pacific association for the study of liver (apasl). international normalized ratio (inr) can also be used as a monitor on blood coagulation function [ ] . hepaplastin test (hpt), reflecting not only blood coagulation mechanism in hepatitis patients but also the function of hepatocytes to synthetize vitamin k dependent clotting factors including factorii (fii), factor vii (fvii), factorix (fix), factorx (fx) synthetically, is a test about liver reserve function; but this indicator can't reflect the change of factorv (fv). when severe hepatitis occur, the function of liver to synthetize above-mentioned clotting factors declines and the time of hepaplastin test (hpt) elongates. with illness progressing, hpt continues elongating. survivors undergoing effective therapies can have gradual recovery in the time of hepaplastin test (hpt). so this test is regarded as the specific test of liver diseases or the optimal indicator reflecting liver reserve function [ ] . the severity of liver damage is positively correlated with the decline degree of prothrombin time activity (pta): the more severe damage occurs in hepatocytes, the more significantly prothrombin time activity (pta) will decline. therefore prothrombin time activity (pta) < % and total bilirubin (tbil)> μmol/l have always been used as the main laboratory indicators to diagnose severe hepatitis domestically. pta < % is also regarded as diagnostic criterion of blood coagulation dysfunction in the guideline of acute-on-chronic liver failure in the asian pacific association for the study of liver (apasl) [ ] . in severe hepatitis, total bilirubin (tbil), total cholesterol, prothrombin time activity (pta) and complications (e.g. rectory hyponatremia, hepatic encephalopathy, hepatorenal syndromes and so on) are all independent risk factors to evaluate prognosis. the lack of any factors including factori (fi), factorii (fii), factorv (fv), factorvii (fvii), factor x (fx), can lead to the decline of prothrombin time activity (pta). moreover the half-life time of those factors are extremely short, factorii (fii) - h, factorv (fv) - h, factorv (fv) - h, factorv (fv) - h, respectively. it means that when hepatocytes suffer from severe serious damage and necrosis in severe hepatitis, prothrombin time activity (pta) will have dramatic decline just in a few days. as a result, prothrombin time activity (pta), characterized by significant advantages in evaluating patients condition and judging prognosis over other laboratory indicators, has been widely used. elongation of aptt prompts the lack of any clotting factor belonging to intrinsic coagulation system or the existence of anticoagulant substances. the incidence of the elongation of aptt reaches to - % in severe hepatitis patients [ ] . fxii:c reflects liver synthetic function. fvii, characterized by the shortest halftime - h, is the first one to be affected when facing liver synthetic dysfunction. on the contrary, fv:c, characterized by a relatively long half-time, is one of the latest factors to be affected and is correlated with the degree of liver damage. it prompts severe liver failure, bad prognosis and even easy death when plasma levels of fv:c under %. some literatures report that fv:c and pta can be used as significant prognostic factors in liver failure and significant screening indicators in liver transplantation. however, the indicators-fxii:c and fv:c-are not listed as routine examination items. they are just selected on the condition of illness demand [ , ] . the main test about anti-clotting factors is the determination of antithrombin iii activity (at-iii:a). in the state of pathosis, the decline of antithrombin iii activity (at-iii:a) is not parallel with the decline of antithrombin iii (at-iii) content, namely the depletion of antithrombin iii activity (at-iii:a) more apparent. owe to this, it has more clinical value to determine antithrombin iii activity (at-iii:a) rather than antithrombin iii (at-iii) content. moreover, anti-clotting factors tests include the determination of protein c activity (pc:a) as well [ ] . serum levels of fdps are very low in normal people. significantly elevating of fdps indicates the existence of hyperfibrinolysis and reflects the occurring of dic indirectly. there are many assay methods including immunization fi test (namely latex particle agglutination test, normal titer< : ), fdps flocculation test, radial immunodiffusion staphylococcal clumping test, indirect hemagglutination inhibition test, enzyme-linked immunosorbent assay (elisa) and so on. if the serum levels of fdps elevate, it indicates acute dic may occur [ ] . plasma protamine paracoagulation test ( p test) and ethanol gel test (egt) reflect the soluble fibrin complexes in plasma. soluble fibrin complexes, combination of fdps and fibrin monomer, can't be solidified by thrombase. but protamine is able to make the complexes isolate and then fibrin monomers separate out again. the paracoagulation test means self-polymerization between fibrin monomers and fdps, then forming macroscopic flocks. ethanol gel test (egt) has the same principle with plasma protamine paracoagulation test ( p test), whereas the former has a lower positive rate. the two methods may have false negative results and false positive results. in contrast, egt has a relatively lower sensitivity, while p test has a relatively lower specificity. for example, relative small molecular mass of the shreds of fdps may lead to negative result using p test. so it is more valuable to compare the two indicators simultaneously [ , ] . euglobulin, a protein (including fibrinogen, plasminogen and other activins except for fibrinolysis inhibitor) separating out from plasma in acid circumstances, can be used to determine whether levels of plasminogen activators increase or not [ ] . when hyperfibrinolysis occurs, plasma levels of plasminogen decline, plasma levels of plasmin increase, and euglobulin suffer from accelerated dissolution by a great number of plasmin. the normal value of euglobulin lysis time (elt) is above h. that is to say dissolution within h means the occurrence of hyperfibrinolysis. domestic population data report the positive rate of elt test reaches - . %, when dic occur [ ] . furthermore there are other tests about fibrinolysis including tissue-type plasminogen activator test (t-pa), plasminogen activator inhibitor test (pai), plasminogen antigen test (plg:ag), plasmin activity test (pl:a), α -plasmin inhibitor test (α -pi) and so on [ , ] . blood platelet count reflects the absolute number of platelet in peripheral blood circulation. according to the reports at home and abroad, platelet count have a significant decline in patients with chronic severe hepatitis. moreover, studies on domestic population find that platelet count ranges from × /l to × /l in peripheral blood of severe hepatitis patients. some studies have already compared the platelet count among early stage, typical stage and late stage in severe hepatitis, and they have turned out to be × /l, × /l and × /l respectively. as a result, it indicates that platelet count is positively correlated with the severity of hepatitis [ ] . except for platelet count, other routine indicators including mean platelet volume (mpv), plateletcrit (pct) and platelet distribution width (pdw) also have significant reference value. when severe hepatitis occurs, the above-mentioned three indicators will be dramatically lower, and they have the tendency to continue declining with illness progressing. what's more, platelet quality tests include platelet aggregation rate, platelet factor validity tests and blood clot retraction test (table . ) [ ] . diagnosis of blood hypercoagulability in the early stage of dic relies on several molecular marks including plasma thrombinogen segment + (f + ), thrombinantithrombin complex (tat) and d-dimer, due to no significant changes in general laboratory tests. this stage is characterized by the elevated levels of those three molecular marks, and levels will increase more significantly with the occurrence of typical dic symptoms. dynamic monitoring of above-mentioned indicators is helpful for early diagnosis of dic [ ] . the stages are mainly characterized by the decline of blood clotting factors (including factorv (fv), factorvii (fvii), factorxii (fxii), factorix (fix), fac-torx (fx)), platelet count and plasminogen, and increasing of fibrinolytic the elevating levels of fibrin(−ogen) degradation products (fdps) and d-dimer; the shortening of euglobulin lysis time (elt) and the positive reaction of p test [ , ] . with the occurrence of dic, there will be a wide range of blood coagulation and highly-activated fibrinolysis in the patient's body [ ] . what's more, abnormal increased soluble fibrin monomer and fdp fragments will exist in plasma [ ] . the level of soluble fibrin monomer complex (sfmc), a complex combined fibrin monomer with fdp, is determined by p test. the p test shows positive with the occurrence of secondary fibrinolysis, whereas it shows negative with the occurrence of primary fibrinolysis. this means p test is negative when there is no blood coagulation. domestic population data report that the positive rate of p tests reach - %. however, the test can't be used as the ideal diagnostic indicator for dic, owing to many affected factors. false positive reactions are mainly found in the following conditions: gastrointestinal bleeding, massive hemoptysis, malignant carcinoma or blood sampling reserved improperly, whereas false negative reactions are usually found at the late period of the stage of secondary fibrinolysis [ , ] . as mentioned above, plasma thrombinogen segment + test (f + ), thrombinantithrombin complex test (tat) directly reflects production of intracorporeal thrombase, which increased in the early stage of dic. plasmin degrades the crosslinked fibrin to release fibrin degradation products and expose the d-dimer antigen, which reflects production of intracorporeal plasmin. d-dimer will have an apparent increase with the occurrence of secondary fibrinolysis, but d-dimer test shows negative when primary hyperfibrinolysis occurs. monitoring the above molecular markers dynamically is helpful to estimate therapeutic efficacy and guide treatment [ ] . as mentioned above, clinical manifestations of blood coagulopathy in patients with severe hepatitis are lack of specificity. the most common manifestation is bleeding, not only little hemorrhage from superficial sites, but also massive hemorrhage from internal sites, such as esophagogastric varices [ ] . the diagnosis of severe hepatitis complicated with blood coagulopathy is mainly based on the results of laboratory tests and clinical manifestations [ ] . according to current guidelines, basic diagnose conditions of dic contain the following points [ ] . . severe or multiple bleeding tendency. . microcirculation collapse or shock which is difficult to explain using protopathy. . a wide range of embolism in skin and mucosa, focal ischemic necrosis and ulcer, or unexplained dysfunction of kidney, lung, brain. . anticoagulant therapy is effective. if severe hepatitis b patients have one of the above-mentioned points except for ( ) and exhibit blood coagulating easily or prothrombin time (pt) shortening over s simultaneously, it can be considered as the early stage of dic in which the tendency of bleeding is not obvious. in addition, if severe hepatitis b patients have two of the above-mentioned four points, dic can be considered as the preliminary clinical diagnosis. furthermore, it can be definitely diagnosed when combined with the aforementioned items of laboratory tests ( table . ). firstly, we should use anti-viral therapy as a mainly method to treat primary disease of severe hepatitis b [ ] . then, we must eliminate the incentive, maintain the balance of water and electrolyte, and correct hypoxia and acidosis. it is very important to focus on massive upper gastrointestinal hemorrhage and disseminated intravascular coagulation when treating coagulation disorders [ ] . gastrointestinal bleeding, including esophageal varices bleeding and non-bleeding esophageal varices, were correlated with coagulation dysfunction and portal hypertension. prevention is still focused on improving the coagulation function, including adequate vitamin k supplements, coagulation factors, fibrinogen, fresh plasma or platelets supplements. it is particularly critical to control diet in patients with severe hepatitis. in which, light and easily digestible diet was recommended, but rough, hard and too greasy food was prohibited. for these patients, appropriate antacids can be used to protect the gastric mucosa [ ] . . general treatment: bed rest is necessary, meanwhile vital signs were closely monitored. the patients can eat liquid diet when bleeding mildly or having no active bleeding. however, abrosia is required when the patients have a heavy bleeding. . fluid resuscitation: firstly, intravenous access should be established rapidly in patients with gastrointestinal bleeding, then, the patients were given intravenous infusion of normal saline, lactated ringer's solution, plasma, whole blood or plasma substitute [ ] . . vaso-active agents: vaso-active agents such as dopamine and alamin may be given to maintain normal blood pressure, if blood circulation is still not stable after fluid resuscitation [ ] . . hemostatic: if mucosal bleeding was caused by portal hypertension, oral administration of norepinephrine and ice normal saline can promote mucosal vascular contraction and hemostasis. in addition, oral administration of yunnan baiyao may be effective [ ] . . acid suppression therapy: the proton pump inhibitors, such as omeprazole, pantoprazole and esomeprazole, are commonly used to inhibit gastric acid secretion [ ] . . reduction of portal pressure: patients with severe hepatitis complicated by gastrointestinal bleeding often accompanied with portal hypertension, so it may be considered to give them drugs to decrease portal pressure. especially in patients with portal hypertension gastropathy, reduction of portal pressure is more important than acid suppression therapy [ ] . . compression hemostasis via using three-chamber double-balloon catheter: after the above treatment, if there is still active bleeding in patients with bleeding esophageal varices, it can be considered to use three-chamber double-balloon catheter [ ] . . endoscopic hemostasis: if the above treatments have no effect in upper gastrointestinal bleeding, endoscopic hemostasis, including endoscopic hemostatic agents spray, endoscopic ligation or endoscopic injection sclerotherapy may be used [ ] . . others: surgery or interventional therapy may be considered, if internal medicine therapy is ineffective [ ] . firstly, we should treat original disease, then improve coagulative function in those patients. in addition, prevention and control of infection, correction of electrolyte disturbance, avoiding the hemorrhage and reduction of allergies and transfusion reactions should be considered [ ] . the key is early diagnosis and early treatment. . anticoagulant drugs: low molecular weight heparin is the most commonly used drug in earlier stage of dic. it is recommended to periodic test blood routine and coagulation, and dynamically observe coagulation status during medicine therapy. others such as dextran, anti-platelet aggregation ticlopidine, salvia injection, urokinase may be effective, but it should be support by more evidence-based medicine [ ] . . plasma and blood products: during the process of dic formation, the patients should be transfused with fresh plasma, each - ml/kg. when they developed the stage of secondary fibrinolysis, prothrombin complex containing coagulation factors, cryoprecipitate and platelets can be supplemented due to a large consumption of coagulation factors [ ] . . others: such as hemodialysis, anti-fibrinolytic -aminocaproic acid and tranexamic acid should be supported by more evidence-based medicine. hepatorenal syndrome (hrs) is a common and serious complication occurring in patients with decompensated cirrhosis and liver failure, who have overt circulatory dysfunction. the -year incidence of hrs in patients with ascites is about % [ ] . hrs may predict a poor prognosis in spite of it being functional reversible [ , ] . the clinical characteristics of hrs were first described in [ ] . in , hrs was defined by a group of international investigators as a progressive renal dysfunction that occurred in severe liver diseases, with features of prerenal failure (low urine sodium concentration and hyperosmolar urine) but without any improvement following volume expansion [ ] . the international ascites club (iac) developed hrs definition in , as a syndrome that occurs in patients with cirrhosis, portal hypertension and advanced liver failure, characterized by impaired renal function with marked abnormalities in the arterial circulation and activity of endogenous vasoactive systems [ ] . hrs is a potentially reversible syndrome that occurs mainly in patients with liver cirrhosis, ascites and all kinds of liver failure [ ] . it's clinical features include impaired renal function, marked changes in cardiovascular function and over activity of the renin-angiotensin systems and the sympathetic nervous. progressive hrs with severe renal vasoconstriction is able to cause a decrease of gfr [ ] . clinically, hrs can be divided into two types ( and ). type- , so-called acute hrs is a rapid progressive form of renal failure defined by doubling of the initial serum creatinine concentrations to a level higher than mmol/l ( . mg/dl) or a % reduction of the first h creatinine clearance to < ml/min within weeks [ ] . it appears mainly in patients with acute liver failure, but often develops after a precipitating event, such as bacterial infections (especially spontaneous bacterial peritonitis, sbp). the prognosis of type is poor with the median survival about month [ ] [ ] [ ] . type- , so-called chronic hrs is a steady or slowly progressive form of renal failure defined by serum creatinine from to mmol/l or from . to . mg/ dl [ ] . type- hrs is mostly related to refractory ascites. survival of patients with type- hrs is generally around - months, which is better than that of patients with type- hrs but shorter than that of non-azotaemic cirrhotic patients with ascites. patients with type- hrs tend to develop type- hrs while infections or other trigger events occurred [ , [ ] [ ] [ ] ]. portal hypertension is the essential factor of haemodynamic changes, which resulted from the development of cirrhosis associated with distortion, compression and even obliteration of the hepatic sinus and vessels [ ] . in patients with portal hypertension, bacterial translocation is increased and intrahepatic hypercontractile stellate cells activated [ , ] . this overall increased resistance to portal hypertensional causes increased local production of various vasodilators such as nitric oxide, leading to splanchnic vasodilation [ , ] . there are several other factors contributing to the splanchnic vasodilation, including hyporesponsiveness of the splanchnic vessels and mesenteric vascular hyperplasia [ ] . in addition, portal hypertension per se can cause renal vasoconstriction by activating sympathetic nervous. for example, when tips is used to reduce portal hypertension and improve renal blood flow, a corresponding reduction in sympathetic nervous activity has been observed [ , ] . as a result of splanchnic vasodilation, blood is accumulated in the splanchnic vascular bed just like a splanchnic steal syndrome [ ] . the combined effect leads to reduction in the effective arterial blood volume (relative hypovolemia) causing a relative inadequacy in the systemic circulation, which triggers a hyperdynamic circulation in these patients [ , ] . vasodilatation induces activation of neurohumoral systems including the reninangiotensin-aldosterone system (rass); sympathetic nervous system (sns); and non-osmotic release of antidiuretic hormone (adh) [ ] . relative hypovolemia initially causes sodium and water retention, increases intravascular volume, and simultaneously increases cardiac output. as cirrhosis progresses, vasodilatation aggravates, which activated vasoconstrictive systems, causing renal vasoconstriction and decreased renal blood flow [ , ] . local release of potent vasodilators such as nitric oxide (no) leads to splanchnic visceral vasodilation, as well as enables the splanchnic circulation against a variety of vasopressor agents, including norepinephrine, vasopressin, angiotensin ii and endothelin [ ] . the resistance of the splanchnic circulation to these vasopressor agents makes the control of arterial pressure during cirrhosis dependent on the extra-splanchnic effects produced by the endogenous vasoconstrictor systems. the role of vasoconstrictors in maintaining haemodynamic stability becomes pivotal as arterial vasodilatation increases during cirrhosis, which makes clear why cirrhotic patients with hrs are prone to develop renal, hepatic and cerebral vasoconstriction [ ] . the reduction of effective arterial blood volume leads to the compensatory activation of various vasoconstrictor systems. normally, the kidneys increase the production of renal vasodilators including prostaglandins and kallikrein to maintain blood flow. however, renal vasodilator production is generally reduced in patients with cirrhosis, thus contributing to renal vasoconstriction. this type of renal hypoperfusion further increases the production of various intrarenal vasoconstrictors such as angiotensin ii and endothelin, causing further decline of renal haemodynamics and renal function, occasionally accompanied by glomerular ischaemia and mesangial constriction [ ] . when blood pressure fluctuates, renal auto-regulation regulatory mechanisms initiate to make sure that the kidneys receive a relatively constant blood supply. when the critical threshold is below mmhg, renal blood flow is proportional to renal perfusion pressure which, in turn, is dependent on mean arterial pressure. in cirrhosis, with the development of liver disease in patients of cirrhosis, the renal auto-regulation curve gradually shifts to the right -which means as liver disease advances, the renal blood flow gradually decreases for each given renal perfusion pressure [ ] . furthermore, lumbar sympathetic blockade increases renal blood flow in patients with hrs, suggesting that the renal sympathetic activity is involved in this outgoing hepatorenal arm. insufficient cardiac output is considered one of the leading causes for renal hypoperfusion in patients with hrs in recent years. despite control of infection, the cardiac output of cirrhotic patients with sbp who developed progressive renal failure was lower than that in similar sbp patients without renal failure. similarly, when patients with non-azotaemic cirrhotic patients who developed hrs are compared with similar patients who did not, it is observed that low cardiac output and high plasma renin activity (pra) were independent predictors of hrs. in addition, in patients developing hrs, the evolvement of circulatory dysfunction leading to arterial hypotension and renal failure occurs in the setting of a continued decline in cardiac output and increase in pra. therefore, effective hypovolaemia occurs when cardiac output decreases, resulting in renal hypoperfusion and hrs [ ] . to summarize, the principal mechanisms leading to renal vasoconstriction include systemic circulation changes, accompanying portal hypertension which are characterized by decreased peripheral vascular resistance with subsequent vasodilatation, hyperkinetic circulation and the activation of compensatory mechanisms, i.e., the sns, raas, and adh. with the progression of cirrhosis, the combined effective of all the above factors will result in the gradual deterioration in renal function. any event that leads to a sudden deterioration in hemodynamics can cause a rapid renal dysfunction, precipitating type hrs [ ] . the diagnostic criteria for hrs have been first defined by iac in [ ] . the main findings include reduced glomerular filtration (creatinine clearance) less than ml/min or serum creatinine increased more than μmol/l after excluding the other causes of renal dysfunction. however, estimation of renal function by using creatinine clearance is not reliable, because these patients have lower levels of serum creatinine and higher renal tubular creatinine secretion compared with filtered creatinine. furthermore, it is often incomplete for the collection of a h urine. iac developed the new definition and diagnostic criteria for hrs in , which ( ) excludes creatinine clearance because of its inaccuracy of renal function estimation and the complicity to perform; ( ) includes renal failure at the time of combined bacterial infection (but absence of septic shock), indicating that hrs can be diagnosed before antibiotic treatment; ( ) determines by using albumin for plasma volume expansion better than saline. ( ) excludes minor diagnostic criteria (urinary indices) because of its poor sensitivity and specificity for the diagnosis [ , , ] . the diagnostic criteria of hrs for patients with liver cirrhosis are as follows: . cirrhosis with ascites; . serum creatinine > mmol/l ( . mg/dl); . no improvement in serum creatinine (decrease to a level of ≤ mmol/l or . mg/dl) after at least days of diuretic withdrawal and volume expansion with albumin. the recommended dose of albumin is g/kg body weight/day up to a maximum of g/day; . absence of shock; . no current or recent treatment with nephrotoxic drugs; . absence of parenchymal kidney disease as indicated by proteinuria > mg/ day, microhematuria (> red blood cells/high power field) and/or abnormal renal ultrasonography [ , ] . there are some other causes of renal failure in patients with cirrhosis that were not included, such as membranoproliferative glomerulonephritis and/or iga nephropathy in patients with chronic liver diseases. these chronic forms of kidney disease can cause acute rises in serum creatinine. determining whether it is a potential kidney disease or hrs that causes a sudden increase in serum creatinine in patients with cirrhosis and chronic kidney disease could be difficult [ ] . naturally, liver transplantation is the only rational solution in cases of advanced liver disease while it is also the treatment of choice for both type- and type- hrs. as calcineurin inhibitors (ciclosporin and tacrolimus) may induced gfr impairment, it is recommended to delay their administration until a partial recovery of renal function is recorded, usually - h after transplantation [ ] . clinically, the haemodynamic associated with hrs as well as neurohormonal abnormalities fade away within one month of transplantation, and the patients recover their ability to excrete sodium and free water. compared with patients without hrs, patients with hrs tend to have more complications, take more days in intensive care units and have higher in-hospital mortality rates after liver transplantation. nevertheless, their -year survival rate is acceptable ( % vs. - % in liver transplant patients without hrs [ ] . the primary limitation of liver transplantation is that most patients with type- hrs die before transplantation due to the shortage of donor liver and their extremely short survival time. reference to the model of end-stage liver disease (meld, including scr, tbil, inr) for organ prioritisation has partially addressed this problem, since patients with hrs have a high priority on the waiting list. in addition, the use of vasoconstrictors and albumin in the treatment of type- hrs can improve survival rate of these patients, and increase the likelihood of their transplantation [ ] . in patients with advanced liver disease and the renal insufficiency, simultaneous liver and kidney transplant (slkt) is taking into consideration. however the reversibility of renal function in some patients when they receive slkt should be taken into account. therefore, to ensure allocation of transplants only to those truly in need, the transplant community proposed an evaluation algorithm in , whose purpose is to determine the presence of kidney disease with structural damage (preferably on biopsy) before giving slkt. in the case of chronic kidney disease, a decreased creatinine clearance at ml/minute or less is considered an indication of slkt. slkt should not be performed for the patients with simple hrs, but for the patients with hrs who become dialysis dependent and without any recovery after to weeks of dialysis [ , ] . vasoconstrictors combined with albumin are the first line of therapy for type- hrs patients. it was recognized long time ago that the effective plasma volume was reduced when patients of advanced liver diseases complicated with hrs, and this led to many attempts to improve the patients' renal function by expanding their plasma volume, including a large dose of albumin or saline perfusion. with the advent of safer compounds including terlipressin, a vasopressin analogue with longer activity, and the α -agonist midodrine combined with octreotide the analogue, vasoconstrictors is widely used in the patients with hrs. these vasoconstrictors are able to ameliorate vasodilatation while increasing effective arterial blood volume, improving renal vasoconstriction and improving renal flow. in order to further increase effective blood volume, vasoconstrictors have been used in conjunction with intravenous albumin. the clinical results from uncontrolled studies including patients with hrs (type- , ) showed that a total of % were observed complete response (mostly defined as a decrease in scr to . mg/dl). interestingly, once the treatment is stopped, hrs relapses only in a few "responders" [ , ] . there were several randomized controlled trials (rcts) published, suggesting that terlipressin was associated with an increase in gfr compared with albumin alone or with a placebo. the rate of hrs reversal in the terlipressin group was higher than that in the control group ( % vs. %). as survival rate was not improved in the two largest rcts, liver transplantation is still the preferred treatment for hrs, but terlipressin seems to serve as a "bridging" treatment. two recent small, open-label rcts suggested that the incidence of hrs reversal and the rate of side effects showed no significant difference between the two groups of norepinephrine and terlipressin [ , , ] . the initial dose of terlipressin recommended in many studies ranged from . to mg per - h [ , ] . if the creatinine level did not decrease by % on the third day, the dose could be increased to mg every h or mg/days by continuous intravenous infusion, respectively. in some studies, the daily dose of albumin was generally - g by a load of g/kg body weight. some mentioned central venous pressure to establish albumin doses and to prevent body fluid from overloading. this treatment was maintained until hrs is reversed, but did not exceed weeks [ ] . about % of patients relapsed after the treatment withdrawal. however, these patients should be given repeated treatment with terlipressin, which is often effective [ ] . several studies have evaluated the role of transjugular intrahepatic portosystemic stent-shunt (tips) in hrs. these studies show that tips help decreasing in scr in most patients, even in a minority of organic renal failure, but it is slower compared to those obtained using terlipressin combined with albumin [ ] . recrudescence of hrs is rare provided the shunt remains patent, while hepatic encephalopathy often comes. it is worth noting that the vast majority of patients included in these studies suffered from alcoholic cirrhosis, many of whom have active alcoholism, and therefore the improvement observed may be caused by the improvement in an acute-on-chronic process. in addition, since all these studies excluded patients with a child-pugh score ≥ , resulting in a lack of data, the efficacy of tips should be further explored in rcts [ ] . extracorporeal albumin dialysis molecular adsorbent recirculation system (mars) is designed for making clearance of water-soluble cytokines (i.e., il- ) and albumin-bound toxins (i.e., bile acids) which is implicated in the pathogenesis of hrs. two studies showed that mars was ineffective in improving survival rate and emic haemodynamics in type- hrs [ , ] . another clinical observation including patients with type- hrs reported a rate of complete renal response of % [ ] . extracorporeal albumin dialysis (ecad) reduces serum creatinine levels, but it is not clear whether this effect is due to a real improvement of renal function or merely a filtration process. several studies demonstrated that patients' systemic haemodynamics improved after ecad, manifested as an increase in systemic vascular resistances and arterial pressure, as well as a decrease in cardiac output, pra and levels of norepinephrine. however, there were too few studies on the effect of ecad on survival in type- hrs patients to draw any definitive conclusions [ , ] . in addition, ecad is very expensive and therefore not suitable for wide and rapid clinical application [ ] . the treatment of type- hrs should take into account the survival rate as well as controlling the ascites. both hrs- and hrs- are indications for the tips treatment. the therapeutic effect of tips is excellent for its better controlled of complications of portal hypertension compared with other treatments. tips have been reported not only to improve renal function in patients with type- hrs but also to treat refractory ascites in patients with type- hrs [ ] [ ] [ ] . the contraindications to the creation of tips are shown in the followings [ ] . • contraindications to placement of a tips: - there were only a few studies evaluated the role of tips in type- hrs and the number of cases was quite low. in most patients, tips could decrease scr, even in some with organic renal failure [ ] . hrs recurrence is rare as long as the shunt remains patent, but hepatic encephalopathy often occurs [ ] . nine patients were followed-up for month after the treatment of tips in a study, eight cases were found with decreased scr decreased and notably controlled ascites. four patients died, two of them died within month, the other two died at months and months respectively. the remaining five patients survived for a long time. although tips can be used in improving refractory ascites which often contributes to type- hrs, data on the effect of tips on survival are still insufficient. therefore, the efficacy of tips should be further explored in randomized controlled trials (rcts) [ ] . the information about combining albumin and vasoconstrictive agents treated in type- hrs is limited. only a few patients with type- hrs have been specifically treated with terlipressin and albumin. in one clinical study, patients with hrs- were assigned to receive this treatment and of them achieved improvement of renal function. however after the treatment withdrawal, hrs patients showed relapsed during the follow-up. the most common side effects during terlipressin therapy are cardiovascular and ischemic and reported as an incidence of nearly %. the high recurrence rate of hrs after terlipressin and albumin treatment discontinuation suggests that they are less effective in treating type- hrs compared to type- hrs [ ] . prevention of hrs is important because it develops at a constant frequency in cases of spontaneous peritonitis (sbp) and advanced liver disease [ , ] . it becomes possible to prevent hrs if sbp is diagnosed and treated promptly [ ] . according to current data, using albumin in combination with antibiotics for the treatment of patients with sbp seems to be warranted but only for those with jaundice or renal dysfunction. the prophylactic use of antibiotics in cirrhosis with gastrointestinal bleeding also seems to be necessary, because the use of antibiotics contributes to reducing incidence of infection and rebleeding whereas improving survival rate. furthermore, the incidence of hrs in sbp patients decreases by albumin administration, and prevention of hrs can also be related to increased survival. the recommended dose of albumin is . g/kg body weight on the first day then g/kg body weight on the third day, a maximum of g and g, respectively. albumin administration is strongly recommended in sbp patients with serum bilirubin levels higher than . mmol/l ( mg/dl) or serum creatinine more than . mmol/l ( mg/dl). a placebo-controlled rct that enrolled the patients with low (< . g/l) ascites protein who also had advanced liver diseases or "renal dysfunction"(defined as scr ≥ . mg/dl or blood urea nitrogen≥ mg/dl, or serum sodium level ≤ meq/l) suggested that oral norfloxacin contributed to a reduced hrs incidence within year ( % vs. %) and an improvement in survival at the end of months [ ] . norfloxacin may ameliorate or prevent vasodilatation by reducing bacterial translocation and overt infections, as well as suppressing plasma renin activity, thereby prevent these patients from developing hrs. the concept that the severity of the clinical course of patients with cirrhosis complicated with serious bacterial infection is related to the degree of an impairment of circulatory function, which has led to new and effective approaches in the prevention and treatment of these complications. in patients with severe hepatitis, multiple causes may lead to disorders of internal environment, mostly manifesting fluid and electrolyte imbalance as well as acidbase imbalance, usually resulting in deterioration, greater complexity and even death. accurately recognizing the occurrence of severe hepatitis with complications such as fluid and electrolyte imbalance and/or acid-base imbalance, and therefore giving appropriate treatment to maintain balance of internal environment, is of great importance for improving prognosis of the patients [ ] . water is the major component of human body. electrolytes are substances that dissociate in solution to form charged particles, orions. body fluid comprise mainly of water and electrolytes and electrolytes comprise mainly of na + , k + , ca + , mg + , cl − , hco − , hpo − and so − . the primary function of electrolyte include: ( ) to maintain osmotic pressure and acid-base balance of body fluids; ( ) maintain nerve, muscle, cardiac cells resting potential, involved in the formation of action potentials; ( ) involved in metabolism and physiological activities [ ] . body fluid include intracellular fluid and extracellular fluid, the latter can be divided into plasma and interstitial fluid. intracellular and extracellular fluid differ in ion components. na + is major cation in extracellular fluid and its main anions are cl − and hco − . k + is major cation in intracellular fluid and its major anion is hpo − . the total number of ions in body fluids is called osmolality, its unit is mosm/l. if the osmolality on both sides of a semipermeable membrane is not equal, water moves toward the side with the higher osmolality. this phenomenon is called osmosis [ ] . osmosis of water can be opposed by applying a pressure across the semipermeable membrane in the direction opposite to that of the osmosis. the amount of pressure required to oppose the osmosis is defined to be osmotic pressure. in spite of various solute concentrations are different in extracellular and intracellular fluid, the osmotic pressure remain equal. normal plasma osmotic pressure is - mosm/l. steady osmotic pressure is the basic guarantee to maintain the fluid balance across the cell membrane. multiple mechanisms in nervous and hormonal system are involved in the regulation of body fluid and electrolyte balance [ ] . ( ) there exists sensation of thirst in central nervous system, which plays an important role in regulating body water. ( ) there is a powerful feedback system for regulating plasma osmolarity and sodium concentration that operates by altering renal excretion of water independently of the rate of solute excretion. a primary effector of this feedback is called antidiuretic hormone (adh) which plays an important role in regulation of renal concentration and dilution to maintain the body fluid homeostasis. ( ) reninangiotensin-aldosterone system (raas) is an important regulator of sodium reabsorption and potassium secretion by the renal tubules. human body fluid environment must be suitable ph value for maintaining normal metabolism and physiological function, under normal conditions, human body take in acidic and basic food and drinking water, produce acids and bases during metabolism and eliminate acidic or basic substances by the kidneys and lungs. in the plasma, the normal ph value ranges from . to . with an average value at . . the regulation of the acid-base balance is accomplished by the buffer system of the body fluid, the respiration of the lungs and the excretion of the kidney [ ] . ( ) blood buffer system is composed by a weak acid and its corresponding buffer base, includes bicarbonate buffer system, phosphate buffer system, plasma protein buffer system, hemoglobin and oxygen synthetic hemoglobin buffer system. ( ) the role of the lung in acid-base balance is to adjust the concentration of plasma carbonic acid by changing the amount of co , so that the ratio of hco − and h co in the plasma is close to normal, so that the ph is relatively constant. ( ) the major role of the kidneys in maintaining acid-base balance is to conserve circulating stores of bicarbonate and to excrete h + . the kidneys maintain ph by increasing urinary excretion of h′ and conserving plasma hco − when the blood is too acidic, or increasing urinary excretion of hco − , and decreasing urinary excretion of h + when the blood is too alkaline. patients with severe hepatitis are prone to develop water retention, with the main manifestation of seroperitoneum (ascites) as well as body weight gain. with the acatharsia of water becoming more serious, oliguria and edema of lower extremities occur. sbp (spontaneous bacterial peritonitis) can also occur, which is manifested with symptoms such as fever and abdominalgia. several factors contribute to ascites include an increase in capillary pressure due to portal hypertension, obstruction of venous and lymph flow through the liver, decrease in colloidal osmotic pressure due to impaired synthesis of albumin by the liver, salt and water retention by the kidney [ ] . some theories have been used to explain the increased salt and water retention by the kidney. because of vasodilation or an actual loss of fluid into the peritoneal cavity, the effective blood volume maybe reduced, which may in turn decrease the renal blood flow leading to a lower glomerular filtration rate (gfr) and an activated rennin-angiotensin-aldosterone system (raas). the diagnosis of water retention depends on typical clinical symptoms such as ascites, pleural effusion, and edema of lower extremities, when the body begins to excess water, blood pressure is increased, which leads to many complications such as congestive heart failure and pulmonary edema. hyponatremia is a common complication in severe hepatitis patients, and always incorporate with the retention of water and sodium, but the total sodium can be decreased, normal or even increased, that is dilutional hyponatremia. in laboratory test, serum sodium is below . mmol/l. the mechanism of hyponatremia probably depends on the following factors: ( ) the decreased function of adh inactivation in liver brings adh increase, enhancing the reabsorption of water in renal tubule, which causes the formation of water retention. this is the main cause of dilutional hyponatremia. ( ) water retention brings about volume extension, causing the aldosterone secretion decrease, which leads to the sodium egestion increase in urine. ( ) some severe hepatitis patients frequently vomit, and can not eat, bringing about a major loss of body fluid and electrolyte. ( ) severe hepatitis patients, serum albumin reduces, combining with the factors such as poor appetite, anorexia, fasting or limit sodium, bring about a state of low permeability in the cell, which causes the extracellular na + moving into the cell. ( ) iatrogenic factors, exhaust potassium diuretic such as hydrochlorothiazide and furosemide and spironolactone have a strong role in the excretion of sodium, so a large number diuretic is liable to hyponatremia in ascites patients, and in the treatment of cerebral edema, a large infusion of mannitol may cause hyponatremia either [ ] . hyponatremia due to the osmotic pressure of extracellular fluid decreased, water moves to the cells, causing cell edema, especially brain edema. so the symptoms of nerve system are the main manifestation in hyponatremia patients [ ] . generally dilutional hyponatremia in severe hepatitis patients develops slowly and progressively, and the symptoms are often covered by primary disease symptoms, like weak, feeble, nausea, vomiting, lethargy, significant body weight increase, pale and moist skin and sometimes saliva, tears increase. improper treatment will bring about a sharp serum sodium decrease in the short term, such as serum sodium rapidly decreasing to below mmol/l, acute hyponatremia syndrome comes, the manifestation include convulsions, coma, hypotension, pulse narrowing, tachycardia, oliguria even respiratory arrest and death. if cerebral hernia happens, corresponding nerve location signs will follow. hypokalemia refers to the condition in which the concentration of potassium (k + ) in serum is less than . mmol/l. it could occur during the whole period in severe hepatitis patients and is more common in early metaphase of disease. hypokalemia can be the result from one or more following medical conditions: ( ) insufficient intake of potassium due to the poor appetite or anorexia in the patients with severe hepatitis. ( ) frequent vomiting leads to excessive loss of stomach acid, which causes alkalosis and extracellular potassium is transferred into cells. ( ) the reduce of effective circulating blood volume can cause to high aldosterone levels and excessive urinary losses of potassium. ( ) the decreased function of aldosterone inactivation in liver brings aldosterone increase, enhancing urinary losses of potassium. ( ) some medications such as diuretics can also cause urinary losses of potassium. the clinical syndromes of hypokalemia are related to the degree of the shortage of intra/extracellular potassium and disorders of other electrolytes and acid-base, but more depends on how soon it occurs. in the early time it shows muscle weakness, first in limbs, then develops to the torso and respiratory muscle. deficiency of potassium also can lead to weak peristalsis, poor appetite, sick and constipation in mild hypokalemia but abdominal distention and paralytic ileus in severe situation. in cardiac syndromes, it mainly presents atrioventricular block and arrhythmia which including premature ventricular contraction or atrial premature beats, sinus bradycardia, paroxysmal auricular tachycardia or junctional tachycardia, even ventricular fibrillation. in hypokalemia state an increasing shift of potassium from extracellular fluids into cells and an obligate loss of potassium from kidney can cause metabolic alkalosis and abnormal acidic urine. long-term hypokalemia also can lead to hypokalemic nephropathy with proteinuria and cylindruria syndrome. the changes of ecg [ ] : in the early stage, flattened t wave and an obvious u wave, st-segment depression can be found, qu interval is widen. in severe situation, a wide pr interval, low voltage, wide qrs interval and ventricular arrhythmia can occur. hyperkalemia refers to the condition in which the concentration of potassium (k + ) in serum is higher than . mmol/l. it is more common in the middle and late period of severe hepatitis. the mechanism of hyperkalemia: ( ) the most usual way lead to hyperkalemia is oliguria or uroschesis which are caused by renal dysfunction among the patients have severe hepatitis with hepatorenal syndrome. ( ) metabolic acidosis and na + -k + -atp enzyme inactive lead to a shift of potassium out of cells also contribute to develop hyperkalemia. ( ) long-term and high dose potassium-sparing diuretics applied during the treatment lead to hyperkalemia is not rare, and easy to get sudden death in patients. hyperkalemia mainly influences myocardium and skeleton muscle. the most dangerous situation is fatal arrhythmia. when the concentration of k + is higher than . - . mmol/l, there are peaked t waves. when it is over - mmol/l, pr interval is widen and p wave is flattened even vanish. when it is up to - mmol/l, t waves and qrs complex can evolve to sinusoidal shape and cardiac arrest. hyperkalemia is a common critical and severe symptom in clinic. when it happens, all of the potassium-sparing diuretics and potassium uptake should be stopped. at meantime, the treatment against the toxicities to myocardium and skeleton muscle should be taken to accelerate a shift into cells and potassium excreting. there also can happen hypocalcemia, which shows neuromuscular excitability, cardiac electrical instability and instable emotion. the concentration of calcium in serum under mmol/l is significant in diagnosis. hypomagnesemia can be found too. it mainly presents similar symptoms as hypocalcemia such as weakness, muscle cramps, increased irritability, tetany and chvostek positive. hypomagnesemia can cause cardiac arrhythmia, when it occurs, the concentration of magnesium is less than . mmol/l, and it should be urgently treated. severe hepatitis patients prone to acid-base imbalance [ ] , mainly to alkalemia. the main types of acid-base imbalance include respiratory alkalosis, metabolic alkalosis, respiratory alkalosis plus metabolic alkalosis, secondly include respiratory alkalosis plus metabolic acidosis, triple acid-base disorders (tabd), metabolic alkalosis plus metabolic acidosis [ ] . history and clinical manifestations provides important clues for the judgment of acid-base imbalance. the result of blood gas monitoring is the decisive basis for judging the type of acid-base imbalance. serum electrolyte examination is an important reference. anion gap (ag) has important diagnostic value in determining the type of acidbase imbalance [ ] . ag = na + -(cl − + hco − ) is a simple formula for the value between the number of cations and anions in serum. its normal value was ± mmol/l. ag can not only help diagnose "potential" metabolic acidosis and to distinguish different types of metabolic acidosis, can also help determine special types of mixed acidosis, and has its unique role in the judgment of tabd. sometimes the indicators of blood gas analysis are normal, the calculation of ag value become the only evidence of diagnosis of metabolic acidosis. in addition, ag value can be used as reference for correction of acid-base imbalance. in severe hepatitis, the change of ag values can be used as an indicator to estimate the complications and prognosis. clinical observations indicate: ag value significantly increased often suggestive of severe infection, kidney dysfunction or severe bleeding, and the prognosis is poor. respiratory alkalosis refers to arterial paco decrease and ph > . as well as compensatory decrease of blood hco − . respiratory alkalosis occurred in early stage of severe hepatitis. in severe hepatitis, respiratory alkalosis related to hyperventilation: accumulated ammonia and other vasoactive peptides excited respiratory center, ascites and pleural fluid increase respiratory rate, hypoxemia excited respiratory center. compensatory mechanisms: co reduction, breathing shallow and slow, so that co retention, h co compensatory rise; when last longer, reduce renal row h + , hco − excretion increased, hco − /h co equilibrium at a low level. most patients have the performance of shortness of breath and heart rate increase. can have vertigo, hand, foot and mouth numbness, muscle tremor, hand and foot convulsions. convulsions associated with low calcium. dysfunction of nervous system is related to the damage of brain function and cerebral blood flow decrease. respiratory alkalosis diagnosis relies on the following: ( ) ph is normal when fully compensated, increased underdecompensation. ( ) paco lower (typically < mmhg or . kpa). ( ) hco − compensatory decline. ( ) ag value may have a slight increase. ( ) blood cl − may increase. metabolic alkalosis refers to the type of acid-base imbalance characterized by an increase hco − in extracellular liquid. the inappropriate application of basic drugs, potassium-sparing diuretics, dehydrating agents, hormones can often induce or aggravate metabolic alkalosis. severe gastrointestinal symptoms, anorexia, vomiting or diarrhea are also the reasons for the occurrence of metabolic alkalosis. compensatory mechanisms: when alkaline substances increased in body, buffer system instantly transfer strong base into weak base, increase hco − consumption, h co increased. inhibit respiratory center, decrease pulmonary ventilation, co retention, hco − compensatory increase. renal carbonic anhydrase activity decreased and h + formation and excretion decreased, nahco reabsorption is also reduced, so hco − /h co compensatory restore to : , ph value is normal. patients with mild metabolic alkalosis usually have no obvious symptoms. many disorders can occur in severe metabolic alkalosis. ( ) functional changes in the central nervous system, the patient may have irritability, confusion, delirium, consciousness disorders. ( ) slow and shallow breathing, hypoxemia. brain tissue is particularly sensitive to hypoxia, thus neurological symptoms first appeared. ( ) hypocalcemia and neuromuscular stress increased, the performance of tendon hyperreflexia, face and muscle twitching, and limbs twitching. ( ) hypokalemia can cause neuromuscular symptoms and arrhythmias. according to ph value, paco , hco − , level of k + and cl − , effective circulating blood volume and performance of primary disease, diagnosis of metabolic alkalosis is no difficult to make. metabolic alkalosis should be divided into two categories based on the urinary level of cl − . ( ) chloride positive metabolic alkalosis: supplement sodium chloride can correct the alkalosis. it indicates that the body has cl − deficiency, urinary cl − < mmol/l. ( ) chlorine negative metabolic alkalosis: alkalosis can not be corrected by supplement sodium chloride, urinary cl − > mmol/l. respiratory alkalosis plus metabolic alkalosis tend to occur on the early stage of severe hepatitis. in most cases, there is no obvious complication, more often metabolic alkalosis happens on the basis of respiratory alkalosis, or the other way around. due to respiratory and metabolic factors are inclined to alkaline change, name as reduce paco and elevated plasma hco − , there is no mutual compensation between them, so it is easy to present as severe decompensation and poor prognosis. main point of diagnosis: ( ) ph value of blood increase significantly. ( ) paco decrease. ( ) hco − increases, the value should be greater than . ×( -paco ) + . . ( ) hypokalemia and hypochloremia are common phenomenon. respiratory alkalosis plus metabolic acidosis is relatively rare. metabolic acidosis can be divided into types: ( ) value of ag is normal (high chlorine acidosis), commonly seen at long-term diarrhea, combined with renal tubular acidosis and a large amount of physiological saline input in patients or water intoxication. ( ) high ag value type (normal blood chlorine acidosis), regularly present in combination of hepatorenal syndrome, lactic acidosis and patients with ketoacidosis. ( ) a hybrid type (high ag merged high blood chlorine), mainly in patients with severe diarrhea following lactic acid or ketoacidosis. when respiratory alkalosis plus metabolic acidosis happens, paco and plasma concentration of hco − are higher than scope of compensation to each other. its characteristics as follows: ( ) the range of blood ph change is not large, normal, slightly higher or slightly lower. ( ) paco reduce to less than . × hco − + or hco − < -( -paco ) × . - . . ( ) ag values can be normal or elevated, the latter is more common. if the elevated blood cl − value is equal to the hco − decrease, ag value normal metabolic acidosis type can be diagnosed; the cases that the rising value of ag is equal to the decline of hco − values can be diagnosis as ag increased metabolic acidosis type. on the third occasion, the increase value of ag is equal to the sum of hco − and cl − drop, the diagnosis is mixed metabolic acidosis. for this type of offset mixed acid-base balance disorders, treatment should be moderate, the measure of the metabolic factors correcting should be precede to respiratory factors, avoid paco quickly returning to normal in the process of treatment, which would lead to blood ph drop rapidly and acidosis more worse. to patients with severe hepatitis, the harm of alkalosis is greater than acidosis, thus the blood ph should be kept slight acidic in a normal state. generally, the target of alkali supplement can be arterial blood ph value recovered to . . respiratory alkalosis tabd refers to respiratory alkalosis, metabolic acidosis and metabolic alkalosis three primary imbalances coexist in the same patient, which is one sort of serious acid-base imbalance, mostly develops in the late stages of severe hepatitis, fatality rate is high. respiratory alkalosis tabd characteristics as follows: ( ) blood ph value depends on the relative severity of these three primary imbalances, which can be normal, or slightly high generally. ( ) reduce paco , its value is less than . xhco − + . ( ) hco − can raise, normal or lower. ( ) value of ag rise significant, and extent of ag raise is greater than the hco − lower. ( ) cl − often lower than normal. the occurrence of metabolic alkalosis plus metabolic acidosis in patients with severe hepatitis is not uncommon. usually it is accompanied with existing lactic acidosis or ketoacidosis, and the patient may manifest frequent vomiting. since the causes for raising and lowering hco − coexist, they tend to cancel one another. the ph and hco − concentration can be normal, increased or decreased, depending on the relative severity of the two kinds of imbalances. severe hepatitis can also be accompanied by metabolic acidosis, metabolic acidosis plus respiratory acidosis, tabd of respiratory acidosis, and etc. pure metabolic acidosis refers to arterial blood ph < . and compensatory decline of paco due to primary decrease of hco − . typical manifestation is known as kussmaul breathing, characterized by deeper and faster breathing, as well as obvious contraction of respiratory muscle, and also ketone-smelled exhaled breath. the patients often flush, companied by increased heart rate and decreased blood pressure. there may be reduced or disappeared tendon reflexes, confusion or stupor. due to respiratory and metabolic factors both towards to acidic changes, there is no respiratory compensation for decrease in hco − , nor renal compensation for increase in paco , hence presenting severe decompensated status. the resulting distinct decrease in ph and vicious circle are the characteristics for metabolic acidosis plus respiratory acidosis. the characteristics for tabd of respiratory acidosis include significantly increased paco , elevated hco − , ag > mmol/l, and significantly decreased cl − . the incidence of the above types of acid-base imbalance is very low in patients with severe hepatitis. once it happens, it should be actively treated with corresponding methods, so that the blood ph quickly restores to the safety range. during therapeutic process against various pure acid-base imbalance, interactions among various treatments need to be taken into consideration, in order to avoiding the possibility that the treatment for one type of acid-base imbalance causes or aggravates another type. in summary, water-electrolyte imbalance and acid-base imbalance have a relatively high morbidity in patients at various stages of severe hepatitis. this often results in deterioration and complication of the disease, evermore, the death of the patient. therefore, the functions of heart, lung, kidney, blood circulation as well as changes of body weight in the patient need to be intently monitored. regular detection of k + , na + , cl − , carbon dioxide combining power (co cp), blood urea nitrogen, creatinine, ph, data about arterial blood gas analysis, and also detailed records of patient's input and output are demanded. during the process of diagnosis and treatment, careful analysis of the history, clinical manifestations and laboratory examination are necessary to achieve correct diagnosis, early prevention and prompt treatment. in normal condition, proper amount fluids can make a lubrication action on organs in peritoneal cavity. but in those patients who have severe hepatitis, especially with cirrhotic portal hypertension, too many fluids over ml can lead to ascites. the ascites can be categorized into uncomplicated ascites and refractory ascites. there is no infection in uncomplicated ascites and won't lead to hepatorenal syndrome, but refractory ascites is in the contrast. the refractory ascites includes diureticresistant and diuretic-intractable ascites. the diuretic-resistant ascites shows no response to diuretic and diuretic-intractable ascites limits the application of diuretic due to the complications induced by diuretic. mg antisterone per day as an initial dose can be given to those patients with moderate ascites, if it goes to no satisfied effect, mg can be added after every days till the maximum dose to mg/d. if the patients show a hyperkalemia or aldosterone antagonist-resistant, nicorol can be combined and with an increasing dose from to mg/d gradually. the patients without edema losing weight should be less than . kg and those with edema should be less than kg per day to avoid electrolyte disturbance or hepatorenal syndrome during the whole treatment period. diuretics should be withdrawn on the patients with severe hepatic encephalopathy, severe hyponatremia, progressive renal failure or severe muscle spasm. the patients should only take the minimum dose of diuretics to maintain the state after the syndrome controlled, or withdraw when it is necessary. due to the poor outcome and living quality, the median survival time of refractory patient is half year. so liver transplantation can be considered in the patients with refractory ascites induced by cirrhosis, which required more cautious before make a diagnosis. generally, if the patients meet the following conditions when they are receiving mg/days antisterone and mg/days nicorol treatment over week and sodium uptake limited in mmol/days, refractory ascites can be diagnosed. ( ) losing weight less than . kg/days over days ( ) sodium uptake is more than elimination ( ) grade - ascites is arisen again after -week long treatment ( ) hepatic encephalopathy, hepatorenal syndrome and severe electrolyte disorder induced by diuretics are shown up. refractory ascites patients without complications can be treated with abdominocentesis, but it will possibly induce circulation failure, and increases the risk of hepatic coma and hemorrhage. so low rate infusion of albumin with abdominocentesis is combined to avoid circulation failure, meantime diuretics also need to give after abdominocentesis. aldosterone antagonist combined with nicorol is a suitable strategy: the dosage of antisterone is increased from to mg/days and nicorol from to mg/days gradually. the goal of this strategy is to maintain the situation without ascites under the minimum dosage. but when severe hepatic encephalopathy and electrolyte disorder show up, which means serum sodium concentration is less than mmol/l, serum potassium concentration is less than . mmol/l, nicorol should be withdrawn, if serum potassium concentration is more than . mmol/l, antisterone should be withdrawn. to those patients who need abdominocentesis repeatedly, transjugular intrahepatic portosystemic shunt (tips) can be considered, but the risk of hepatic encephalopathy will be higher and the outcome is poor. so the patients with severe liver and renal failure, cardiorespiratory function failure or active infection should be in cautious. the mean survival time of refractory ascites patients complicated with hepatorenal syndrome is months, preventive antibiotics combined with albumin is an option for these patients. to those patients who have already showed hepatorenal syndrome, terlipressin combined with albumin could be useful. meantime, abdominocentesis, tips or artificial liver supporting treatment can improve patients' living quality in short term, but long-term outcome won't be good, so liver transplantation should be execute as soon as possible. in general, medicine can improve the symptoms in short term but with poor outcome, liver transplantation is more meaningful. patients with hypovolemic hyponatremia can have a supplement with sodium and decrease the dosage of diuretics, patients with hypervolemic hyponatremia can restrict fluids uptake (less than ml/d) and combined with vasopressin v receptor blocker or antidiuretic hormone receptor blocker. currently, vaptans, tolvaptans, conivaptan and satavaptan have already applied in clinical practice. there was research showed that vaptans could improve - % patients' symptoms significantly after patients took it for week to month, and main side reaction was thirsty. the patients complicated with hepatic encephalopathy should be used vaptans with cautious due to its high risk in dehydration and hypernatremia. meantime, vaptans is metabolized by cyp a, so rifampin, barbital and phenytoin can decrease its effect, and ketoconazole, clarithromycin can increase its plasma concentration. tolvaptans can give some relief but increase the risk of hemorrhage. satavaptan will decrease patients' survival rate. so the proper treat period and side reactions of these drugs in long-term using need to make clear. potassium uptake should be withdrawn immediately after hyperkalemia occurs, emergency treatment for detoxicating potassium should be taken to protect cardiac. the treatment depends on the plasma concentration of potassium. the treatment for patients with fulminant hepatitis b with cirrhosis complicated with hyperkalemia is to restrict the uptake of potassium, improve the microcirculation, correct renal filtration decrease induced by hepatorenal syndrome and increase the elimination of potassium. there are also some patients have abnormal distribution of potassium due to hypoxia, acidosis, catabolism, and deficiency of energy supplies, which leads to intracellular potassium is transferred into extracellular. the treatment for these patients is to correct hypoxia and acidosis, high glucose, insulin and atp are administered to boost glycogen synthesis to transfer potassium from extracellular to intracellular. peritoneal dialysis and plasmapheresis can be given to the patients with intractable hyperkalemia. hypokalemia can present during the whole period of fulminant hepatitis b, it will occur more often on the early and middle stage. long-term inappetency and abdominal distension lead to insufficient potassium uptake. nausea, vomiting and diarrhea lead to increase of potassium losing. renal filtration rate decreasing and aldosterone increasing lead to potassium elimination increase. complicated with alkalosis and anabolism increase also can cause hypokalemia. the treatment for hypokalemia should focus on comprehensive therapy, correcting alkalosis, increasing potassium supply, improving microcirculation. disturbance of acid-base balance occurs quit often in hepatitis b patients, especially with alkalosis. during the early stage, it can present in pure respiratory alkalosis, also can complicated with metabolic alkalosis. during the middle and late stage, both of above symptoms and metabolic acidosis occurs concurrently. treating idiopathy and correcting hyperventilation is a treatment for respiratory alkalosis. arginine hydrochloride injection is used to treat metabolic alkalosis to avoid secondary metabolic alkalosis. the general regulation is prefer acid to base, till ph value of arterial blood back to . . in prevention of disturbance of acid-base balance, the effective strategy is to correct hypokalemia and hypochloremia, control vomiting. meantime, controlling infection, endotoxemia and upper gastrointestinal hemorrhage are necessary. hepatic encephalopathy (he) due to metabolic disturbance is a complex neuropsychiatric syndrome caused by severe liver dysfunction or disorder and is one of the common complications and causes of death in severe liver diseases. patients with he mainly present with neuronal or mental abnormalities and disturbance of consciousness, even coma and death. the clinical manifestations and the severity of the disease vary because of its complex pathogenesis. hepatic encephalopathy is the result of acute and chronic hepatic failure caused by cirrhosis or various kinds of portosystemic shunt (pss) created. a diagnosis of he can be made after excluding encephalon diseases. the syndrome is caused by metabolic disorders and is potentially reversible. he clinical features differ due to the wide degree and range of neuropsychiatric symptoms that vary from subtle abnormalities detected only by intelligence tests or electrophysiological methods geared for detecting personality changes to abnormal behavior, intellectual impairment, and even different degrees of consciousness disorders. he was previously known as hepatic coma, but that is only one of the worst severe signs of he and does not represent all types of he. in , the world congress of gastroenterology (wcog) suggested that based on the cause he can be divided into three types (a, b, and c) [ , ] . type a: type a is acute liver failure-related he and the symptoms occur within weeks. in subacute liver failure-related he, the symptoms of he occur within - weeks with or without predisposing factors. type b: patients with type b he have obvious pss and normal liver histology without associated intrinsic liver disease. these clinical manifestations are similar to those in patients with he and cirrhosis. the pss may be spontaneous or caused by surgical or interventional procedures [ ] . common causes of pss include congenital vascular malformation, intrahepatic or extrahepatic portal vein obstruction (including trauma, carcinoid, and bone marrow hyperplastic disease caused by a high coagulation state due to portal vein branch embolization and thrombosis) and generation of portal hypertension by oppression of lymphoma, metastatic tumors, and bile duct carcinoma. type c: type c he is related to chronic liver diseases, with cirrhosis being the most common type, and is generally accompanied by portal hypertension and pss. type c he is mainly caused by liver function failure, rather than by pss. according to the clinical manifestations, duration and characteristics, type c can be divided into three types: episodic he, persistent he, and minimal he [ ] . episodic he, related to chronic hepatic disease, is defined as a disturbance of consciousness and cognitive change in a short time and can be alleviated by spontaneous remission or drug treatment in the short term, which cannot be explained by a relevant preexisting mental disorder. episodic he can be divided into three types according to the presence of known risk factors: ( ) incentive type: there is a clear history of predisposing factors; ( ) spontaneous type: there is no history of predisposing factors. ( ) recurrent type: he attacks more than two times within a year. persistent he related to chronic hepatic disease is defined as an occurrence of continuous neural mental abnormality, including cognitive decline, disturbance of consciousness, coma and even death. persistent he can be further divided into three types according to the severity of the disturbance in the patient's self-control and self-discipline: , mildest type, namely west haven level ; , severe type: namely west haven level - ; and , therapeutic resistance type: medication can alleviate he quickly, but withdrawal can aggravate he rapidly. patients with minimal he, with normal clinical manifestations and routine biochemical tests, have mild cognitive and psychomotor deficits detected by neuropsychology and neural physiology tests, and these patients usually have a history of chronic hepatic disease [ ] . the prevalence of minimal he in patients with cirrhosis is - %. patients with minimal he with reduced physical and mental ability have gained more and more attention recently because they have a high risk of accidents when engaged in occupations involving mechanical, or driving work. the pathogenesis of he has not been fully elucidated so far, and many theories have been put forward. it is generally believed that he is caused by acute and chronic liver failure and/or pss. when toxic substances absorbed by the intestines cannot be detoxified and cleared by (or through) the liver, they directly enter into the systemic circulation and pass through the blood-brain barrier to reach the brain tissue and cause central nervous system dysfunction. a variety of the risk factors mentioned above can result in he. hyperammonemia is still recognized as one of the most important factors, especially in he related to chronic liver disease, liver cirrhosis and/or pss. according to the ammonia intoxication theory several factors including false neurotransmitters, such as γ-aminobutyric acid/benzodiazepine (gaba/bz) receptor complex, an imbalance in the ratio of branched chain amino acids to aromatic amino acids, brain cell edema, astrocyte dysfunction, mercaptan, short chain fatty acid toxicity and manganese deposition are all involved in the occurrence of he [ ] . ammonia intoxication caused by an ammonia metabolism disorder is the most important factor in the pathogenesis of he [ ] . ammonia comes mainly from the gut and the generation and absorption of ammonia increase in a serious liver disease when excess ammonia cannot be cleared sufficiently by ornithine cycle due to serious damage to liver parenchyma. when pss occurs, intestinal ammonia directly enters the systemic circulation without liver detoxification, resulting in increased blood ammonia. high levels of blood ammonia can enter the brain through the blood-brain barrier and generate central nervous system toxicity by interfering with cerebral energy metabolism, neurotransmitter and nerve cell membrane ion transport; increasing cerebral edema; and changing gene expression (such as stellate cell glutamate carrier, stellate cell structural protein, glial fibrillary acidic protein, peripheral benzodiazepine receptor and aquaporin- ) and inducing the mitochondrial permeability transition (mpt). the main way of removing ammonia from the brain is through urea cycle. during glutamine synthesis, glutamic acid is formed from ammonia and α-ketoglutaric acid and the glutamic acid combines with ammonia to generate glutamine. this process requires atp and consumes a large amount of α-ketoglutaric acid, which interferes with the brain energy metabolism and causes an energy supply shortage in brain cells. glutamate is an important excitatory neurotransmitter in the brain, and lack of glutamate increases inhibition in the brain. glutamine synthetase is present in astrocytes, where glutamic acid is detoxified to glutamine. glutamine is a strong intracellular osmotic agent, and increases in glutamine can lead to brain cell swelling. reports have identified a strong correlation between the content of glutamine in cerebrospinal fluid (csf) and the degree of he [ ] . during he, excess ammonia under the effect of glutamine synthetase, not only reduces the formation of active glutamate but also consumes a lot of energy, leading to the accumulation of glutamine, which increases intracellular osmotic pressure and causes brain cell swelling. swollen astrocytes with impaired function further affect ammonia metabolism, reduce the ability of neurons to efficiently uptake or release extracellular ions and neurotransmitters, and stimulate glial cell synthesis of neurosteroids by upregulating their expression of the peripheral-type bz receptor (translocator protein, kda). neurosteroid is an endogenous bz that can enhance gaba nerve tension and cause symptoms in patients with he [ ] (fig. . shown that the metabolic rate of cerebral ammonia in he patients is increased. increased levels of blood ammonia enter the brain through the blood-brain barrier. brain dysfunction also occurs even if blood ammonia levels appear normal; this partially explains the occurrence of he in the case of normal blood ammonia and invalidates he treatment by simply reducing blood ammonia. in addition, increasing evidence suggests a synergistic effect between blood ammonia and its metabolic disorders with systemic inflammation, nerve steroids, oxidative stress, nitrification stress, manganese poisoning, and gaba/bz [ ] . the main inhibitory neurotransmitter in the mammalian brain is gaba. plasma gaba is derived from the conversion of glutamic acid by glutamate decarboxylase in intestinal bacterial. notably, gaba has dual role. on one hand, during liver function failure and pss, the removal of gaba in liver is significantly decreased; on the other hand, gaba can directly enter the systemic circulation bypassing the liver, resulting in increased concentration of gaba in blood. the concentration of gaba in csf and brain tissue increases as more gaba crosses the abnormal blood-brain barrier. in addition, endogenous bz was found in the blood and csf, and the gaba receptor on the membrane surface of the brain's postsynaptic neurons increased significantly in some patients with he and in animal models. this receptor not only combines with gaba but also binds to barbiturates (barb) and bz on different parts of the receptor surface; thus, it has been named the gaba/bz complex receptor or the super receptor complex. when liver function is severely impaired, the binding affinity of this complex receptor to its three ligands is also increased. binding of gaba, barb, or bz with the complex receptor can promote entry of chloride ions from neuronal membrane ion channels into the cytoplasm of postsynaptic neurons, causing membrane hyperpolarization and nerve conduction inhibition. he symptoms were relieved in about % of patients treated with a gaba receptor antagonist or bz receptor antagonist, and gaba/bz and ammonia were reported to act synergistically in he. recently, some studies focused on peripheral type bz receptors, which are different from central gaba [ , ] . some questions, including the source of endogenous bz, and the correlation between the increased degree of gaba or bz and the disease, remain to be answered. therefore, therapy targeted at reducing the blood ammonia concentration in patients with he and significantly reducing the increased gaba nerve tension seems reasonable [ ] , but may not be completely effective. treatment effects of reducing ammonia vary, because of the different levels of ammonia in he patients that can be produced by the interaction between various known or unknown factors and the different effects of bz receptor antagonists. this theory is related to the metabolism of aromatic amino acids (aaa), the precursors of true neurotransmitters, including norepinephrine and dopamine. due to the reduction in the liver's detoxification function or formation of pss, the amines (phenylethylamine and tyramine) produced in the intestine cannot be cleared completely, resulting in elevated concentrations of these amines in the systemic circulation and increased levels in the brain through the blood-brain barrier. under the effect of β-hydroxylase, phenethanolamine and β-hydroxytyramine (β-dopamine) are generated from phenylethylamine and tyramine, respectively and are similar to norepinephrine and dopamine in chemical structure. these amines can be taken up, stored and released by adrenergic neurons in the brainstem reticular structure. phenethanolamine and β-hydroxytyramine are called false neurotransmitters because of their low physiological effects on the postsynaptic membrane, which is about / of norepinephrine. when these false neurotransmitters accumulate in the nerve synapse, they can outcompete or replace normal neurotransmitters, resulting in a disorder of nerve conduction. it was reported that plasma aaa (such as phenylalanine, tyrosine, and tryptophan) increased and branched-chain amino acids (bcaa, such as valine, leucine, isoleucine) decreased in patients with decompensated liver cirrhosis, leading to an imbalance of amino acid metabolism. aaa are decomposed and metabolized in the liver, and liver failure decreases aaa decomposition resulting in an elevated concentration of aaa in the plasma. insulin can promote bcaas entering muscle, which is then broken down and metabolized in the skeletal muscle instead of the liver. insulin inactivation is decreased in patients with liver failure, promoting a large number of bcaas entering the muscle tissue and decreasing the concentration of bcaas in plasma. finally, the bcaa/aaa ratio is reduced from a normal - . : to : or lower. the above process reduces the bcaa concentration, but increases the aaa concentration, leading to an increase in synthesis of false neurotransmitters and reduction of the normal neurotransmitter [ ] [ ] [ ] . the epidemiological data suggests that manganese poisoning and he extrapyramidal have common clinical symptoms. the liver is an important organ for manganese excretion. the concentration of blood manganese can be increased when liver function is affected, during pss, or when excretion of bile is reduced. manganese content in plasma was sharply increased in more than % of patients with acute hepatitis and liver cirrhosis and the density of globus pallidus increased in the brain basal ganglia of he patients (partially - times higher by mri). based on histological results, the above changes were caused by manganese deposition, which disappears after liver transplantation. it has been suggested that manganese deposition may cause dopamine dysfunction. deposition of manganese not only cause direct brain injury, it can influence the function of -hydroxytryptamine ( -ht), norepinephrine and gaba neurotransmitters; impair astrocyte function; and have a synergistic effect with ammonia. however, there is no reliable correlation between the concentration of serum manganese and he severity, which may be due to the chronic deposition of manganese [ ] . the characteristic change in mri imaging as the deposition of manganese remains to be verified. the effectiveness of manganese removal to improve the symptoms and neurological signs of patients with he needs further validation. the synergistic toxic effects between toxins (ammonia and mercaptan) and short chain fatty acids [ ] , the -ht hypothesis, the effect of helicobacter pylori urease, opioids, endotoxin, tumor necrosis factor, melatonin, and hepatitis b virus termed additional theories of he syndrome. this theory also suggests the same hypothesis mentioned in the above theories. due to the extensive amount of liver cell damage caused by acute liver failure in type a he, the residual liver cells cannot effectively remove toxins leading to central nervous system dysfunction. type a he, known as non-ammonia encephalopathy, is endogenous he without clear causative agents. simple type b he is rare in mainland china; the liver can clear limited metabolic toxins in patients with chronic liver failure or pss, but once these toxins exceed the compensatory capacity of the liver, type c he occurs. the occurrence of type c he is largely related to the following risk factors, which are the most important factors in the prevention and treatment of he. patients with chronic liver failure or pss are less tolerant to the protein found in food, especially animal protein. a large amount of ammonia and aaa are produced by the decomposition of intestinal bacteria, which can induce he. oral ammonium salts, urea, and methionine can induce he by increasing the absorption of nitrogenous substances and elevating blood ammonia. intestinal production of ammonia can be increased by hemorrhage in the intestine ( ml of blood contains - g protein). at the same time, because of the lack of isoleucine in the blood, after digestion and absorption of a hemorrhage, extra blood leucine and valine increase bcaa decomposition by enhancing the activity of bcaa dehydrogenase, thereby exacerbating the imbalance in the bcaa/aaa ratio. loss of blood volume, cerebral ischemia and hypoxia also increase the sensitivity of the central nervous system to ammonia and other toxic substances [ ] . infections such as spontaneous peritonitis, pneumonia, and urinary tract infection can increase tissue decomposition and production of ammonia. secondary sepsis or sirs induce he through tnf-α, il- , il- and other inflammatory factors, exacerbates oxidative stress, and increases the blood-brain barrier permeability of ammonia and other toxic molecules to liver and brain [ ] . studies have shown that sirs is directly related to the deterioration of he in patients with liver cirrhosis, and its extent and mortality increase with the deterioration of sirs [ ] . similarly, sirs is a common factor in triggering chronic liver failure characterized by he and renal failure. in a study of patients with liver cirrhosis, artificially-induced hyperammonemia by oral administration of glutamine may have worsened the results of psycho-mental testing in cases of sepsis patients; while brain toxicity was not obvious after the inflammation was relieved, the observation of decreased cytokine levels indicated that infection and induced inflammatory mediators enhanced brain toxicity of hyperammonemia. accordingly, some researchers suggested that sirs could be an independent pathogenesis of he rather than a risk factor [ ] . hyponatremia can affect the intracellular osmotic pressure and lead to brain edema, which induces he. hypokalemia is often associated with metabolic alkalosis [ ] . mass use of diuretics or extraction of ascites can also cause alkalosis. ammonia is easily absorbed by the intestinal tract or through the blood-brain barrier inducing he [ ] . a variety of reasons can cause pre-renal azotemia such as hypovolemia, anorexia, diarrhea, limiting the amount of liquid, mass use of diuretics, or extraction of ascites. hepatorenal syndrome or other causes can result in renal azotemia. pre-renal azotemia and renal azotemia caused by hepatorenal syndrome or other causes can increase the concentration of ammonia in the blood. several other predisposing factors can contribute to he such as constipation, hypoglycemia, the use of sedatives and proton pump inhibitors, and epilepsy. after the occurrence of constipation and intestinal obstruction, the patient's intestinal mucosa is exposed to ammonia longer thus increasing the absorption of ammonia. hypoglycemia can reduce brain deamination. the binding of sedatives, hypnotics and the brain gaba/bz receptor produce an inhibitory effect on the brain. it was reported that proton pump inhibitors increase the risk of he in patients with cirrhosis in a population study [ ] . another study also suggested that epilepsy was associated with an increased risk of he in patients with cirrhosis [ ] . patients with type a he often have no obvious anatomical abnormalities in their brains, but - % of patients have brain edema, which may be a secondary change of the disease. hypertrophy and hyperplasia of the original plasma astrocytes in gray matter and subcortical tissue can be found in patients with type c he. patients with longer course of the disease will exhibit brain atrophy (especially in patients with alcoholic cirrhosis) of different degrees, thinning of the cerebral cortex, loss of neurons and nerve fibers, and deep cortical sheet necrosis, even the cerebellum and the base may also be involved. the majority of patients with cirrhosis may have different degrees of he at some stage in the course of the disease. the incidence of he in patients with liver cirrhosis is at least - % in mainland china while the incidence of post-tips (transjugular intrahepatic portosystemic shunt) he is - %. if patients with chronic liver disease have he, the outcome is poor; the one year survival rate is lower than % and the year survival rate is less than % [ , ] . the incidence of mild he is . % in mainland china in patients with liver cirrhosis, . % in patients with child-pugh a, . % in patients with child-pugh b, and . % in patients with child-pugh c. the incidence of mild he is not significantly associated with cirrhosis; however, with the increased degree of decompensated liver cirrhosis, the incidence of mild he increase. several studies have found that the incidence of depression and anxiety in patients also increased, with the increase of liver function damage, the incidence also increased, and the outcome is poor [ , ] . the clinical manifestations of he vary, because of the difference in the nature of underlying disease, the degree of liver cell damage, the speed of injury and incentives. they are not specific to he compared with other metabolic encephalopathies. early pathological changes of he are mild he. the neuropsychological and intelligence tests detect mild form of he, which exhibit no clear clinical symptoms and often develop symptomatic he. the main clinical manifestations seen in acute liver failure induced by type a he are rapid-onset jaundice, bleeding, decrease in prothrombin, and eventually, change in mental status that can start as mild confusion but progress to coma and even death. type c he is characterized by chronic recurrent episodes of changes in personality and behavior [ ] , stupor and coma, which is often accompanied by increased muscle tone, hyperreflexia, hepatic flap, ankle clonus or positive babinski sign and nervous system abnormalities. most patients in the early stages relapse, but then their symptoms become persistent. he often has a variety of risk factors such as consuming a high-protein diet or discontinuing treatment of he. patients with type c he not only have the clinical manifestations of encephalopathy, they also have chronic liver injury, cirrhosis and other clinical manifestations [ ] . observation of encephalopathy dynamic changes is beneficial for early diagnosis, treatment and analysis of treatment efficacy. he can be graded and quantified according to the degree of disturbance of consciousness, nervous system performance and eeg changes. according to the edition of the "consensus on the diagnosis and treatment of hepatic encephalopathy" in china, he is divided into - periods, but each period can be overlapping or distinct but each period can be overlapping (table . ). at present, scholars have stressed that the occurrence of he is a continuous progression of the disease and should be viewed as a continuum of a wide range of neuropsychiatric abnormalities, rather than isolated clinical stages. according to the traditional west haven criteria diagnosing grade he is based on clinical signs and physician assessments, resulting in diagnostic criteria confusion [ ] . (fig. . ) . covert he is diagnosed by a variety of neuropsychological and intelligence tests; the evaluation of overt he widely uses the modified west haven semi-quantitative grading table for the analysis of patients with neuropsychiatric state (table . ), the glasgow coma scale for the analysis of the degree of consciousness of patients, and the simple he severity rating scale for the disease in addition to abnormal liver function (such as increased bilirubin, enzyme bile separation, and decreased prothrombin activity) commonly used auxiliary examinations for he diagnosis include: determination of ammonia, amino acid analysis of plasma and csf, psychological intelligence test, neurophysiological test, electroencephalogram and neuroimaging. the normal level of fasting venous ammonia is - μg/l (serum) or - μg/l (whole blood) and arterial ammonia concentration may be . - times that of venous ammonia. generally, the determination of arterial ammonia is common in clinical practice than intravenous determination; however, if venous blood has been transported on ice box and detected in a timely manner after proper collection, the result is expected to be as effective as arterial detection. ammonia levels are increased in type b and c he, but are normal in type a he. thus, he cannot be ruled out based on having a normal ammonia level. the increased level of ammonia was reported to be associated with the degree of type a he, but significant overlaps in different clinical stages of patients were also found [ , ] . therefore, ammonia detection is not routinely recommended in the diagnosis of he. notably, we need to rule out falsely elevated levels of ammonia caused by lab error, renal failure, complete parenteral nutrition, gastrointestinal bleeding, the use of steroid hormones and other extrahepatic factors. the fischer ratio (bcaa/aaa) is used as a marker of he, the plasma bcaa levels decrease while aaa levels increase; resulting bcaa/aaa: < (normal > ). it was reported that the concentration of glutamate in csf in he patients is increased compared to healthy controls. the concentrations of phenylalanine and tyramine in csf were also significantly increased, and the level of phenylalanine was closely related to the degree of he [ ] . recently, it was reported that h-nuclear magnetic resonance spectroscopy could select biomarkers for these diseases, such as in patients with he [ ] , but this is not commonly used clinically because the the characteristic manifestations of cognitive dysfunction in patients with covert he are lack of attention, working memory problems, and deficits in executive function. therefore, various intelligence tests are used to assess the subtle changes in a patient's cognitive or precise movement, which is important for the diagnosis of covert he, but not for overt he. [ ] . at present, computer-aided psychological tests such as information and communication technology (ict), cognitive drug research test (cdr), and critical flicker fusion test (cff) are not influenced by the factors mentioned above and easily operated, which can be used as an alternative choice for pen and paper tests. ict with sensitivity % and specificity % was one of the most commonly used tests to diagnose minimal he. cff was originally used to detect the critical flicker frequency of alert patients, reflecting brain conduction dysfunction. based on a spanish study of cases, including patients with cirrhosis and healthy controls, cff was a sensitive method to diagnose covert he with sensitive, simple and reliable advantages [ , ] . because the diagnosis of minimal he has just started, the related diagnostic value still needs to be further evaluated. an abnormal eeg is often observed before biochemical abnormalities or mental abnormalities [ ] . the main abnormalities by eeg are slowed rhythm, sporadic or universal θ wave ( - times/s) and the occasional α wave ( - times/s). with the deepening of consciousness, symmetrical δ-wave and three-phase waves appear on both sides simultaneously. this change usually occurs on both sides of the forehead and the top, gradually moving backwards. although these eeg changes are not specific to he and can appear in uremic encephalopathy and other metabolic encephalopathy, the severity of changes have a good correlation with clinical stages of he. computer analysis of eeg frequency distribution, such as artificial neural network-expert system (aness) and short epoch dominant activity cluster analysis (sedaca), is more objective and valuable in diagnosing minimal he than conventional eeg [ ] . there are many kinds of evoked potential tests, including visual evoked potential (vep), brainstem auditory evoked potential (baep), somatosensory evoked potential (ssep) and endogenous event evoked potential (event-related potentials, erps) p , of which the p is the most sensitive test for the diagnosis of he. compared with intelligence tests, neurophysiologic tests, independent of age and education background, are more objective. however, they are only used in clinical studies and are limited by equipment, and professional operation. based on cerebral ct and mri, brain edema can be found in patients with type a he while brain atrophy in the frontal cortex, and the t -weighted signal enhancement in the globus pallidus can also be found, which may be associated with manganese deposition. detected by h-magnetic resonance spectroscopy (h-mrs), the metabolic changes of he patients in the brain include increased levels of glutamate and glutamine, and decreased levels of inositol, taurine and choline [ ] . using fluid attenuation inversion recovery (flair) and diffusion weighted imaging (dwi) techniques, diffuse t -weighted signal enhancement is found in the hemisphere white matter and corticospinal tracts, which may be associated with cerebral ischemia. however, the sensitivity and specificity of the above-mentioned imaging abnormalities remain unknown, and the correlation with he staging is not clear. therefore, the main significance of cranial nerve imaging is to exclude cerebrovascular accident, intracranial tumors and other diseases, rather than diagnose he. there is no gold standard diagnostic criteria of he, and diagnosis is mainly based on the exclusion of other diseases. but one should consider the following five factors [ ]: several forms of hepatic diseases may lead to different kinds of he. type a he is caused by acute hepatic failure, but without chronic hepatic disease. type b is caused by pss, but without any history of hepatic diseases. type c is caused by serious hepatic diseases and/or widespread pss, such as cirrhosis, liver cancer, post-tips and so on. psychiatric symptoms can be found such as change of mood and personality, dementia, behavior disorder and disorientation. drowsiness alternating with excitability, hypermyotonia, asterixis, ankle clonus, insanity and coma are physical signs could be present in progressed patients. some patients may lack related physical signs and psychiatric symptoms, but have deficits in ability of learning, understanding, concentration, and quick verbal response. upper gastrointestinal hemorrhage, ascites tapping, excessive diuresis, high protein diet, medicine (such as sedatives) and infection could lead to he. previous he symptoms could be helpful for the diagnosis. type a usually does not have any risk factors. metabolic encephalopathy includes ketoacidosis, hypoglycemia, uremia, pulmonary encephalopathy, serious electrolyte disturbances and toxic encephalopathy. nervous system diseases include intracranial hemorrhage, infection or tumors, mental diseases and excessive use of sedatives [ ] . but one should also watch out for the coexistence of he in these situations. an overt he should be considered if ( ), ( ), ( ), and ( ) coexist; and covert he is based on ( ), ( ), ( ), and ( ) [ ] . then, based on the degree of neuropsychiatric symptoms, determine the stage of he, or for he classification refer to the west have semi-quantitative classification table or ishen scores. the flow chart of diagnosis is shown in fig. . . he is a complex metabolic disorder caused by many factors and comprehensive measures should be taken to cure it from different aspects. according to the clinical type, inducements and the severity of the disease, different plans of treatment should be designed for he. at present, the treatment of overt he generally includes the following aspects: ( ) supportive treatment; ( ) identification of possible concurrent encephalopathy and removal of other precipitants; ( ) cause of treatment; and ( ) empirical treatment (fig. . ). the point of nutritional therapy is to promote anabolism, inhibit catabolism, and maintain a positive nitrogen balance, rather than simply limiting protein intake. to reduce the source of ammonia, it has been suggested that patients with he should limit protein intake. in critically ill patients, it has been suggested that they should stop all protein intake and, after the disease improves, gradually increase protein intake to the maximum clinical tolerance. these recommendations are now being questioned because most cirrhotic patients are malnourished and all long-term protein-restricted diets increase the severity of malnutrition. in addition, a negative nitrogen balance increases mobilization of skeletal muscle, resulting in a reduction in ammonia metabolism that may increase blood ammonia levels. recent studies have shown that normal ingestion of protein . g/(kg • d) can also improve the health-related quality of life (hrqol), especially in mhe [ ] ; and have no adverse effects on the recovery of blood ammonia and he fig. . the flow chart of diagnosis of he compared with the restricted protein intake. according to the guidelines of the european society of enteral nutrition in , the intake of protein should be guided by the following principles: patients with acute phase he on the first day should be put on a prohibited protein diet and given glucose to ensure energy supply and those who cannot eat food may be fed through a nasogastric tube without short-term fasting; patients with chronic he do not need to fast and their intake of protein should be - . g/(kg • d); oral or intravenous use of bcaa and essential amino acid preparations can be administered to adjust the balance of aaa/bcaa, promote the balance of nitrogen, and also reduce the risk of he recurrence [ ] ; probiotics and prebiotics can enhance the body's tolerance to protein; plant protein is superior to animal protein because it contains methionine, has less aaa, and more bcaa, but it also contains cellulose, which is conducive to maintain the normal flora in the colon and acidize the intestinal tract, shortening the transit time of the colon and reducing absorption of ammonia. the above points need further verification. additional supportive treatments include: maintaining adequate hydration, electrolytes and acid-base balance; ensuring an energy supply of - kal/(kg • d), which should be composed of - % sugar, - % protein, and - % fat; administering appropriate vitamins and trace elements; treating for hypokalemia, hyperkalemia, hyponatremia, hypocalcemia, hypomagnesemia and metabolic alkalosis as needed; strengthening the basis of treatment with the appropriate infusion of fresh plasma or albumin, increased plasma colloid osmotic pressure; treating for hypoxemia and cerebral edema; and preventing and treating any bleeding and bacterial infection. type c he has a variety of precipitents. actively finding and eliminating triggers can effectively prevent the development of he, such as esophageal variceal bleeding that can develop into he. active hemostasis, anemia correction, and removal of intestinal blood are also conducive to controlling he. in addition, active control of infection, correction of water and electrolyte imbalance, elimination of constipation and improving renal function are essential to control he. anesthetics, painkillers, sedatives, sleeping pills, and other drugs should be strictly controlled. patients with mania or convulsions can reduce the use of diazepam and scopolamine, and the frequency of administration can also be reduced for promethazine, chlorpheniramine, and other antihistamines. toxic substances causing he mainly come from the intestine. thus, in order to prevent and control he, it is very important to clean the intestinal tract to reduce the generation and absorption of ammonia and other toxic substances. saline or weak acidic solution enemas (such as a dilute acetic acid solution), or oral or nasal feeding of % magnesium sulfate ( - ml) can be used to clear intestinal hemorrhage, intestinal impaction, and other toxic substances. an enema composed of non-absorbable lactulose ( - ml) plus water ( ml) is also useful, especially when applied for type b he. a recent clinical trial suggested that polyethylene glycol was more effective than the current standard first-line therapy in treating these patients [ ] . another two studies showed that polyethylene glycol was more effective than the standard lactulose therapy in treating patients with acute he by cirrhosis [ , ] . other available drugs include pear liquors, mannitol, rhubarb, and so on, but excessive use of these substances may lead to dehydration and aggravate he. non-absorbable disaccharides include lactulose and lactitol. lactulose, a kind of synthetic ketone disaccharide, cannot be broken down in the stomach and small intestine due to a lack of enzymes that can break down galactose in the digestive tract. after entering the colon, lactulose can be broken down into acetic acid and lactic acid with the help of gut bacteria, leading to a reduction in the colonic ph and inhibition of the absorption of ammonia in the intestine. these non-absorbent disaccharides are decomposed into organic particles in the intestinal tract, which can increase the osmotic pressure of the intestine, and their acidic products stimulate the intestinal wall and can slightly promote intestinal excretion. these non-absorbent disaccharides, acting as prebiotics in the intestine, can inhibit the growth of bacteria, which can produce ammonia and urea, finally reducing the production of ammonia and reversing low-grade cerebral edema when combined with rifaximin [ ] . however, probiotics can benefit patients in the long-term [ ] . oral or nasal feeding ( - ml, or times daily) was recommended to adjust the daily defecation appropriately, about - times daily. main adverse reactions include abdominal discomfort, abdominal distension, abdominal pain, loss of appetite, nausea, vomiting, and diarrhea. lactulose can even be used in patients with diabetes or lactose intolerance when the purity of non-absorbable disaccharide was high (≥ %), but is not used in patients with intestinal obstruction. numerous randomized controlled studies showed that lactulose or lactitol can significantly alleviate overt he and improve the patient's cognitive function and quality of life [ , ] . lactulose is still the first-line therapy of anti-he, although its effect on improving the survival rate of patients is uncertain. antimicrobial agents can be used as a substitute for non-absorbable disaccharides in treating acute and chronic he. in the past, oral aminoglycoside antibiotics, such as neomycin, which are rarely orally ingested, were used to inhibit the overgrowth of bacteria in the intestine. however, recent randomized placebo-controlled studies have shown that neomycin may not benefit patients with he compared with placebo-treated patients and that long-term use of neomycin may lead to increased ear and renal toxicity risk and impair the function of small intestinal mucosa [ ] . metronidazole can inhibit anaerobic bacteria in the intestine and alleviate he, but long-term use may lead to disruption in the intestinal flora, gastrointestinal discomfort, or neurotoxicity. rifaximin, a derivative of rifamycin with a broad-spectrum, has a potent inhibitory effect on intestinal bacterial growth, is a minimally-absorbed oral antibiotic and only plays a role in the gastrointestinal part. administration of rifaximin ( mg, twice a day) can significantly prevent the occurrence of he compared with placebo-treated patients [ ] [ ] [ ] ; rifaximin was equivalent to or better than lactulose and neomycin in treating patients with chronic he [ ] . a study indicated that rifaximin-α in combination with lactulose was a cost-effective therapy for patients who had experienced at least two prior overt he episodes [ ] , and this therapy could also improve the driving ability of patients with covert he without toxicity to the auditory nerve and renal function [ , ] . thus, rifaximin has been recommended by the us food and drug administration (fda) for the prevention of recurrent he. the efficacy of rifaximin and relation between longterm use of rifaximin and intestinal flora in the treatment of he needs to be further investigated. however, a recent study in mice showed that rifaximin beneficially alters intestinal ammonia generation by regulating intestinal glutaminase expression in mhe [ ] . the study may provide a new opportunity to study intestinal flora in the treatment of he. microecologics with bifidobacterium and lactobacillus can regulate intestinal flora structure to inhibit the growth of bacteria that produce ammonia and urease. in combination with prebiotics, microecologics can reduce the production and absorption of intestinal ammonia and other toxic substances [ , ] . in a recent openlabel study, patients with cirrhosis were randomized to three groups and treated with lactulose ( - ml daily), probiotic capsules, or with both drugs. after a month of treatment, patients with he showed better results in the neuropsychological test, p auditory evoked potentials, and blood ammonia. however, there was no difference in the therapeutic effect among the three groups [ , ] . clinicians commonly use sodium glutamate, potassium glutamate, arginine hydrochloride and potassium magnesium aspartate, but the exact efficacy is highly controversial at present and effective drug reduced ammonia is described below. (a) l-ornithine-l-aspartate lola, a dipeptide, can lower blood ammonia by promoting ammonia consumption and the synthesis of urea, glutamic acid and glutamine in brain, liver and kidney [ , ] . ornithine, a substrate of the ornithine urea cycle, can increase activity of carbamyl phosphate synthetase and ornithine carbamyl transferase, and promote urea synthesis. n-methyl-d-aspartate (nmda) is a substrate of glutamine synthesis, and the conversion of glutamic acid to glutamine in the body can remove blood ammonia [ ] . nmda is also involved in nucleic acid synthesis in liver cells and indirectly improves the metabolism of the krebs cycle process in liver cells to facilitate the repair of liver cells. clinical studies show that, compared with a placebo group, g/days lola intravenously could noticeably reduce fasting blood ammonia (fnh ), postprandial blood ammonia, and improve the mental status of patients with he [ ] . patients with oral lola also had improved he examination results for the digital connection test, the flapping tremor, and eeg results [ ] . in addition, glycerol phenylbutyrate (gpb) can safely reduce the incidence of he as well as ammonia in patients with cirrhosis and he. the results showed that gpb had therapeutic potential in this population [ ] . zinc is an important cofactor in urea cycle enzyme catalysis. a study in he patients showed that serum zinc concentration is reduced, and showed a negative correlation with the blood ammonia concentration; the serum ammonia level is much lower after zinc supplementation in patients, and he can be improved in some patients. a new study suggests that antioxidant and zinc supplementation can improve mhe in patients with liver cirrhosis [ ] . oral zinc preparation can also reduce absorption of divalent cations such as manganese in the intestine; however, it has not been determined if zinc has a positive therapeutic effect on he. (c) sodium benzoate sodium benzoate can lower the blood ammonia concentration by activating the urea cycle for ammonia detoxification and promoting urinary ammonia. randomized controlled studies showed that sodium benzoate had the same efficacy as lactulose in treating patients with he. the recommended sodium benzoate dose is g twice a day; nevertheless, few patients can tolerate this dose because of its high gastrointestinal side effects [ ] . a recent study showed that tranilast could protect patients from thioacetamide-induced acute liver injury and alleviate he [ ] . endogenous bz analogues combine with the inhibitory neurotransmitter gaba receptor to depress the action on the cns, and is one of the occurring hallmarks of he pathogenesis. a large-scale clinical study on he cases showed that the improvement rate in brain function in treatment and control groups were % and %, respectively [ ] . the study showed that treatment of he with receptor antagonists such as fluorine marcie is feasible. a meta-analysis which included casecontrol studies of patients show that fluorine marcie can noticeably improve he, but didn't show any long-term benefits or improve patient survival rate. so fluorine marcie should only be considered for he patients who had used bz. although the reduction of dopamine neurotransmitter activity is also one of the pathogenesis, the application of bromocriptine, levodopa, has been unable to bring more benefits besides partly improving symptoms of patients. oral or intravenous infusion with a bcaa-based amino acid mixture can theoretically correct an imbalance in amino acid metabolism and control false neurotransmitter formation in the brain [ ] . a meta-analysis which included five studies showed that intravenous bcaa did not reduce the mortality rate of he. three studies with bcaa did not reduce the mortality rate of he; however, two larger studies (randomized controlled study about patients with liver cirrhosis in cases and cases, respectively) show that the application of bcaa not only reduced the occurrence of he and liver failure, but also improved the nutritional status, liver function and survival rate in patients. another study showed that bcaa could stimulate liver cell regeneration thus reducing the occurrence of liver failure. supplementation with a bcaa-rich amino acid mixture showed improved restoration of the patients' positive nitrogen balance, and increased the patient's susceptibility to protein food, improving cerebral perfusion [ ] . considerable progress has been made to understand treatment of he, studies of basic and clinical research are underway using newly discovered treatment strategies, such as toll-like receptor antagonists (with the ability to reduce systemic inflammation and oxidative stress) as well as non-steroidal anti-inflammatory drugs (ibuprofen) [ , ] , nmda antagonists, anticholinesterase. however, research using gene therapy should not be ignored [ ] . after tips, lola can significantly reduce the increase of venous ammonia concentration in patients with he [ ] . and the positive dietary intervention can significantly reduce the incidence of he [ ] . for patients with refractory he, embolization of pss is a safe and effective treatment strategy [ ] . improving liver function antiviral treatment with nucleos(t)ide analogues can reduce or eliminate liver inflammation and necrosis, promote the regeneration of liver cells, and help restore the functions of hepatic metabolism and detoxification in chronic liver failure caused by hepatitis b virus. artificial liver support systems can be divided into three types including nonbiological type, biological type and mixed type. the non-biological liver support system is the most widely used type, and consists of hemodialysis, hemofiltration, plasma exchange, blood perfusion, plasma adsorption, and the molecular adsorption recirculation system (mars) [ ] . the artificial liver support system can replace the partial function of the liver, remove the poison accumulated in the body, create conditions that allow for the regeneration of liver cells and provide enough time to wait for liver transplantation for patients with he. an artificial liver support system can be used to treat acute and chronic he, but patients with overt stage he should be especially careful with plasma exchange. liver transplantation remains the only promising therapy for patients with an acute liver failure or endstage liver disease. liver transplantation is an effective means for all kinds of persistent and severe he; however, in patients with he there is a significant increase in mortality among patients awaiting liver transplantation [ ] . recently, it has been reported that cognitive function was not fully recovered after liver transplantation in some patients with severe he [ ] . the key point in improving the prognosis of he is early recognition and timely treatment. active treatment should be given when diagnosing covert he. theoretically, for patients with serious pss, interventional therapy, surgery or permanently/temporarily and partially/totally blocking the pss can improve the patient's symptoms. the use of this therapy should be carefully weighed because it can increase the risk of gastrointestinal bleeding in case of portal hypertension. covert he has gained an increasing amount of attention in recent years. patients with covert he do not exhibit obvious signs and symptoms; however, their quality of life is reduced because of reduced operational ability or sleep disorders. without treatment, covert he will progress to overt he over time. the population with high risk should be examined and treated early, especially those engaged in potentially dangerous occupations. the following solutions can be referred to: (a) adjusting dietary structure (vegetable protein is the main intake); (b) oral administration of lactulose ( - ml, - times daily); (c) oral administration of rifaximin ( mg, twice a day); (d) oral administration of lola ( g, times a day); (e) oral administration of baaa; and (f) oral administration of probiotic preparations [ , , ]. although medical technology has made great progress and the research into he is also increasing in recent years, the pathogenesis of he is still unclear. due to a lack of specific methods, combination treatment is still the main therapy for he. it is generally believed that the onset of he may be a result of the synergistic effects of many factors. therefore, it is difficult to implement and draw convincing conclusions from randomized controlled trials with a single intervention targeting a specific pathogenesis and risk factor. ongoing issues remain, such as standardizing the research design of he treatment and evaluating the efficacy of he treatment more scientifically and objectively. some clinical studies may bring new hope for he treatment by new ongoing strategies of targeted systemic inflammation, oxidative stress, and neurosteroids. in addition, the key point in improving the prognosis of he is early recognition and timely treatment. active treatment should be given when diagnosing covert he. it is difficult to popularize the cognitive dysfunction detection methods of latent he. so the key and difficult point is to develop new method of assessments for clinicians in the future. jia shang hepatopulmonary syndrome (hps) is a syndrome of shortness of breath and hypoxemia induced by vasodilation in the lungs of patients with a variety of acute and chronic liver disease. essentially primary liver disease, pulmonary vasodilation and arterial oxygen lack of co-triad constituted. due to abnormal increase of vasodilators、ventilation/ blood flow disproportion and pulmonary hypertension caused by liver disease, the hypoxemia (pao < . kpa) ( mmhg) is included in hps. when fluckiger reported a -year-old syphilis female patient as early as in , he described cirrhosis, cyanosis and clubbing at the same time, while he was not aware of the intrinsic relationship between these clinical manifestations [ ] . in , snell reported decreased arterial oxygen saturation (sao , less than %) with abnormal hemoglobin in patients with liver parenchymal lesions and biliary obstruction, and years later, he proposed that such a phenomenon was associated with decreased affinity of o with hemoglobin. in , rydell and hoffbauer reported the detailed clinical diagnostic and treatment process of a -year-old male with "juvenile cirrhosis", and found multiple arterial-venous anastomoses in the lungs during autopsy, which he thought contributed to clinical cyanosis mainly. this provided a histological basis for the patient, and people conducted a large amount of studies thereafter. in , berthelot et al. injected opaque glue into the pulmonary vascular beds at the time of biopsy after the patient's death for the first time, and he found abnormal small arterial dilation in the lungs of the patient with cirrhosis, which he termed lung spider nevus. the term hepatopulmonary syndrome (hps) was first proposed by kennedy and knudson in [ ] . after nearly years of studies in a large number, people gradually developed a clear understanding of the mechanisms underlying its pathogenesis. in , eriksson used the term functional hepatopulmonary syndrome for the first time. in , the famous liver disease expert sherlock formally used this diagnostic term in his monograph hepatobiliary diseases, which has been recognized by many scholars [ ] . hps can occur in patients of any age groups, and various literature reports show conflicting incidences of hps in patients with cirrhotic portal hypertension, with the average incidence of various chronic liver diseases being about - %. the incidence of cirrhosis in patients is high, and - % of patients can additionally develop mild arterial hypoxia and - % develop arterial hypoxia. in the study by binay on indian cirrhotic populations arising from hepatitis b mainly, the incidence of this disease is relatively low ( . %) [ ] . the differences in incidence were mainly attributable to the different diagnostic criteria adopted. schenk et al. studied the incidence of hps by performing transthoracic contrast echocardiography (ttce), pulmonary function tests and blood gas analysis on patients with cirrhosis patients. the results showed that the incidence of hps patients in whom alveolar-arterial partial pressure of oxygen (aapo ) was used as an indicator of hypoxemia was significantly higher than those in whom arterial partial pressure of oxygen (pao ) was used [ ] . when arterial partial pressure of oxygen (pao ) was reduced to reflect hypoxemia, hps incidence was % when < mmhg and % when < mmhg, respectively. while when increase in alveolar-arterial partial pressure of oxygen (aapo ) was used to reflect hypoxemia, the incidence of hps was high, with % when > mmhg and % when > mmhg, respectively. hps is most common in cirrhosis due to various causes. pulmonary vascular abnormalities and arterial hypoxemia can occur in a variety of acute and chronic liver diseases, and this is true mainly when it comes to cirrhotic patients due to chronic liver diseases, especially cryptogenic liver cirrhosis, alcoholic cirrhosis, hepatitisinduced cirrhosis and primary biliary cirrhosis. besides, hps can also occur in chronic hepatitis, acute severe hepatitis, cholestasis, ɑ-anti-trypsin deficiency [ ] , tyrosinemia, wilson disease, and non-cirrhotic portal hypertension (such as idiopathic portal hypertension and schistosomiasis cirrhosis, etc.). arterial hypoxemia can also occur in extrahepatic portal vein occlusion. the observation of these patients suggests that portal hypertension may be the main factor for the pathogenesis of hps. hps can also occur in non-cirrhotic portal hypertension, and even cirrhosis-and portal hypertension-free chronic viral hepatitis. in , binay et al. found that patients with progressive liver failure with hyperdynamic circulation are most likely to suffer from hps, while they did not find the correlation with the severity of liver cirrhosis. hps is, in essence, hypoxemia due to anomaly in pulmonary vascular dilatation and arterial oxygenation when liver disease occurs. arterial hypoxemia occurs as the result of insufficient oxygenation by blood cells in the blood when blood flows through the lungs, or a proportion of blood fail to flow through the alveoli [ ] . since primary heart and lung diseases have been excluded when hps occurs, the abnormal pathways that red cells may pass through include: ( ) passing through the pleural and hilar bronchial vessels while not reaching the alveoli; ( ) blood flows directly into the pulmonary veins due to the high pressure portal system in the mediastinum, thereby bypassing the pulmonary circulation; ( ) flowing directly into the pulmonary veins through the expanded alveolar capillaries or the pulmonary-venous fistula. alveolar telangiectasia may be more important to the formation of hypoxemia, and existing study data show that the development of hps is at least associated with the systemic hyperdynamic state, portal hypertension, hepatic encephalopathy, hepatorenal syndrome and pulmonary hypertension [ ] . therefore, it is believed that the main causes of hps are systemic metabolism and hemodynamic disorders, and that it is involved in the formation of systemic metabolism and hemodynamic disorders, which is of important pathophysiological significance. . the basic pathological change of hps is pulmonary vascular dilatation, which is manifested as: (a) dilation of anterior capillaries in a large number. (b) formation and opening of the pulmonary basilar arterial -venous communicating branches. (c) formation of pleural "spider mole", which is mainly manifested as dilation of anterior capillaries. in autopsies, it was found that the basic pathological changes in patients with liver cirrhosis and other chronic liver diseases were extensive pulmonary vascular dilatation and arteriovenous communicating branches. some people found the pathological changes through vascular shaping, with pleural vasodilation at the basal aspect of the lungs or the formation of subpleural spider nevus. domestic professor gu changhai summarized these pathologic changes in as arterial dilation within the pulmonary acinus in a pattern of inhomogeneous distribution, thin-walled blood vessels, - μm in diameter, in the lower lobes of the whole lungs, extensive dilation of pulmonary vascular beds adjacent to the alveolar gas at the anterior capillary level, and significantly expanded pulmonary artery branches and pulmonary capillaries up to μm in diameter. electron microscopy showed thickened pulmonary capillaries, pulmonary arterial walls and the basal layers of small veins. . factors that affect the dilation of blood vessels: the mechanisms underlying pulmonary vascular dilatation have not yet fully elucidated, and the possible influencing factors include: (a) increased activity of vascular dilators various acute and chronic liver diseases, liver cell failure and metabolic disorders, particularly reduced inactivation of vasoactive substances which can enter directly into the systemic circulation through abnormal anastomotic collateral vessels, result in disorder of the systemic hemodynamics and increased contents of vasodilators in the blood circulation. just as visceral congestion in patients with portal hypertension, they can act on the intrapulmonary vessels, causing pulmonary vascular dilatation and pulmonary congestion. substances that cause vasodilation include glucagon, prostaglandin, vasoactive intestinal peptide, nitric oxide, angiotensin, bradykinin and endotoxin, etc. (b) reduced vasoconstrictors or decreased sensitivity of intrapulmonary vascular beds to the endogenous vasoconstrictors, such as norepinephrine, endothelin, atrial natriuretic peptide, vasopressin, serotonin and tyrosine, etc. the contents of the substances are not absolutely reduced because maybe their sensitivity is reduced. when chronic liver disease occurs, the anterior communicating branches of the originally closed non-functional capillaries may be opened, and a disorder occurs in the hypoxic pulmonary vascular systolic dysfunction which should have been normal, and it is only % of the normal state [ ] . (c) neurological factors cirrhotic patients show sympathetic nerve hyperactivity, but after the formation of portal hypertension, their sympathetic nerve function may be damaged, which play an important role. animals with portal hypertension often show abnormal pressure responses and reduced sensitivity of blood vessels to norepinephrine, resulting in increased cardiac output, and dilated pulmonary vascular volumes. besides, hemodynamics within the lungs is also a manifestation of the body's hyperdynamics. (d) decreased reactivity of intrapulmonary blood vessels to hypoxia recent inert gas dispersion tests show that cirrhotic patients with over two spider nevus are manifested as not only liver damage, but also decreased systemic intrapulmonary vascular resistance, decreased reactivity of blood vessels to hypoxia and dilated pulmonary vessels. however, it was also found using pulmonary angiography that in spite of the dilated vessels at the ending of arteries, the responses of vessels to oxygen were almost normal, which did not support this view. (e) intrahepatic angiogenesis or dysplasia may also be one of the factors for the formation of hps. to date, the mechanisms underlying pulmonary vascular dilatation caused by hps is still unclear. however, long-term administration of intrapulmonary vasoactive substances can cause significantly increased intracellular cyclic adenosine monophosphate (camp) and/or cyclic guanosine monophosphate (cgmp), resulting in hypoxic pulmonary vasomotor dysfunction and pulmonary artery dilatation, which may be an important cause of this disease and also pulmonary manifestations of systemic hyperdynamic circulation. due to the significant dilation of the pulmonary capillaries and the anterior capillaries, some of the blood around the capillaries in contact with the alveoli can still undergo exchanges with gases, while the central blood, due to the increased diffusion distance from the alveoli, leads to insufficient gas exchange, resulting in insufficient arterial oxygenation and thus a series of hypoxemic manifestations. to date, the pathogenesis underlying the pathogenesis of hps has not yet been elucidated. in view of the above pathophysiological changes and current studies, it is believed that the disease may be caused by insufficient ventilation, diffusion disorder, ventilation/blood flow imbalance and decreased oxygenated hemoglobin affinity, or the above factors in combination. under normal circumstances, insufficient ventilation due to a variety of reasons causes insufficient oxygen inhaled into the alveoli and reduced blood oxygen exchanges, which can result in hypoxemia [ ] , such as chronic bronchitis, foreign bodies in trachea, atelectasis and respiratory muscular paralysis, etc. and the presence of insufficient ventilation in patients with chronic liver disease and cirrhosis or not is still controversial. in , fujiwara studied the lung function in patients with decompensated liver cirrhosis and reported that vital capacity (vc), functional residual capacity (frc) and respiratory reserve volume (evr) in the patients were significantly reduced, that r/t was mildly increased, and that there was no changes in s forced expiratory volume (fev ). therefore, it was believed that mechanical compression and insufficient ventilation due to pulmonary interstitial edema in patients with liver cirrhosis was the main reason for impaired lung function. subsequently, edison et al. studied the pulmonary function of patients with decompensated liver cirrhosis, and found that their vc, maximum ventilation volume (mvv), frc, total lung volume, and r/t were significantly reduced, and they believed that patients with cirrhosis had obvious obstructive and limited insufficient ventilation, which were mainly caused by compression of lung tissue due to increased abdominal pressure, elevated diaphragm and increased chest volume and pressure when patients had additional ascites, and atelectasis [ ] . however, decreased fev resulted from compression of small trachea due to pulmonary interstitial edema and vasodilation, and early closure of expiration. theoretically, all of the above factors can lead to insufficient ventilation, one of the factors resulting in this disease. this was also substantiated by significantly increased arterial partial pressure of oxygen and decreased co partial pressure in cirrhotic patients with pleural effusion after pleural effusion extraction and recovery from atelectasis. however, there are also some people who do not think that hypoxemia results from insufficient ventilation, but because cirrhotic patients are not complicated by high concentrations of co when their arterial partial pressure of oxygen is decreased [ ] . this is likely because when patients have hypoxemia, compensation of hyperventilation causes arterial blood co partial pressure not to increase, and results in decreased paco or even respiratory alkalosis. besides, in some patients without decompensated liver cirrhosis, arterial hypoxemia can also occur. even it has been found that the lung function tests in patients with decompensated liver cirrhosis are normal. therefore, the majority of scholars currently believe that insufficient ventilation is not the main cause of hypoxemia in cirrhotic patients. for patients with hps, the inert gas exclusion technique should be performed to prove that there is a disorder in the diffusion of oxygen, which is determined by the basic pathological changes of hps -pulmonary vasodilatation. pulmonary angiography can show small spider-like to obviously cavernous diffuse vasodilation within the lungs. due to the significant dilation of the pulmonary capillaries and the anterior capillaries, the diffusion distance of the blood flow in central blood vessels and the alveoli is increased, preventing the gases in the alveoli from entering the pulmonary capillaries, thereby affecting the gas exchanges. studies have shown that hypoxemia often occurs in patients with cirrhosis or aggravates during exercises, and it is believed that diffusion disorder or limitation of oxygen occurs in patients. in fact, factors that affect o diffusion do occur in patients with cirrhosis, but they are still not sufficient to explain the apparent hypoxemia. although vascular dilatation occurs at arterial endings in patients with hps, their arterial partial pressure of oxygen can be reduced while inhaling air and increased when they are given oxygen inhalation, which further proves that although diffusion disorder does exist and it plays a role in the formation of this disease, the role is not important. to engage in gas exchanges is the most important biological function of lung tissues, and this gas exchange must be completed when there is an appropriate ventilation/blood flow ratio. under the normal circumstance (normal adult resting state), the most appropriate ventilation/blood flow ratio physiologically is . . changes in the ratio due to any cause can affect the gas exchange, and the imbalanced ventilation/blood flow ratio in hps patients with hypoxemia is mainly because of pulmonary vascular dilatation and arteriovenous shunt [ ] . . intravascular vascular dilatation: pulmonary vascular dilatation has been confirmed pathologically and by angiography. dilated blood vessels in the lungs leads to gas diffusion disorder. besides, since the oxygen molecules in the air can not be diffused to the central dilated blood for the gas exchange, causing decreased ventilation/blood flow ratio and pulmonary arterial partial pressure of oxygen. this decreased ventilation/blood flow, together with increased amount of reactive cardiac output, shortens the duration of blood that flows through the capillary network and insufficient oxygenation [ ] . excessive ventilation can in part enhance patients' pao . if the alveolar oxygen partial pressure is increased at this time, some oxygen molecules can reach the central areas of dilated blood vessels, increasing the arterial partial pressure of oxygen. therefore, it is called the diffusion -perfusion disorder or pulmonary arteriovenous functional shunt rather than the true lung shunt. . arterial -venous shunt: intrapulmonary vascular fistula and pleural spider nevus can occur in cirrhosis of the liver and allow the pulmonary arterial blood to circumvent gas exchange and directly flow into the pulmonary vein so that patients may develop hypoxemia. this hypoxemia can not be corrected by oxygen inhalation and represents the true pulmonary shunt, which has been confirmed by pulmonary histopathology, angiography, transthoracic echocardiography and other examinations. it is now believed that pulmonary vascular casting is still the most direct evidence for determining the arterialvenous shunt. this intrapulmonary arterial -venous shunt is the main cause of abnormal ventilation/blood flow ratio and insufficient gas exchange. although pleural spider nevus can also cause arterial -venous shunt, it generally does not suffice to cause significant hypoxemia due to the small amount of shunt. in addition, studies in recent years also show that portal-pulmonary vein shunt in a small amount occurs in some patients with cirrhosis, in whom blood flow circumvents the alveolar gas exchange and enters directly the systemic circulation. this can also cause ventilation/blood flow abnormalities, causing insufficient gas exchange . airway closure: in , ruff et al. proved that cirrhotic patients had significantly increased closed volume (cv) and total amount of closed gas (cc) and increased gases trapped in the lower fields of the lungs, resulting in an extremely low ratio of ventilation/blood flow, and they believed these were due to reduced airway ventilation [ , ] . in , furukawa et al. measured the lung function of patients with liver cirrhosis and did not find abnormalities; however, most patients had flow -volume abnormalities and significantly increased cv, suggesting the closed the airway in advance and decreased ratio of ventilation/blood flow, which might important causes of hypoxemia. . decreased affinity of oxygen with hemoglobin: some reports showed that patients with cirrhosis (mostly alcoholic cirrhosis) patients had mild systemic vascular or pulmonary vascular dilatation, normal pao , mild hypocapnia, mild right shift of the oxygenated hemoglobin dissociation curve, normal amount of carbon monoxide diffusion [ ] , and mild imbalance of the ventilation/pao blood flow, indicating that the right shift of the oxygen dissociation curve due to decreased affinity of oxygen with hemoglobin in the patients. this was possibly caused by the increased concentration of , -diphosphate glyceride in red blood cells, which, however, is not an important factor in the occurrence of hypoxemia [ ] . in summary, hypoxemia can result from many factors, while none of the factors can completely explain the pathogenesis underlying the disease. since the basic pathological changes in patients with hps are intrapulmonary vascular dilatation and opening of arterial -venous communicating branches, together with recent findings, it is suggested that the diffusion disorder of the alveoli and pulmonary capillaries and ventilation/blood flow imbalance may coexist, and are the main cause of hypoxemia in this disease. other factors may aggravate hypoxia and are secondary factors. therefore, it is believed that the disease occurs as result of the above factors. the pathological features of hps are dilation of capillaries in the anterior aspect of the lungs and telangiectasia. autopsies show arterial-venous short circuit within the lungs, vasodilation and thickened pulmonary muscles [ ] . at the same time, arterial hypoxemia is common in liver diseases, often attributable to a variety of factors (such as ascites, hepatic pleural effusion, and copd in patients with alcoholism); it shows unique pathophysiological characteristics under specific circumstances of hps. its prominent features are dilation of micro-arteries in the anterior aspect of pulmonary capillaries and true capillaries (the normal diameter of these vessels is to μm, which can reach - μm when patients rest), with the number of dilated vessels increased macroscopically. some patients show arterial-venous communicating between the pleura and lungs, vascular anastomosis in the liver and the lungs, and thickened walls of small veins and capillaries. pulmonary vascular dilatation is promoted, and mixed venous blood quickly or directly enter the pulmonary veins through the anastomosis in the lungs, leading to oxygenation defects. increased nitric oxide (no) is a key cause for pulmonary vasodilation, and whether other mediators, such as heme oxygenase-derived carbon monoxide, are causes of pulmonary vasodilatation are not yet confirmed. abnormal arterial oxygenation seriously affects the survival of patients, and is an important indicator that determines the timing and risk of liver transplantation as well as an important basis for the grading of severity of hps. causes of deaths associated with hps are often multifactorial and are associated with basic liver diseases, and there are few cases of respiratory failure due to severe hypoxemia. hps is a triad composed by intrapulmonary vascular dilatation and insufficient arterial oxygenation due to primary liver disease, and it is mainly clinically manifested as primary liver disease and pulmonary lesions. hps can occur in various liver diseases, mostly in chronic liver disease, especially cirrhosis caused by various causes, such as cryptogenic cirrhosis, alcoholic cirrhosis, liver cirrhosis, viral cirrhosis, postnecrotic cirrhosis and biliary liver cirrhosis, etc. the most common clinical manifestations include liver palms [ ] , spider nevus, jaundice, ascites, hepatosplenomegaly, gastrointestinal bleeding and abnormal liver function, etc., while they are not significantly correlated with hps. some patients with clinically stable liver disease may also develop the clinical manifestation of progressive pulmonary insufficiency. since hps patients have no primary cardiopulmonary diseases, most ( - %) patients gradually develop respiratory manifestations on the basis of various liver diseases, such as cyanosis, dyspnea, clubbing, orthodeoxidation and platypnea, etc. among them, progressive dyspnea is the most common lung symptoms of hps. binay et al. believed that cyanosis was the only reliable clinical sign, and that platypnea and orthostatic hypoxia are the most characteristic manifestations. pulmonary examinations generally show no obvious positive signs [ ] . a small number of patients (about - %) can present complaining dyspnea on exertion in the absence of clinical manifestations of a variety of liver diseases, to which attention should be paid clinically so as to prevent misdiagnosis. the domestic researchers gao zhi et al. reported that patients presented to hospitals with cyanosis, palpitation after exercises and shortness of breath; meanwhile, they found that the patients had clinical manifestations of liver cirrhosis [ ] (such as liver palms, spider nevus, hepatosplenomegaly and ascites), which were conducive to the diagnosis of this disease. if liver disease patients have other lung diseases (such as chronic bronchitis, emphysema and pneumonia, and pleural effusion, etc.), then significant respiratory symptoms may occur. therefore, differential diagnosis should be made. data show that the duration of initial dyspnea to the conformed diagnosis in patients with hps average - years; that is to say, about % of patients already have dyspnea at the time confirmed diagnosis. . orthodeoxidation: pao is decreased by > % when patients switch from the supine position to the standing position. . platypnea: when patients switch from the supine position to the standing position, they have palpation, chest tightness and shortness of breath, and when patients resume the supine position, the above symptoms are improved [ ] . krowka reported that about - % of patients with hps had the above two manifestations because vascular dilatation in the patients was mainly distributed in the middle and low lung fields. when patients switch from the supine position to the standing position, the blood flow in the middle and lower lobes of the lungs is increased under the action of gravity, aggravating hypoxemia [ ] . although the two manifestations are not unique to hps, they suggest the significant abnormality in the patients' pulmonary vascular system. if patients with a variety of liver diseases present with the above two manifestations, further examinations are needed for confirmation. patients may present with liver palms, hepatosplenomegaly, spider nevus and ascites; patients show palpitation, chest tightness, shortness of breath when switching from the supine position to the standing position due to hypoxemia. schenk et al. defined the values of pao for the diagnosis of hps, thinking that pao < mmhg suggested a high possibility of hps, and that for pao < mmhg, the diagnosis of hps could be made [ ] . pulmonary function tests mainly showed significantly decreased vc, frc, mvv and fev , but sometimes the total lung volume and fev were normal. chest x-ray, ct scan and transthoracic contrast echocardiography (ttce) hps patients are mostly normal on chest radiography or show diffuse small millet shadows predominantly in both lower lobes of the lungs, nodular shadows in both lower lung interstitium, dilated pulmonary arterial trunks, and thickened pulmonary markings, whereas these manifestations have no specific values to the diagnosis of this disease. ct scan shows certain diagnostic values in that it demonstrates distal vasodilation and even pleural blood vessels, and can suggest the presence of hps. arteriovenous communicating occurs in hps patients due to pulmonary vascular dilation, indicating that subclinical pulmonary vascular dilatation and abnormal gas exchanges occur in cirrhotic patients with normal pao [ ] . contrast echocardiography: when hps is suspected, transthoracic echocardiography can be used as a preliminary screening to determine whether the intrapulmonary vascular dilation occurs or not. the microbubble contrast material in the right atrium, after intravenous injection of dioxane isotonic saline, will develop images in the left atrium through the dilated vascular beds after - cardiac cycles, while the microbubbles cannot pass through the normal capillaries (normal capillaries are < - μm in diameter). approximately % of patients with cirrhosis had positive changes on contrast echocardiography, while only a small proportion of patients are in line with the diagnosis of hps due to the influence of dilated blood vessels. if contrast echocardiography is positive for liver cirrhosis or portal hypertension patients with hypoxemia and cardiopulmonary diseases in them can be ruled out, then the diagnosis of hps is established. this means is used to confirm the diagnosis of intrapulmonary vasodilatation. the pulmonary vascular abnormalities in hps patients are as follows: ( ) diffuse spider nevus images. patients of this type have severe hypoxemia and erectile hypoxia and respond well to inhalation of % oxygen; ( ) cavernous or spotted arterial dilatation mainly seen in the basal aspect of the lungs. patients during this period respond poorly to % oxygen; and ( ) intermittent local arterial malformation or communicating branches, isolated earthworm-like or bulk images. in addition to severe hypoxemia and erect hypoxia, patients of this this type respond extremely poorly to the inhalation of % oxygen. when hypoxemia is caused by hps and cardiopulmonary diseases concurrently, cm tc-maa scan can determine hypoxemia resulting from hps more likely. radiolabeled albumin cm tc, which is administered through intravenous injection, is about pm in diameter. when pulmonary vascular shunt occurs, a proportion of the polymerized albumin passes through the lungs and enters the systemic circulation, the intake of albumin by other organs can be simultaneously determined by scintigraphy [ ] . therefore, the amount of shunt can be calculated. a study showed that cm tc-maa scan was positive for hps patients with pa < mm hg, while the scan was negative for chronic obstructive pulmonary disease (copd) patients with the same degree of hypoxemia, a result indicative of the good specificity of this means. compared with contrast echocardiography, cm tc-maa scan, in spite of its low sensitivity, can be used for the diagnosis of hps in patients with copd. pathological examinations are the most reliable means for the diagnosis of hps, whose basic pathological change is pulmonary vasodilation manifested as diffuse anterior capillary dilation or discontinuous formation of arteriovenous branches [ ] . in addition, pulmonary perfusion scan and right cardiac catheterization are also valuable for the diagnosis of hps to a certain extent. there is no unified standard for hps diagnosis to date. diagnosis should be based on clinical manifestations plus imaging evidence of pulmonary angiography. (c) the domestic scholars gao zhi et al. thought in that the diagnosis of this disease should be based on the following manifestations in patients, hepatosplenomegaly, ascites, liver palms, spider nevus, dyspnea on exertion, hypoxia while breathing in the supine and orthostatic positions, increased mesenchyma in the basal aspects of the lungs and vascular markings on chest radiography, patchy or nodular shadows, dilated basilar pulmonary vessels and increased pulmonary vascular branches on ct, severe hypoxemia or not on blood gas analysis, increase in alveolar -arterial oxygen gradient by ≥ kpa ( mmhg), and % diffusion disorder on pulmonary function tests [ ] . in addition, shunt-related examinations should be performed, such as cm tc-maa scanning, contrast-enhanced two-dimensional echocardiography and pulmonary angiography, etc., while the last one does not show the same sensitivity as that of the former two because the small blood vessels in the lungs may not develop on angiography. most hps patients have a slow onset of the disease, are difficult to treat, and have a poor long-term prognosis, with a mortality of more than % after years. therefore, early diagnosis of the disease and its differential diagnosis are vital to improving the prognosis of patients. first of all, the previous liver and lung diseases in the patients should be ruled out, such as chronic obstructive pulmonary emphysema, pulmonary infection, interstitial pneumonia and silicosis, etc. at the same time, cirrhosis with pulmonary hypertension, infections secondary to pleural effusion, interstitial pulmonary edema, atelectasis and hyperventilation syndrome, etc. need to be ruled out. hps should be differentiated mainly from the following diseases: . liver cirrhosis following pulmonary heart disease: this is mainly because pulmonary diseases result in cardiac insufficiency and thus increased pulmonary venous pressure. repeated or long-term existence of liver congestion can lead to central venous hypertrophy and lobular central connective tissue hyperplasia, and further progression of the lesion will lead to the formation of liver cirrhosis following pulmonary heart disease. patients with pulmonary cirrhosis often have a long history of chronic lung disease and signs of cardiac insufficiency, such as edema of lower extremities, palpitation, shortness of breath and other symptoms. this patient has no history of chronic lung disease or edema of lower extremities, making him inconsistent with liver cirrhosis following pulmonary heart disease. . left heart insufficiency: both hps and left heart insufficiency can cause severe dyspnea and hypoxemia. a history of liver disease or evidence of chronic liver damage and decreased po can be found in patients with hps, especially such characteristics as orthostatic hypoxemia and intrapulmonary vascular dilatation. patients with left heart insufficiency have a history of heart disease, orthopnoea, pink frothy sputum and moist rales in the lungs, etc. this patient is inconsistent with such manifestations. . primary pulmonary hypertension: after inhalation of pure oxygen, hypoxemia in most of patients with hps will be significantly alleviated. the effects of oxygen are poor in patients with hypoxemia [ ] , and hps is manifested as orthostatic hypoxemia. the characteristics of hemodynamics of patients with hps are hyperdynamics and normal or decreased pulmonary artery pressure and pulmonary vascular resistance, while those in hypoxemic patients are increased. . others: ductus arteriosus, eisenmenger syndrome and pulmonary embolism, etc., need to be differentiated from this disease, and comprehensive judgments should be provided based on other clinical data of the medical history. hps patients can also have the above-mentioned diseases, and careful and meticulous examinations are needed for differentiation. because hps is developed on the basis of original liver disease, the frequency of its occurrence and its severity are mostly associated with the liver cell function of patients, while there are also hps patients in whom chronic liver disease is relatively stable and liver functions are normal. besides, pleural effusion, ascites and infections secondary to pulmonary edema after liver function decompensation can aggravate patients' respiratory function injury. therefore, under the current circumstances in which there are no effective measures for hps, active and effective treatment of primary liver diseases is the basis for the treatment of hps. therapy of primary diseases, including correction of hypoproteinemia, elimination of pleural effusion, improvement of liver function and treatment of complications, etc., can improve tissue oxygenation and improve arterial oxygen saturation. on this basis, the following treatment can be given. oxygen therapy also helps the differential diagnosis of pulmonary shunt: if pao is resumed after oxygen inhalation, then the diagnosis of intrapulmonary vascular dilatation (ipvd) can be made; for patients with partial improvement, pulmonary anatomical shunt and functional shunt may coexist; for patients in whom the oxygen therapy proves inefficacious, pulmonary arteriovenous fistula is a possible diagnosis. it is now believed that once the diagnosis is established, treatment should be given as soon as possible. in the early stage of correcting hypoxemia in patients with mild conditions, even in patients in whom the critical value of hypoxemia (pao , - kpa ( - . mmhg) is reached and who have ascites, the hemoglobin saturation may still be less than % when patients are in activities or even sleep. that is to say, nasal catheter oxygen inhalation at - l/min is needed so as to improve hypoxemia [ ] . with the development of the disease, oxygen flow needs to be gradually increased, and intratrachea oxygen supply can be offered when necessary. during the late stage, patients can receive pressurized oxygen through a ventilator or a hyperbaric oxygen chamber. for patients whose conditions are severe, the efficacy of oxygen therapy alone is not obvious. . vasoactive drugs vasoactive drugs for the treatment of patients with hps are most studied; however, since its pathogenesis has not been clarified to date and primary liver disease is difficult to reverse, it is hard to define the clinical efficacy of these drugs. the commonly used drugs include: used aerosolized ephedrine hydrochloride for the treatment of patients with hps, and the preliminary efficacy was significant. the mechanisms were that ephedrine could excite the pulmonary vascular α receptor, resulting in contracted bronchial mucosa and pulmonary capillaries and alleviated bronchial edema, so that the dilated blood vessels within the lungs were contracted and intrapulmonary shunt was reduced. meanwhile, the bronchial β receptors were excited and the bronchi were dilated so as to improve the ventilation/blood flow ratio and relieve hypoxia. further studies are merited. (f) others: there have been reports on sympathomimetic drugs (isoproterenol) and β-blockers (propranolol), etc. that improve the symptoms of hps. theoretically, vascular endothelin, estrogen suppressor (tamoxifen) and so on can relieve the spider nevus and pulmonary vascular dilatation in patients with liver cirrhosis and improve their respiratory symptoms, while further studies are needed. no is most studied currently, and there are reports indicating that no synthesis inhibitors can increase pulmonary vascular resistance. alexander et al. used no for the treatment of severe hypoxemia in patients after liver transplantation, and obtained good results. durand et al. also reported that an hps patient was cured by inhaling no, while its mechanisms and clinical efficacy needed to be further confirmed. . pulmonary embolism it is generally considered that pulmonary vascular dilatation can vanish after liver transplantation in hps patients who are normal on pulmonary angiography or who have cavernous vessels on imaging [ ] ; for patients who show diffuse pulmonary vascular dilation features on pulmonary angiography, embolization is usually not adopted since patients' lesions are extensive and the efficacy is poor; for patients with isolated and severe pulmonary vascular dilation or arterial-venous communicating branches, local pulmonary embolism therapy can yield a satisfactory effect. . liver transplantation it is currently considered that liver transplantation is still a possible fundamental measure for the treatment for hps. in the past, it was believed that serious hypoxemia was an absolute contraindication against liver transplantation, while recent studies show that liver transplantation is preferred for patients who have good alveolar gas diffusion function, who can respond well to pure oxygen inhalation and who can undergone oxygenation safely during anesthesia. recent reports further prove that hypoxemia can be cured after liver transplantation [ ] . through literature review and case reports, krowka et al. believed that progressive hypoxemia in hps could be used as an indication of liver transplantation. temporary hypoxemia following liver transplantation can be adjusted by using no and taking the head-down supine position and the alternate lateral decubitus position. and for hps patients who fail to respond to the inhalation of pure oxygen, who have direct pulmonary arterial communicating branches on pulmonary angiography and who have severe clinical hypoxia, liver transplantation cannot improve their hypoxic status, has limited efficacy, or even increases intraoperative and postoperative risks. therefore, liver transplantation should not be performed on them. tips was an effective method for the treatment of hps, and its effects of improving symptoms, enhancing oxygenation and reducing intrapulmonary shunt could last up to months. riegler et al. performed tips on an hps patient with diffuse intravascular dilatation who was not suitable for vascular embolization, and the results showed significantly increased pao and significantly improved hypoxemia. however, coley et al. also reported that a patient failed to respond to tips, and therefore, its exact effects remain to be studied. . other treatment options one patient with hps was once treated with garlic, and months later, his oxygenation was significantly improved and his symptoms were relieved. there are also patients who receive plasma replacement therapy, which has limited effects on the oxygenation of patients with hps [ ] . to sum up, no effective treatment options are currently available for hps. since the basic cause of hps is liver cell failure, the usual cause of patients' deaths is not lung failure, mostly complications such as gastrointestinal bleeding, renal failure, hepatic encephalopathy and sepsis. therefore, we consider that the therapy of primary liver disease is particularly important. oxygen inhalation alone can be given in the early stage of hypoxemia, or conservative treatment can be provided if additional drugs are effective. liver transplantation is the best solution whenever possible. it is generally accepted that liver transplantation is the most promising regimen with confirmed efficacy. if oxygen inhalation is less satisfactory and patients are diagnosed with local intrapulmonary vascular dilatation or arterial -venous fistula by such means as pulmonary angiography, pulmonary embolism should be carried out as soon as possible. for patients with additional obvious portal hypertension, tips treatment can also be given. the interval from chronic liver disease and cirrhosis in patients to the confirmed hps due to such respiratory symptoms as anoxic dyspnea is usually several years or even more than years [average interval, ( . ± . ) years], and a small number of patients can develop such a disease acutely in the short term. besides, signs of chronic liver disease can be traced in patients complaining breathing difficulties. once hps is established, obvious hypoxemia has occurred already. it confers a poor prognosis, and most patients die within - years often due to other complications of liver disease [ , ] . if patients' oxygenation is satisfactory and they have undergone liver transplantation, or with the improvement in liver function, their hypoxemia can be resolved or improved of its own volition with good prognosis. if patients' oxygenation deteriorates severely and they have a very poor prognosis, most of them will die in the short term. hps often progresses slowly. although it is not a direct cause of death in patients with cirrhosis, it can significantly aggravate the disease. therefore, cirrhotic patients, especially those with positive liver palms and spider nevus as well as patients with portal hypertension, should be careful of the possibility of hps. timely detection and symptomatic treatment (such as low flow oxygen inhalation) can improve the prognosis of patients. active and effective treatment of primary liver disease forms the basis for the prevention of this disease. education of common sense should be given to patients with liver diseases so as to avoid factors inducing hps in their life. for patients with liver disease, mild hps should be found as soon as possible and appropriate treatment should be given. jia-quan huang and dong xu lipopolysaccharide (lps) is a constituent of bacteria cell wall which plays an essential role in the pathogenesis of septic shock by generating endogenous mediators such as nitrous oxide, cytokines, superoxide anions, and lipid mediators. despite the recent advances in antibiotic treatment and hemodynamic monitoring, septic shock still remains a serious disorder that is associated with a high mortality rate, to more comprehensive definition of the mechanisms that underlie innate immunity against bacterial pathogens, lps has been extensively studied [ ] . the pathophysiological consequences of bacterial sepsis are contributed by the dysregulation of these same mechanisms. before we can hope to design effective anti-sepsis therapies, greater insight into the nature of host interactions with lps is extremely essential. the gram-negative bacterial envelope is composed of two bacterial membranes, outer and inner membrane. the outer membrane consists of the following substances, like lipopolysaccharide (lps), several kinds of outer membrane proteins, lipid a and metal ions. for most gram-negative bacteria, lps is a major component in the outer monolayer of the outer membrane which works like a tight shield. the shield is composed by unique molecules, such as polysaccharide, or long chain of sugar, and lipid a. during the process to evoke the signaling events of lps, lipid a plays a pivotal role. the entire lipid component of lps molecule, however, is required for optimal activity [ ] . the basic principles of lps bioactivity are nowadays well understood. endotoxins do not elicit their toxic effects -as we might suspect and as it is known for many proteinous exotoxins which can kill host cells or inhibit cellular functions. rather, lps requires the active response of host cells. according to present knowledge we get, lps interacts with various host cell types through lipid a, those cells include mononuclear cells, thrombocytes, endothelial and smooth muscle cells, and polymorphonuclear granulocytes, among which macrophages/monocytes are of particular importance. through the lps-induced activation, macrophages produce many substances, like bioactive lipids, reactive oxygen species, and in particular, cytokines such as tumor necrosis factor a (tnf), interleukin- , il- , il- , and il- . it appears that when low levels of mediators are produced, beneficial effects (e.g., induction of resistance to infection, adjuvant activity) are elicited and when high levels of mediators reach the circulation that detrimental effects (e.g., high fever, hypotension, irreversible shock) are induced. however, when the host organism is in a hyperreactive state lps, low mediator concentrations may also become harmful. the hyperreactivity to endotoxin may be caused by exo-toxins, chronic infection, and by growing tumors, and interferon-γ. to function properly, organism requires an immune system that must detect pathogens, from viruses to parasitic worms, and distinguish them from the organism's own healthy tissue. the immune system can be classified into humoral immunity versus cell-mediated immunity or the innate immune system versus the adaptive immune system. when microbes invade organism, the innate response is usually triggered by pattern recognition receptors, which recognize components that are conserved among broad groups of microorganisms, or when injured, damaged, or stressed cells send out alarm signals, many of which (but not all) are recognized by the same receptors as those that recognize pathogens. the innate immune system defenses are nonspecific, meaning that the system responds to pathogens in a generic way. this system does not confer long-lasting immunity against a pathogen. in most organisms, the innate immune system is the dominant system of host defense [ ] . they activate innate immune responses by identifying some conserved non-self molecules, so as to protect the host from infection,. bacterial lipopolysaccharide (lps), an endotoxin which is found on the bacterial cell membrane, is considered to be the prototypical pamp. lps is specifically recognized by toll-like receptor (tlr) , a recognition receptor of the innate immune system. the interaction of the lipid a moiety of lps with macrophages appears to be especially important because subsequent cellular activation results in the release of systemically active pro-inflammatory molecules, which in turn mediate systemic toxicity. lps has extreme potential in macrophages activated at concentrations of lps as low as pg/ml [ ] . host-defense peptides (hdps) could be a possible alternative solution since they possess the antimicrobial, antiseptic, and immunomodulatory properties [ ] . endotoxins lipopolysaccharide is released not only from dead gram-negative bacterial, but also from the growing ones. endotoxins are very stable molecules, which are resisted to extreme temperatures and ph values in comparison to proteins. endotoxins are shed largely during cell death as well as growth and division. they are highly heat-stable and are not destroyed under regular sterilizing conditions. endotoxin can be inactivated through exposed at temperature of ° c for more than min or ° c for more than h. acids or alkalis of at least . m strength can also be used to destroy endotoxin in laboratory scale [ ] . gut microbiota is composed of strict anaerobes, facultative anaerobes and aerobes. recent reports suggest the existence of over , bacterial species in the human gut microbiota. an important characteristic of gut microbes is their heterogeneity [ ] . the composition and the frequency of the microbiome changes with the different segments of the elementary tract. the composition is influenced by the environment, consumed diet and host factors. endotoxin to surrounding tissues and organs or blood shift that shift pathway include: ( ) via the portal vein, liver into the systemic circulation; ( ) through the intestinal tract into the lymphatic system lymphatic; ( ) through the intestinal wall into the peritoneal cavity and then absorbed into the bloodstream. under physiological conditions, although a small amount of toxins continued to parenteral shifted via the portal vein into the liver, but it does not cause endotoxemia; mild in gram-negative bacilli infections, although bacteria continue to release to the tissue or blood endotoxin, but it does not give rise to a strong inflammatory response, the above are dependent on the presence of an effective mechanism within the body to remove toxins and detoxification. the liver is the main organ of clearance of endotoxin, and the spleen, also removes toxins. molecules in lps removed include cationic antimicrobial peptides (cationic antimicrobial peptides, cap), acyloxy acyl hydrolase (acyloxyacyl hydrolase, aoah) lipoprotein binding protein and anti-lps antibodies are important endotoxin clearance methods [ , ] . liver blood endotoxin clearance, primarily through kupffer cells, hepatocytes internal toxin endocytosis achieved, but the specific metabolic process is not entirely clear. mediated by kupffer cells engulf toxins may be scavenger receptor, it may be a molecular weight of , and , protein; mediate phagocytosis liver toxin structure may be the lectin-like receptor (lectin-like receptor). endotoxin receptor hepatocyte sinusoidal plasma membrane on the surface: after one to one binding, is taken up within the liver cells endocytosis way to microtubule-dependent vesicular transport through the liver cells, transported to the liver cells bile duct surface to exocytosis into the bile duct, and then discharged into the biliary system through the intestine [ ] . splenic macrophages containing approximately % of the body to settle within the organization, and macrophages are important endotoxin removal cells. when endotoxin intravenously into the body, except gathering in the liver, a lot of endotoxin can be quickly gathered and taken up into macrophages in the spleen, the spleen and therefore equally important endotoxin removal organs. in addition to its clear role in the performance of its direct effect, but more importantly, spleen macrophages is the precursor cells of kupffer cells in the liver, having a very big impact on removing toxins within the liver [ ] . mechanisms for removing toxins in the intestine are related to the intestinal villus tip epithelial cells. under normal circumstances, injected into the intestinal, endotoxin in intestine does not enter intestinal epithelial cells, but after intravenous injection of endotoxin, endotoxin may enter intestinal epithelial cells inside. therefore, endotoxin receptor may identify ways by endotoxin and (or) simple diffusion way into the intestinal epithelial cells. endotoxin way into the intestinal epithelial cells intravenously may have two: ( ) displaced from the lamina propria macrophages to intestinal epithelial cells basolateral; and enter intestinal epithelial cells from the side; ( ) including lower toxin, intestinal lamina propria macrophages, intestinal epithelial cells release large amounts of no and oxygen free, resulting in intestinal lamina propria microvascular injury, increased permeability, extravasation of endotoxin and ultimately displaced into the outer intestinal epithelial cells. villus tip epithelial cells within a stronger uptake of toxins, thus endotoxin can be started from the crypt, moving along the intestinal villi, and finally to the top of the inner hair cells of the intestinal epithelium. uptake of endotoxin villus tip epithelial cell loss, while the endotoxin into the intestine, which constitutes one of the effective clearance mechanisms of endotoxin [ , ] . plasma lipoproteins in endotoxin detoxification mechanisms play an important role, in which the lipopolysaccharide binding protein (lipopolysaccharide binding protein, lbp) and high-density lipoprotein (high density lipoprotein, hdl) play a major role. endotoxin into the bloodstream within minutes there were half white blood circulation due to binding to the edge of the pool or the pool is cleared, the remaining residual endotoxin and rapidly bound to plasma lipoprotein is inactivated. in plasma, lipoproteins and endotoxin when hdl plays a major role in its binding of endotoxin to reach more than % of the total, hdl, and thus research endotoxin important [ ] . cationic antimicrobial peptides are an ancient ingredients in the natural evolution of the immune system, including bactericidal/permeability-increasing protein (bactericidal/permeability-increasing protein, bpi), cathelicidin, lactoferrin, defensins and other substances, with not only the activity against gram-negative bacteria, but also the ability to combine internal toxins. cationic antimicrobial peptides are mainly in regular contact with the pathogen site mammalian skin, digestive tract, respiratory tract, and inherently express or express induced by pathogens and their products in the blood, secretions and neutrophil granules. cationic antimicrobial peptides have two types of three-dimensional structure; one is a α-helix, having such a molecular structure include cathelicidin and lactoferrin; the other is β-fold, including mammals α and β-defensins, etc. [ ] aoah aoah is a glycoprotein produced by white blood cells with weight of . - , , the large subunit of , and small subunits of . million to , , the large and small subunits connected by covalent disulfide bond. aoah as a lipase with hydrolysis for toxin, can selectively hydrolyze the secondary acyl chain on lipid a acyl groups acyloxy. when hydrolyzing endotoxins, both the large and small subunits of aoah play an important role, and both are indispensable. in addition to directly destroying toxin, the deacylated lps after the hydrolysis of endotoxin by aoah is also involved in aoah's detoxification mechanism on endotoxin; the material can accumulate and inhibit endotoxin-induced inflammatory response in the cell. however, due to a limited number of secretion by local infiltration of leukocytes, the internal detoxification of toxins is also limited [ ] . when the body respond with endotoxin, one trigger inflammation, on the other hand can be cleared to produce or activate, specific polysaccharide inactivate toxins, including antimicrobial-specific polysaccharide antibody and anti-core polysaccharide antibody. after two antibodies binding with endotoxin, and then with the fc receptors on the cell membrane, inner source of the toxin-mediated, so that the endotoxin inactivated intracellularly. anti-endotoxin antibodies can interfere with toxin within lbp binding, thus preventing lbp endotoxin transport [ ] . in the body's defense system, the shift from the inhibition of intestinal endotoxin ingredients include intestinal mucosal mechanical barrier, intestinal mucosal immune barrier, the normal intestinal flora and liver hepatocytes and kupffer cells, in which intestinal mucosal mechanical barrier, intestinal mucosal immune barrier, hepatocytes and kupffer cells play a direct inhibitory effect, while the normal flora plays an indirect inhibition. the intestine is huge "endotoxin library", a special anatomical location determines the intestinal mucosa must be an effective defense barrier. immunological barrier intestinal barrier formed by the epithelial cells of mechanical barrier and secretory iga (siga) and the like components [ ] . intestinal epithelial cells and tight junction formation mucosal mechanical barrier, is a significant barrier in the intestine endotoxin translocation defense to maintain its integrity is it to play an important role in defense guarantee. in severe trauma, burns, infection, considerable loss of body fluids, hypovolemia, cause the body ischemia and hypoxia. in order to maintain blood pressure, to ensure that the blood supply to the heart brain and other vital organs, compensatory splanchnic vascular contraction, including gastrointestinal ischemia and hypoxia longer time than other organs, even after shock patients after resuscitation to restore normal hemodynamics, stomach intestinal still in a state of shock occult. therefore, when the intestinal microvascular perfusion recovery, intestinal ischemic/reperfusion injury, epithelial cells produce large amounts of reactive oxygen species and other media, resulting in intestinal epithelial cell apoptosis, destruction of tight junctions between cells, thus rapidly increasing intestinal permeability mechanical barrier function weakens, thus contributing to the intestine of the endotoxin absorbed through the intestinal wall to parenteral tissue displacement [ ] . intestinal mucosal intestinal immunology barrier is a defense of the invasion of pathogens and endotoxin important line of defense, siga plays an important role in intestinal mucosal immunity. siga is an important component of the protection of the intestinal mucosa, both to prevent bacteria in the intestine mucosal surface colonization, but also in endotoxin. studies have found that e. coli o infection suffered intestinal mucosa, the anti-endotoxin core polysaccharide-specific siga secretion, in convalescent patients has been particularly evident, suggesting siga endotoxin to prevent the transfer of the body has a protective effect. in addition, studies suggest that nitric oxide (nitrogen monoxide, no) preventing endotoxin translocation in intestinal mucosal barrier oxide formed locally. under physiological conditions, nitric oxide synthase (inducible nitric synthase, inos) expression only in the respiratory epithelium, the pregnant uterus and ileal mucosa and a few other parts. induced by endotoxin including but under normal colonic epithelial cells also express inos and catalytic synthesis of no, are formed in the local oxidation barrier to prevent bacterial translocation colon, thus effectively preventing bacterial translocation, also indirectly prevents endotoxin shift [ ] . under physiological conditions, intestinal flora forms a relatively balanced microecosystem. flora distribution in the intestine has certain rules: deep close to the intestinal mucosal surface, parasitic anaerobic bifidobacteria and lactobacilli, these anaerobic bacteria are sugar coated, relatively stable, known as membrane flora; middle class bacteria, streptococcus digest, veillonella and excellent bacilli; the surface of e. coli and enterococci, can swim in the intestine, known as cavity flora. the antagonism between the layers flora, mutual cooperation, to maintain a dynamic equilibrium, in which the film anaerobic flora is a very important body's natural defense barrier that can prevent opportunistic pathogens such as e. coli colonization in the mucosa, but also can inhibit the overgrowth of opportunistic pathogens. intestinal flora micro-ecosystem is a very sensitive system, in severe stress or long-term systemic administration of large doses of broad-spectrum antibiotics, etc., the film significantly reduced the number of anaerobic bacteria, e. coli and other bacteria thrive conditions and continuous release of endotoxin to the intestine, since the film flora defense decreased, these opportunistic pathogens to colonize the intestinal mucosa, resulting in intestinal mucosal barrier damage, followed by the occurrence of intestinal bacteria, endotoxin translocation [ ] . under normal circumstances, the liver is one of the major barriers preventing endotoxin translocation, via the portal vein into the liver hepatocytes and a small amount of the toxin can be kupffer cell depletion. in conditions such as stress, not only liver cell dysfunction, so the ability to reduce endotoxin detoxification and collaterals between the portal vein and the vena cava, causing an overflow of endotoxin from the liver into the systemic circulation. endotoxin absorbed into the bloodstream, which in turn may increase the intestinal epithelial cells of the intestinal microvascular endothelial cell damage and, in a vicious cycle [ ] . when the body's defense system to produce responses in endotoxin, the innate immune system plays a major role. pathogens can be identified conserved receptors called pattern recognition molecules (pathogen-associated molecular patterns, pamps), including endotoxins of gram-negative bacilli, peptidoglycan grampositive cocci, lta and other cell wall composition and gram-negative bacteria, gram-positive bacteria such as dna. although a variety of pattern recognition molecules of different chemical structures, but they have similar characteristics: ( ) characteristic structure in which different types of pathogens in a relatively constant conserved; produced by a pathogen, the host body without these molecules; survival or disease-causing pathogen is generally the essential, such as mutations, death or loss of a pathogen will pathogenicity. natural immune system to recognize the receptor molecule called pattern recognition receptors (pattern-recognition receptors, prrs), including cd , toll-like receptor family (toll-like receptor, tlrs) and scavenger receptors. but in recognition of toxins, some differences exist between the different kinds of cells [ ] . macrophages in addition to expressing cd , tlrs and scavenger receptors and other associated endotoxin receptors on the cell surface, but in the cytoplasm also express the protein molecules nod recognizing toxins. cd , tlrs are key receptors that mediate endotoxin within macrophage activation; and scavenger receptor has relationship with macrophage clearing and inactivating toxins [ ] . kupffer cell is the main cell that clears the endotoxin in the liver. under physiological conditions, although there is still a small amount of bacteria and endotoxin via the portal vein into the liver, but kupffer cells will clear. kupffer cells are the most resident macrophages in the liver and are the largest number of resident cells in tissues. there is a theoretical speculation that if kupffer cells and on is very sensitive to endotoxin as other macrophages, the cell will be in constant activation, but in fact when kupffer cell engulfs, removes endotoxin, its itself is not activated by endotoxin, which suggests that in the treatment of endotoxin, kupffer cells have different mechanisms with other macrophages: kupffer cells treats endotoxin mainly depending on its phagocytosis. in the absence of serum, the phagocytic effects of kupffer cell on endotoxin can play a normal; and with the appropriate increase in endotoxin concentration, the phagocytic activity of kupffer cell was enhanced. the effect has something to do with phosphorylation events of two protein tyrosine residue individually weighting . million and . million. cd is the main receptor that mediates endotoxin activating kupffer cell, scavenger receptor is kupffer cells' important defense of receptor mediated kupffer cell to remove and inactivate endotoxin. there are four stages of the activation of kupffer cells, of which cd is the characteristic marker of cellular activation and function change. ( ) the stationary phase; the performance of less number of kupffer cells, small shape, a number in the hepatic sinusoids, cd staining negative; ( ) reaction period: for the local kupffer cells stimulate hyperplasia and systemic mononuclear macrophage intrahepatic accumulation; ( ) pre excitation period: kupffer cell phenotype occurred transformation period, expressed cd cell membrane receptor, kupffer cell functional changes; ( ) the activation period: the performance of nuclear transcription factor nf kb activation, cellular secretion cytokines [ ] . the cd (cd membrane-bound, mcd ) and tlr endotoxin were activated by the neutrophil surface to activate the neutrophils by binding with the receptor. in addition to the expression of the high affinity endotoxin receptor cd , the surface of the neutrophils also expressed the low affinity endotoxin receptor l-and the activated cells were activated by the receptor. in addition, the integrin is considered to be a low affinity endotoxin receptor for the surface of the neutrophils [ ] . it is generally believed that the expression of mcd was not on the surface of endothelial cells and serum soluble cd (soluble cd (scd ) is mediated endothelial cell recognition of lps molecules, and tlr is involved in lps induced endothelial cell activation. lbp was transported to scd by endotoxin, and tlr was activated by lps/scd and activated endothelial cells in the endothelial cell membrane. scd in addition to mediated endothelial cell activation and also mediated by endothelial removal of endotoxin and lps/scd /lbp form trimers and binding to endothelial cells, following the lps/scd endogenise, thus the removal of endotoxin [ ] . the epithelial cells of the intestinal mucosa were consistently associated with the bacterial and its products, and the bacteria and its products could stimulate other types of cells and induce inflammatory response, but did not induce intestinal epithelial cell defense, this feature for colonic epithelial cells is particularly important, because if can react to the normal intestinal flora in intestinal epithelial cells, it will cause adverse effects on the body. but this does not mean that intestinal epithelial cells are immune cells, when suffered pathogens and their products invasion, intestinal epithelial cells produce normal response. description: intestinal epithelial cells with normal differentiation of natural bacteria and pathogenic ability, and the recognition system of subcellular localization. there are different endotoxin recognition mechanisms in the intestinal epithelium and the myeloid cells [ ] . uncontrolled inflammatory responses (uncontrolled inflammatory response) has a relationship with infection, bacteremia, septicemia, sepsis, systemic inflammatory response comprehensive sign (systemic inflammatory response syndrome (sirs), compensatory anti-inflammatory response syndrome (compensatory antiinflammatory response syndrome, cars) and other related terms used for a long time, but also has an essential difference. out of control including inflammatory reaction syndrome (msas anti-inflammatory response syndrome, mars), a dynamic process of sirs and cars and the mixed antagonistic response syndrome, at present clinical many diseases occurrence and development are closely related. uncontrolled inflammation is a common pathological phenomenon in clinic, which is the important mechanism of the development of the complication after trauma. lps is one of the main factors that induce the uncontrolled inflammatory reaction in the most common. lps receptor on the monocyte/macrophage surface is the he initial factor for the body to recognize and start inflammatory reaction, also is one of the key links for the induction of uncontrolled inflammatory response. the concept of uncontrolled inflammatory response refers to inflammatory disorders then resulting in multiple organ dysfunction syndrome (multiple organ dysfunction syndrome. mods), emphasizes the importance of the of balance inflammatory/anti-inflammatory mechanism in the body, changes the limitations that in the past we only attached importance of the pathogenic effects of inflammatory factors. it is believed that the response of the body to the inflammatory factor is the dominant factor in the development of the whole body inflammatory reaction and mods. this concept is more focused than the previous focus on the dynamic changes of the whole process of inflammation. this can be divided into two types of mods: one is the early stage after the injury, that is, the speed hair style. the main blame is a strong inflammatory reaction induced by proinflammatory factors, and the other is a late phase of the disease, which is "late style", mainly due to the immune paralysis or worse immune disorders caused by cars or mars. the inflammatory is actually a kind of medium disease mainly caused by the chain reaction of cytokine. endotoxin is thought to be one of the most important predisposing factors in the chain reaction and can be referred for chain reaction "trigger" (trigger). endotoxin induced inflammation mechanism is mainly mediated by pamps that can induce cytokines such as il- , tnf alpha and other active molecules synthesis, the formation of the cytokine network, has a very important role in the occurrence and development of infection. the excessive activation of cytokines can cause septic shock, and is the leading cause of death in patients with bacterial infections. accordingly, prrs plays an important role in innate immunity and inflammation, and it can distinguish the pathogens from self organization through prrs organism, which is characteristic of immune response [ ] . inflammatory response syndrome systemic (sirs) is a systemic inflammatory response caused by any pathogenic factor to the body. the concept is first proposed by coris in . august american college of chest physicians and critical care medicine to present the diagnostic standard of sirs, think to have the following each of the two or more than two, sirs can be established: ( ) temperature > deg c or < deg c; ii heart rate beats/min; ( ), the breathing frequency > times per minute or arterial blood carbon dioxide into pressure (paco ) < . kpa mmhg; ( ) peripheral white blood cell count > × /l or × /l < or immature myeloid cells > %. what should be paid attention to is that sirs is a common athophysiological state of body with severe inflammatory reactions, and should be differentiated from some abnormal factors such as leukemia or cause increase or reduction of white cells after chemotherapy. although the naming of sirs has been generally concerned, but some scholars have raised objection to the concept, for example sirs has following problems: the sensitivity and specificity of the diagnostic criteria is poor, has the same meaning with the "critical"; can not understand the pathophysiology of the original disease; is difficult to guide clinical trials and practice. the production of sirs can be divided into two cases, the sirs caused by the infection and the non infectious sirs. from the point of view of the clinical development process, sirs can be followed by injury immediately aroused, then known as the "single phase velocity hairstyle; also to start local, and later developed into a systemic sirs, namely after the initial shock is brief period of stability, later gradually intensified when sirs is called" dual phase delayed onset. either of the factors or the clinical development process, the systemic inflammation of the control of the uncontrolled, and ultimately can lead to mods [ ] . the intestine is the biggest bacterial and endotoxin warehouse in the body. in severe trauma, systemic infection, intestinal ischemia and liver disease, there may be the occurrence of endotoxin. the main source is due to intestinal gram-negative bacteria in the excessive growth and reproduction, or due to increased intestinal permeability lps entry into the portal vein increased. if hepatic kupffer cell phagocytic function is low, the amount of endotoxin over the liver ability to remove endotoxin can "flood" (spill over) into the body of the loop and the endotoxemia formated. because of the endotoxin from the gut, so it is called intestinal endotoxemia (intestinal endotoxemia, ietm) [ ] . hepatitis b patients are often accompanied with ietm. its formation mechanism is: the production and absorption of intestinal endotoxin increased. there is a large number of gram negative bacteria in the body's normal intestinal, so endotoxin in intestinal contents is very high, but the intestinal mucosal epithelial cells have stronger resistance to toxins so that endotoxin is not easy to run through the intestinal mucosa into the blood, even a small amount of endotoxin breaking through the intestinal mucosal barrier into the portal vein, will be swallowed up by the kupffer cells in the liver. severe hepatitis b when the intestinal flora disturbance, endotoxin increased, increased intestinal hyperemia, edema and the permeability of the intestinal mucosa; endotoxin itself can damage the mitochondria and lysosome of intestinal epithelial cells, leading to epithelial cell autolysis; endotoxin can cause intestinal microvascular contraction of the intestinal mucosa, reduce blood, intestinal ischemia, hypoxia, cause the intestinal mucosal barrier function decreased, increased the absorption of endotoxin; severe hepatitis, due to intrahepatic bile acid and bilirubin deposition in kupffer cell phagocytosis was inhibited, resulting in the removal of endotoxin in the endotoxin decreased; through the door body circulation circuit into the systemic circulation, resulting in blood within liver cells to escape kupffer toxin the phagocytosis and clearance, which aggravate endotoxemia; the endotoxin can also pass through the celiac lymphatic system into systemic circulation by thoracic duct. in addition, severe hepatitis patients with sepsis, spontaneous bacterial peritonitis, etc., in the release of endotoxin, so the formation of the endotoxin is exogenous [ ] . in viral hepatitis and other basic diseases complicated with ietm and liver function failure are closely related and endotoxin can directly cause arterial vasoconstriction, the organ ischemia; endotoxin can activating endogenous clotting system, coupled with kupffer cell dysfunction, decrease delimination of blood coagulation or fiber soluble substances, easily lead to dic, so as to damage to multiple organs. endotoxin activated phospholipase a mediated membrane phospholipid degradation and lipid peroxidation, which is an important part of liver cell damage. nolan has pointed out that the effect of kupffer cell dysfunction induced by intestinal endotoxemia on liver and body, far more than the direct action the endotoxin, and production and release of inflammatory mediators and factors from kupffer cells activated by endotoxin are closely related. the occurrence of ietm affect hepatic energy metabolism, resulting in liver cell damage and necrosis, also caused hepatic microcirculatory disturbance, performing liver hemorrhagic necrosis. on the basis of severe hepatitis, it can accelerate liver function failure [ ] . hepatic cellular jaundice the acute and chronic liver function is often accompanied by intrahepatic cholestasis jaundice, and ietm plays an important role in the occurrence of intrahepatic cholestasis.. endotoxin involves in liver cell damage mainly through activation of phospholipase a , and inhibits the activity of na + − k + − atpase on liver canalicular membrane to make the bile excretion disorder, and then to cause intrahepatic cholestasis in the liver cells. endotoxin can start the peroxidation of liver parenchymal cells mitochondrial membrane lipid so that an increase in the content of oxygen free radical in blood, resulting in the disorder of energy generation, atp was reduced, so that the active uptake, metabolism and secretion of bile acid by liver parenchymal are short of energy, resulting in cholestasis [ ] . the liver disease with ietm often causes the coagulation dysfunction, the serious person appears the different degree bleeding, in particular the severe liver disease patient may concurrent dic, endangers the life. violi found that, when liver dysfunction in patients with ietm, the expression of tissue factors on the surface of macrophages and endothelial cell factor induced by endotoxin increased, promoting the synthesis of tumor necrosis factor (tumor necrosis, factor, tnf) to increase, and thrombin generation increasing, activation of coagulation system in about % of patients, followed by hyperfibrinolysis, suggesting that ietm in liver dysfunction can be used as the warning signal as the activation of coagulation and fibrinolysis system and activate, and with the increased hepatic lesions, plasminogen activation decreases with endotoxin levels increased, thus plasminogen may decline with endotoxin induced by chronic consumption of dic on the microstructure of ii related factors, new blood can not correct. in fact, before variceal rupture bleeding, patients with liver function severely damaged already have the gastrointestinal mucosa extensive ischemia and erosion, which is the potential causes of bleeding, ietm in the process. and gastric h + at this time can occur abnormal reverse diffusion and stimulate mast cells to release histamine, may lead to mucosal blood vessel dilation and permeability enhanced. as a result, hemorrhage, edema; histamine and directly stimulate the secretion of gastric acid, so that an increase in the number of h + and reverse diffusion, lesions persisted, form a vicious circle [ ] . it is important for the formation of ascites in ascites due to the obstruction of the hepatic vascular outflow tract obstruction. the initial vascular response to endotoxin was the rapid obstruction of the hepatic venous outflow tract and increased the portal pressure which may be related to endotoxin induced swelling of kupffer cells, liver cells with microvilli swelling, platelet aggregation and fibrin deposition effect, while others think that is endotoxin of blood vessels of the liver has a direct effect. ietm continuous damage to the liver cells, resulting in albumin synthesis and of hormones such as aldosterone de live function obstacles, thus affecting the renal function, and led to the emergence of the refractory ascites plays an important role in the process [ ] . patients with severe liver disease always are complicated with the functional renal failure, hepatorenal syndrome (hrs). the patients with severe liver disease can be associated with the pre-renal azotemia and acute renal tubular necrosis, and there is a certain relationship with ietm. the clinical studies showed that the levels of no --no and endotoxin in serum of patients with liver cirrhosis were significantly higher. at the same time, plasma renin activity, aldosterone and vasopressin levels increased and urinary sodium excretion decreased. the mechanism about ietm induced by hrs is not clear, may be related to the following factors: leukotrienes (leukotrienes, lts) lts can lead to renal vasoconstriction, increased renal vascular resistance, reduced renal blood flow and renal blood redistribution, decreased glomerular filtration rate induced by hrs, in ietm lts generation and release increased obviously. in addition, liver dysfunction, liver's uptake, inactivation and excretory function of lts decline, causing blood concentration increased; the thromboxane a (thromboxane a txa )/i (prostaglandin i , prostaglandin pgi ) can contract renal arterioles, decrease glomerular filtration rate, while pgi and pge (prostaglandin e , pg e ) is caused by on the role of txa . in patients with severe hepatitis with ietm, elevated systemic levels of pgi , reducing the renal vascular resistance, leading to renal vascular resistance txa , reduce the renal blood flow and glomerular filtration rate, promote the formation of hrs; third, nitric oxide, nitrogen monoxide) nono can through vasodilatation of the systemic, resulting in effective circulating blood volume reduction and evoked hrs; endothelial endothelin (et) et caused renal cortical blood priming hrs; and platelet activating factor (growth factor (paf) endotoxin and platelet activating factor (paf) can lead to a decrease in cirrhotic rats cardiac ejection fraction, reducing blood flow to the kidney, and paf antagonist can improve hemodynamics changes [ ] . the mechanism of endotoxin induced hepatic encephalopathy is not clear. we have known that lps can increase the permeability of blood brain barrier, promote intestinal toxic substances through the blood brain barrier (bbb), damage mitochondrial oxidative metabolism in brain cells, reduce oxygen utilization in patients with liver cirrhosis and disrupt energy metabolism of brain cells, induce brain edema. the clinical symptoms of chronic severe hepatitis with endotoxemia included in addition to fatigue, anorexia, tiresome of the oil, nausea, vomiting, yellow skin and sclera and, performance of endotoxemia. endotoxin can cause the release of histamine, -hydroxytryptamine ( -ht), prostaglandin, bradykinin, resulting in micro circulation expansion, venous blood volume reduction, decreased blood pressure, inadequate tissue perfusion, hypoxia and acidosis, and main symptoms and signs are: fever, elevated white blood cell count, bleeding tendency, heart failure, renal dysfunction, hepatic injury, nervous system symptoms and shock [ ] . improve liver function this is the basic treatment of ietm. liver function improved can strengthen mononuclear phagocyte system function to help the endotoxin removal. it can also decrease portal vein pressure to relieve intestinal congestion, edema, hypoxia, improve the intestinal microenvironment, reduce production and lymph reflux, and lower door shunt. these all contribute to the prevention and treatment of ietm. clean the gut saline is available as enema if severe liver disease, which helps reduce intestinal endotoxin generation and absorption. decompensated cirrhosis is often accompanied by small intestinal bacterial overgrowth and intestinal flora disturbance. thus, the promotion of intestinal flora back to normal state help prevent and treat intestinal endotoxemia. a variety of bifidobacterium, lactobacillus can be selected. a synthetic disaccharide, it is not digested and absorbed in the small intestine, but can be broken down into lactic acid, acetic acid and other small molecules by the bacteria into the colon. such acidification of the intestine reduces the generation and absorption of endotoxin, and promotes the growth of intestinal bacteria, stimulates bowel movements so as to increases tool frequency and so on. in addition, the lactulose may have internal direct inactivation of toxins, prevents activation of macrophages to release cytokines. oral absorption of antibiotics can effectively suppress the generation of intestinal endotoxemia. patients with liver cirrhosis taking oral polymyxin e or neomycin, the level of plasma lps and no -/no -horizontal declines in synchronization. polymyxin b has an internal direct antitoxin effect [ ] . it play a role by reducing intestinal absorption of toxins, inactivating toxins and inhibiting those lps-induced media by monocyte macrophages. the new anti-endotoxin therapy including interrupt endotoxin synthesis, binding or neutralizing its activity, preventing its interaction with the host effector cells, or interfering with toxin-mediated signal transduction pathways. therapeutic formulations include endotoxin analogs, antibodies, subunit vaccines, polymyxin combination column, recombinant human protein, small molecule inhibitors of endotoxin synthesis and intracellular signal transduction. bacteriophage producing a piece of short nucleotide sequence which plays the role of an antisense rna, blocking the synthesis of bacterial lps synthase. current clinical studies carried out an experiments in cloning of human anti-endotoxin lipid a light and heavy chain variable region. it laid the foundation for the next screening and expression that recombination between dna of antibodies and phage's succeeded. the clone is one kind of anti-polymyxin b (polymyxin b) monoclonal antibody of the igm class. it can play the role of anti-endotoxin shock by imitating the surface antigen structure of lipid a so as to substitute receptor antagonist of lipid a and lps blocking the causative link that the endotoxin induces inflammatory mediators [ ] . isolating antibodies having a high affinity of various g-bacteria to prepare a chimeric monoclonal igg antibody sdz - which have a therapeutic effect on the human endotoxemia. anti-endotoxin core glycolipid monoclonal antibody (antimonoclonal antibody r ) can prevent and treat the metabolic disorders in peritoneal infection with mods; it plays a significant role in conditioning in high catabolism, and can significantly improve metabolic disorders under the condition of abdominal infection associated with mods. bactericidal/permeability-increasing protein (bpi) bpi is a human endogenous protein, found primarily in neutrophils primary particles. its molecular amino-terminal and carboxy-terminal appear v-shaped structure planar symmetry. many amino acids to form a hydrophobic capsule hold lps's lipid a. it was reported that bpi has an obvious protective effect on intra-abdominal infection induced sepsis, which might be related to its antagonism against endotoxin [ ] . reconstructing hdl (high density lipoprotein, hdl) hdl can be used as an endogenous lps scavenging system, binding of bacterial endotoxin with high affinity to form a stable hdl-lps. lps-hdl complexation may contribute to a reduction in endotoxic activities in vivo by preventing lps (lipid a) from generating important transmembrane signals after binding to cells [ ] . e is the first generation lipid a analogue, which is derived from the lipid a structure from the endotoxin of rhodobacter capsulatus. it can block lps in cell culture without any endotoxin-like activity. e can protect mice from lethal doses of lps, and viable e. coli infections in combination with antibiotics. in human healthy volunteers who are exposed to intravenous lps. e also blocks the endotoxin response [ ] . an anti-cd antibody cd has a very important role in monocyte-macrophage cell signaling. since epitopes of lps on the cell membrane at the binding site is the same material with soluble cd , so we can develop a monoclonal antibody interfering with lps binding to cd and blocking to pass activation signals from immune effector cell [ ] . tyrosine kinase and mitogen-activated protein kinase are involved in lps cellular signal transduction. build anti-endotoxin (lps) single-chain antibody gene and attempt to make it express in e, coli. the scfv gene was successfully constructed and gst scfv fusion protein highly expressed in e. coli was obtained. glucocorticoids, including the synthetic glucocorticoid dexamethasone, are recognized for their anti-inflammatory properties and have the ability to inhibit the production of proinflammatory cytokines such as tnf-α. in intestinal ischemia-reperfusion methylprednisolone pretreatment can prevent endotoxemia. combined lps and dexamethasone treatment at h significantly changed tnf-α [ ] . this points that glucocorticosteroids added before or during stimulation of macrophages can prevent tnf release, after which the administration would be invalid. in fact, it is very difficult to use corticosteroids before tnf release. current clinical anti -tnf antibodies and tnf antagonist use exists, and before many scholars have obtained a more satisfactory results of blocking or neutralizing excessive tnf with anti-tnf. although the clinical symptoms improved, yet the survival rate is not higher than expected. the effect of anti--tnf clinical application needs further evaluation. the first proposed concept is sequential organ failure, based on which multiple organ failure (multiple system organ failure, msof or mof) is put forward, and in diagnostic criteria is developed, but it reflected the end-stage, denied reversibility, ignore the dynamic development from organ dysfunction to failure. therefore, the us accp/sccm proposed to replace the concept with mods. mods emphasized an early phase of organ dysfunction before overt failure occurred. it is defined as "the presence of altered organ function in an acutely ill patient such that homeostasis cannot be maintained without intervention." [ ] organ dysfunction may be relative, or can be absolute; with extension of time, mods can increase or reverse. thus, the term. mods was coined to indicate the wide range of severity and the dynamic nature of this disorder, which contributes to early diagnosis and treatment of patients and is more in line with clinical practice. there are two relatively distinct (although not mutually exclusive) pathways by which mods can develop: in primary mods, there is a direct insult to the organ that becomes dysfunctional. examples of such direct insults include gastric aspiration in the lungs or rhabdomyolysis in the kidney. this direct insult causes: an inflammatory response that is localized, at least in the beginning, to the affected organ. secondary mods is a consequence of trauma or infection in one part of the system that results in the systemic inflammatory response and dysfunction of organs elsewhere [ ] . secondary mods means not directly caused by damage, but to experience the "second strike", the first blow can make the immune system in a preactivated state under which inflammation lost control and significant sirs appears, then the following second strike can quickly cause multiple distant organ dysfunction and easily form sepsis with the basis of sirs/cars. this type of mods often develops as the following model: the original cause → stress → immune wpw → sirs/cars → infection → sepsis → mods → mof. the process of severe hepatitis, can cause intestinal damage and massive release of endotoxins into the blood, leading to intestinal endotoxemia (ietm). endotoxin is a powerful trigger for complement activation, and then these complement can activate "cascade effect" (cascade) to release oxygen free radicals, prostaglandins, endorphins, paf, cytokines and other inflammatory mediators to cause cytotoxicity, the error of microcirculation and tissue metabolism, eventually leading to the occurrence of mods. in this case, we emphasize mods originated in continuous, uncontrolled inflammation and factors causing systemic inflammatory response can be induced mods, including bacteria, fungi, parasites, viruses, toxins and other infectious agents. it is a debate of long-century problems that whether endotoxin has effects on hearts. the late s a large number of experiments prove endotoxins has a role of heart. it enables reduction of coronary blood flow, decrease of total coronary vascular resistance, and have meanings in heart failure. studies have shown that endotoxin can damage myocardial mitochondria, muscle paddle net, muscle membrane, contractile proteins etc., leading to cell membrane system damage, energy metabolism. past research in cardiac dysfunction under endotoxin shock did not attract enough attention, for they think that the heart is the final failure among all organs so that clinical treatment value is little. now people's awareness has completely changed that heart dysfunction can occur early in sepsis or septic shock. therefore, early identification and prevention of the occurrence and development of cardiac dysfunction may have some clinical value for treatment of septic shock and other serious complications [ ] . clinical gram-negative bacteria sepsis often complicated by adult respiratory distress syndrome, or server development of mods. in this pathological process, the high biological activity of endotoxin plays an important role. endotoxins often involving the lungs firstly, and the pathogenesis of non-injury may be associated with the direct damage on endothelial cell by endotoxin through complement pathway and induction of cytokines [ ] . the liver is the major site of endotoxin on clearing and detoxifying and the place where clinical gram-negative bacteria sepsis often can be complicated, and it is also the primary organ suffering attacks by toxins. it is generally believed that toxins in the liver circulating mainly from the gut and liver dysfunction is closely related to the formation of endotoxemia. histological examination revealed the inner toxins damage the liver cells, showing sinusoidal congestion, dieldrin expansion chamber, kupffer cell swelling, endoplasmic reticulum、mitochondria swelling, crest destroy and lysosomal activation etc. [ ] . the liver is the body's largest metabolic organ, and many cases of acute liver failure often involve other organ complications, which has become an important factor in determining the prognosis. endotoxins may cause the reduction of liver nutritional blood flow, mitochondrial oxygen metabolism, interfering with sugar metabolic pathways in liver leading to metabolic disorders. various damaging factors (such as gastrointestinal disorders, ischemia, immunocompromised, dysbiosis) promote absorb of intestinal bacteria and toxins and displacement via the portal vein, the lymphatic system into the systemic circulation. on the one hand these infectious agents can directly damage liver cells, or mediate hepatic injury whether by kupffer cell; on the other hand it can induce systemic inflammatory response by the monocyte-macrophage cells to release the media, both leading to organ perfusion disorder, affecting protein synthesis and energy metabolism, eventually resulting in severe sepsis, mods and even death. the effect of endotoxin on kidneys is not clear yet. in the early stage, it can affect the kidneys, decreased its blood flow renal via vasoconstriction. when endotoxemia is complicated by renal failure, the mechanism of glomerular filtration rate decreasing is unclear. the pathophysiology of aki in sepsis is complex and multi-factorial and includes intrarenal hemodynamic changes, endothelial dysfunction, infiltration of inflammatory cells in the renal parenchyma, intraglomerular thrombosis, and obstruction of tubules with necrotic cells and debris [ ] . the relationship between endotoxin and dic is quite complicated. dic is considered to be an important incentive of mods, especially patients with severe sepsis with dic having a highly possibility of developing mods, what's more, the prognosis is very poor, and the mechanism is multifaceted. endotoxin can start the endogenous coagulation system directly or via activating factor xii (hageman factor) by damaged endothelial cells, also can act on the monocyte-macrophage cells, stimulate the release of tissue factor to trigger the extrinsic coagulation pathway [ ] . in clinical practice it has been noted that serious infections is very possible to be complicated by gastrointestinal failure. the intestine is the biggest reservoir of bacteria in the body and leakage of bacteria or microbial products, notably lps, from the lumen of the gut into the systemic compartment, leads to initiation or amplification of a deleterious inflammatory response and mods [ ] . after endotoxins challenging the gastrointestinal mucosa, it initially shows mucosal telangiectasia, interstitial edema and hemorrhage. microcirculation leading to damage of lysosomal and release of proteases in the cell, cell degeneration and necrosis. in addition, mucosal cellular energy metabolism decrease, h + reverse diffusion, prostaglandins and bradykinin further aggravate mucosal damage. at the same time destroy of the gastric mucosal barrier also make a lot of bacteria and endotoxins pass through the gastrointestinal mucosa, migrate to the blood circulation, the lymphatic system and the abdominal cavity etc., leading to systemic multi-system organ damage. clinical characteristics of mods . organ failure usually do not result directly from the primary injury. there is a certain time interval from the primary injury to organ failure. . not all of infection have bacteriological evidence, and more than % of patients and autopsy found no infected lesions. thus, to identify and treat the infection may not be able to improve the patient's survival. . mods may have perfectly healthy organ involved, and it is ferocious and rapidly progresses. once happened, it is difficult to depress in the event almost, so often with a high mortality rate. . in pathology, mods lacks features, the affected organ only showing acute inflammation, such as inflammatory cell infiltration and so on, and these changes are very inconsistent with severe clinical manifestations, and once restored, patients do not have any clinical sequelae. . mods is closely with shock and infection. shock, infection, injury (including trauma and surgery, etc.) are the three main causes of mods. . generally the later period of shock will typesetting idc and mods, and the order of occurrence of mods usually is the lungs, liver, kidney, gastrointestinal tract, finally the heart. the characteristic clinical manifestations . instability of circulation due to a variety of inflammatory mediators have effects on the cardiovascular system, the circulation is most likely involved. almost all cases, at least in the course of the early and middle will be in highpower type of cycle of "high ranked low resistance". cardiac output up to l/ min or more and low peripheral resistance cause shock and need vasopressors to maintain blood pressure. . high metabolic systemic infection and mods are usually accompanied by severe malnutrition. its metabolic mode has three salient features: ( ) persistent high metabolism, metabolic rate up to . times more than normal; ( ) abnormalities of energy pathway. in starvation, the body obtain energy mainly through the decomposition of. however, with systemic infection, the body will get energy by breaking down proteins while the use of sugar is limited and fat utilization may increase early, fall later; ( ) poor response to exogenous nutrient, supplement of exogenous nutrition can not effectively prevent itself consumption, which suggests that a high metabolism itself has a "mandatory" also known as "autophagy metabolism." high metabolic may have serious consequences. first, protein malnutrition result from it will cause serious damage to the structure and function of the enzyme system of organs; secondly, imbalance of branched-chain amino acids and aromatic amino acid which makes the latter formate into a pseudo-neurotransmitter, then further lead to dysfunction of nerve. . hypoxia in tissue cells at present many scholars believe that the high metabolic and circulatory disorders often cause oxygen supply and oxygen demand does not match, so that the tissues of bodies are in a hypoxic state, mainly clinically manifesting "oxygen supply dependency" and "lactic acidosis. ". currently mods still lacks an unified diagnostic criteria, and any one of the diagnostic criteria of mods is difficult to reflect the entire contents of organ dysfunction, so in clinical practice we can select one according to our own specific situation. . the main contents from national critical care medicine conference standard in are: ( ) respiratory failure: r > /min; pao < . kpa; pco > . kpa; pao /fio ≤ . ( mmhg); p (aa) do (fio . ) > . kpa ( mmhg); x-ray of chest shows alveolar consolidation ≥ / lung (which have more than three or three); ( ) renal failure: except prerenal factors, little or no urine, serum creatinine, increased blood urea and nitrogen levels, exceeding more than twice the normal value; ( ) heart failure: systolic blood pressure < mmhg ( . kpa), sustained more than h; ci < . l/(min · m ); ventricular tachycardia; ventricular fibrillation; degree atrioventricular block; resuscitation after cardiac arrest (with which three or more); ( ) liver failure: total bilirubin> μmol/l; liver enzymes increased more than times compared with the normal; prothrombin time > s; with or without hepatic encephalopathy; ( ) dic: platelets × /l; prothrombin time and partial thromboplastin time prolong . times, and fibrin degradation products increase; systemic hemorrhage; ( ) brain failure: glasgow score below means coma, and less than points means brain death. . the sooner the primary diseases or the primary risk factors are eliminated or controlled, the greater the possibility of organ recovery is. . to effectively rescue and debride as soon as possible, prevent infection, prevent ischemia-reperfusion injury, use a variety of supportive care; . to reduce stress response, mitigate and shorten high metabolism and the magnitude and duration of glucocorticoid receptor; . to pay attention to the patient's breathing and circulation, as soon as possible to correct hypovolemia and hypoxia; . to prevent infection is an important measure of preventing mods; . if possible, improve the nutritional status of patients. . early treatment of any starting organ failure. mods is a problem in the medical field with an acute onset, rapid progression, and high mortality rate. so far for mods, there is no specific treatment, but through clinical monitoring, early detection of possible organ dysfunction, early intervention, and taking effective measures can slow down or block the course, 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around the globe. when the outbreak of this apparently novel infectious disease termed severe acute respiratory syndrome (sars) came to an end in july , it had caused over probable cases worldwide and more than deaths. starting in march , the world health organization (who) organised an unprecedented international effort by leading laboratories working together to find the causative agent. little more than one week later, three research groups from this who-coordinated network simultaneously found evidence of a hitherto unknown coronavirus in sars patients, using different approaches. after koch’s postulates had been fulfilled, who officially declared on april that this virus never before seen in humans is the cause of sars. ever since, progress around sars-associated coronavirus (sars-cov) has been swift. within weeks of the first isolate being obtained, its complete genome was sequenced. diagnostic tests based on the detection of sars-cov rna were developed and made available freely and widely; nevertheless the sars case definition still remains based on clinical and epidemiological criteria. the agent’s environmental stability, methods suitable for inactivation and disinfection, and potential antiviral compounds have been studied, and development of vaccines and immunotherapeutics is ongoing. despite its grave consequences in humanitarian, political and economic terms, sars may serve as an example of how much can be achieved through a well-coordinated international approach, combining the latest technological advances of molecular virology with more “traditional” techniques carried out to an excellent standard. severe acute respiratory syndrome (sars) is the latest in a series of emerging infectious diseases, and certainly one of the most widely publicised. this acute and often severe respiratory illness seems to have emerged in southern china in late (world health organization, c) . it soon caused considerable international alarm, after several index cases had given rise to outbreaks of sometimes ଝ this review is dedicated to all those who were prepared to risk their lives to provide care to sars patients and control the first pandemic of the st century. * corresponding author. tel.: + - - - ; fax: + - - - . e-mail address: annemarie.berger@em.uni-frankfurt.de (a. berger). enormous scales, and when the disease's ability to spread to distant areas within a very short period of time became obvious (world health organization, d) . a definition was developed for suspected and probable sars cases, based on clinical and epidemiological criteria; it has since been modified on several occasions. while sars demonstrated very vividly that in the modern world with an enormous volume of intercontinental traffic, infectious agents may be spread rapidly across the globe, it also serves as an example of how modern technologyprovided there is the necessary will, determination, and coordination to make best use of it-may help in combating such threats with unprecedented speed and enormous success. sars is characterized clinically by fever followed by respiratory signs and symptoms which may lead to rapidly progressive respiratory failure. as of september , people have been notified to the world health organization (who) as fulfilling the criteria for "probable sars", and of these, have died from sars (http://www.who.int/csr/ sars/country/ /en/). what made sars-in contrast, e.g. to influenza-notorious is its propensity to cause hospital outbreaks; some of these have affected over people, including health care staff, other patients and visitors . in contrast to many other emerging viral infections such as ebola, hantavirus pulmonary syndrome, and nipah, sars also clearly demonstrated its ability for easy and rapid geographic spread. this is because the sars agent affected a generally rather mobile population, and because those infected normally remain well enough to travel for several days after onset of infectivity. on march , the who set up a worldwide network of virological laboratories investigating sars cases (world health organization, a) . the investigations conducted by the members of these networks were coordinated by who's department of communicable disease surveillance and response (csr) through normally daily telephone conferences and a password-protected internet website. thus results and planned further studies were communicated and views and comments exchanged almost in "real-time" which made possible the rapid progress in elucidating the aetiological agent. in its final form, this network comprised participating laboratories from ten countries (world health organization multicentre collaborative network for severe acute respiratory syndrome diagnosis, ). investigations had soon ruled out a novel influenza virus strain, possibly of avian origin, as the cause of sars, and then focussed on members of the paramyxoviridae family, including human metapneumovirus (hmpv), and chlamydia-like organisms, including chlamydia pneumoniae. however, further investigations did not confirm these findings; the said agents were indeed found in a number of sars patients but not in all (who multicentre collaborative networks for severe acute respiratory syndrome (sars) diagnosis, ). almost nobody knew at that stage that virologists in beijing had already discovered a new virus in samples from some of the earliest sars patients. however, the official line in china at the time was that the novel "atypical pneumonia" was caused by chlamydia (enserink, a) . nevertheless, before the end of march, laboratories in hong kong, germany, canada, and the united states of america found evidence of a novel coronavirus in patients with sars by cell culture, electron microscopy, and by polymerase chain reaction (pcr) using primers at low stringency designed for other agents followed by sequencing (peiris et al., b; drosten et al., a; ksiazek et al., ; poutanen et al., ; drosten et al., b) . these results could not rule out that very thorough and extensive testing had by chance led to the discovery of a novel agent that was not responsible for the new illness but rather an "innocent bystander". however, the sequences obtained in different parts of the world were shown to belong to the same, previously unrecognised, coronavirus (ruan et al., ) . it could also be shown that sars patients underwent seroconversion against this coronavirus, using cells infected with patient isolates as antigen for indirect immunofluorescent antibody tests (drosten et al., a; ksiazek et al., ; fig. ) . furthermore, no evidence of present or past infection with this agent could be detected in limited surveys of healthy control individuals not suffering from sars (ksiazek et al., ) . this strengthened the case for the novel coronavirus being the cause of sars, but only after it had been shown to cause a similar illness in artificially infected macaques could it be regarded as fulfilling all four of koch's postulates ; world health organisation multicentre collaborative networks for severe acute respiratory syndrome diagnosis, ) . on april , , less than a month after the laboratory network had been brought into existence, who officially announced that a new coronavirus, never before seen in humans or animals and now provisionally termed sars-associated coronavirus (abbreviated as sars-cov), was the cause of sars . coronaviruses are large, enveloped, positive-stranded rna viruses with a diameter of - nm. most but not all viral particles display the characteristic appearance of surface projections, giving rise to the virus family's name (corona, latin, = crown). they have the largest genomes of all rna viruses. based on their unique transcription strategy that involves the formation of "nested" mrna molecules (cavanagh, ) . within the coronaviridae, the genera torovirus and coronavirus (type species: infectious bronchitis virus, ibv) are distinguished. a unique feature of coronavirus genetics is a high frequency of rna recombination as a result of discontinuous transcription and polymerase "jumping" (lai and cavanagh, ) . one example is the porcine respiratory coronavirus (prcov), which evolved in the early s from the enteropathogenic porcine transmissible gastroenteritis coronavirus (tgev), known since the s (pensaert et al., ) . through a large deletion in the s gene, the virus acquired an altered tissue tropism, causing mild respiratory infections. based on homologies on the amino acid sequence level, the known coronaviruses can be divided into three groups. table gives an overview of coronavirus species, group assignment, host species, disease manifestation and availability of a vaccine. there are more than a dozen known coronaviruses affecting different animal species; whereas group i and ii coronaviruses affect various mammals, those in group iii infect birds. some of these cause major problems in the livestock industry or may affect companion animals; therefore, considerable efforts have been devoted to their control, including development of active immunisation. negative-stain transmission electron microscopy of respiratory samples from sars patients and of infected cell culture supernatants reveals pleomorphic, enveloped virus-like particles with diameters of between and nm (fig. ) . most but not all viral particles showed the characteristic coronavirus-like surface features (ksiazek et al., ) . in contrast to most coronaviruses, which infect only the cells of their natural host species and a few closely related species, the sars-cov is able to infect different cell cultures, such as african green monkey (cercopithecus aethiops) kidney cells (vero) and the human colorectal adenocarcinoma cell line (caco- ), causing a massive cytopathic effect (cpe) after as little as days or days (fig. ) . it should be mentioned that these cell lines were not commonly used for the isolation of human respiratory viruses. interestingly during cell culture passages of the frankfurt isolate a virus variant emerged with a nucleotide deletion of bases in the orf b . the biological significance of this finding remains to be elucidated. complete genome sequences of sars-cov were first published by a canadian laboratory and the centers for disease control (cdc), atlanta (marra et al., ; rota et al., ) . as of end-october , full genome sequences are available on http://www.ncbi.nlm.nih.gov/genomes/ sars/sars.html. the genomic data available so far from several sars-cov strains suggest that the novel agent does not belong to any of the known groups of coronaviruses, fig. . electron microscopy image of sars-cov particle from infected cell culture supernatant after ultracentrifugation, % formalin fixation and negative staining with uranyl acetate (photograph by h. r. gelderblom, robert koch institute, berlin, germany). including the two human coronaviruses (hcov) oc and e (drosten et al., a; marra et al., ; peiris et al., b; rota et al., ) , to which it is only moderately related (fig. ) . the sars-cov appears to be neither a mutant of a known coronavirus nor a recombinant between known coronaviruses (holmes, ) . it has been proposed that the new virus defines a fourth lineage of coronavirus (group iv) (marra et al., ) (fig. ) . however, more recently it was suggested that sars-cov may be an early split-off from the group lineage . the sequence analysis of sars-cov suggests that it is an animal virus with a still unknown natural host species that has recently developed the ability to productively infect humans. a genetically very close but not identical virus was found in wild animals (masked palm civets and a raccoon dog) from a wildlife market in guangdong . but uncertainties remain over the exact source of this virus; the animals sampled could have been infected from humans or another animal species (cyranoski and abbott, ; normile and enserink, ) . sequence analysis of different sars-cov isolates reveals two distinct genotypes. one genotype was linked with infections originating from hotel m in hong kong, the other one comprises isolates from hong kong, guangdong and beijing that had no association with hotel m (ruan et al., ; tsui et al., ) . to date, there is no information as to whether different sars-cov strains may have different degrees of virulence. there is little doubt that sars originated from guangdong province of southern china (breiman et al., ) . the first cases retrospectively identified as sars occurred there in november . interestingly, amongst these early cases there seems to have been a significantly higher percentage of food handlers, chefs, etc. than in the general population, lending further support to a zoonotic origin. the worldwide spread of sars-cov was triggered through a single infected individual from guangdong who spent some time in hong kong before succumbing to sars (chan-yeung and yu, ) . during that time he unwittingly infected several others that in turn gave rise to a series of outbreaks (centers for disease control and prevention, ) . through sometimes several generations of transmissions, this event carried the virus to different hong kong hospitals and communities as well as to vietnam, singapore, canada, the united states of america, and beyond to a total of countries and areas of the world (world health organization, d). the virus travelled in infected humans and was passed on over several generations, as reflected in the genetic relatedness of isolates from these countries. although china was late in admitting it, the sars-cov had unsurprisingly also been spread within mainland china; in the end, the worst affected area was the capital, beijing, with cases in total, which surpasses the count for guangdong with by far (world health organization western pacific region, country office china: http://www.wpro.who.int/wr/chn/chn sars.asp). the incubation period of sars is short, ranging from to days. large studies consistently noted a median incubation period of days (booth et al., ; lee et al., ; tsang et al., ) . however, the time from exposure to the onset of symptoms may vary considerably . the who continues to conclude that the current best estimate of the maximum incubation period is days (who update -sars case fatality ratio, incubation period, http://www.who.int/csr/sars/archive/ a/en/). based on the latest data, the case fatality ratio is estimated to be < % in persons aged years or younger, % in persons aged - years, % in persons aged - years, and greater than % in persons aged years and older ; who update -sars case fatality ratio, incubation period, http://www.who.int/csr/sars/ archive/ a/en/). pregnant women with sars appear to have a worse prognosis and a higher mortality. therefore, early delivery or termination of pregnancy should be considered in those who are seriously ill with sars. for women who are relatively well with sars, however, there seems to be no reason for elective preterm delivery, such as reducing the risk of materno-fetal transmission (wong et al., a) . compared with adults and teenagers, sars seems to take a less aggressive clinical course in younger children (hon et al., ) . multivariable analysis showed that the presence of diabetes, advanced age or other comorbid conditions were independently associated with a poor outcome (booth et al., ; donnelly et al., ; fowler et al., ) . at the present time, with no new cases-apart from the isolated laboratory-acquired one-having been reported since june , sars-cov has apparently been driven out of the human population (world health organization, d) . in the meantime, who has issued a consensus document on sars epidemiology (who department of communicable disease surveillance and response, ). the pattern of geographic spread of sars was similar across all affected areas: typically, a patient with sars arrived from a previously affected area, was not identified as such when hospitalised, and thus infected health care workers, other patients and hospital visitors; these then infected their close contacts, and then the disease moved into the larger community (hawkey et al., ) . the virus seems to be spread predominantly by respiratory droplets over a relatively close distance , however, at least under some circumstances direct and indirect contact with respiratory secretions, faeces or animal vectors may also lead to transmission (hong kong department of health, ; who environmental health team, ; tsang et al., ; ng, ) . shedding of sars-cov in faeces and urine also occurs but its significance is unknown. the duration of infectivity is still unclear. faecal shedding seems to last for several weeks; this however does not necessarily mean that there is sufficient excretion of infectious viral particles to infect other individuals (peiris et al., a) . practising stringent droplets and contact precaution significantly reduces the risk of infection after exposure to patients with sars. therefore, the protective role of the mask suggests that the main route of transmission is by droplets . sars-cov spreads more efficiently in sophisticated hospital settings. evidence suggests that certain procedures, such as intubation under difficult circumstances and use of nebulizers increase the risk of infection . the only case of laboratory-acquired sars-cov transmission so far occurred in singapore in september . it involved a postgraduate who worked in a virology laboratory. subsequent investigation showed inappropriate laboratory standards (who severe acute respiratory syndrome (sars) in singapore-update , http://www.who.int/csr/ don/ /en); no secondary transmission arose from this case. it demonstrates the need for optimal biosafety precautions in laboratories working with sars-cov; these constitute the only places on earth where sars-cov is currently known to still exist and might be at the source of a re-emergence. blood transfusions or administration of blood products have not been implicated in transmission anywhere. this is despite the demonstration of viraemia during the clinical phase of the illness, albeit at low to moderate titres (drosten et al., a) . nevertheless, the potential of blood-borne transmission led to the early implementation of measures such as exclusion of possibly exposed individuals from the donor pool. the sars-cov is only moderately transmissible. a single infectious case will infect about three secondary cases (lipsitch et al., ; riley et al., ) . nevertheless, the clusters of cases in hotel and apartment buildings in hong kong show that transmission of the sars-cov can be extremely efficient. attack rates in excess of % have been reported. one common observation in various areas was the occurrence of so-called "super-spreaders", i.e. individuals that transmit the infection to at least ten others (world health organization, b). these "super-spreaders" were mostly very ill and often died from sars, and invariably serious lapses in infection control precautions had occurred during their management. so far there is no evidence that differences in virus strains may be responsible for the "super-spreader" phenomenon. there is also no firm evidence suggesting that subsequent transmissions led to clinically less severe illness, possibly through attenuation of the virus. it is also unclear why children are relatively under-represented amongst sars cases, and why on average they seem to suffer less severe sars illness. studies on the stability of the new sars-cov demonstrate the virus is more stable at room temperature than the previously known human coronaviruses (sizun et al., ) . the virus has been shown to survive for up to hours on plastic surfaces and up to days in diarrhoea. nevertheless the virus loses infectivity after exposure to different commonly used disinfectants and fixatives. heat exposure at • c quickly reduces infectivity (world health organization (who): first data on stability and resistance of sars-cov compiled by members of who laboratory network available at http://www.who.int/csr/sars/ survival /en/index.html). as defined by the who, a person is suspected to have sars if she has documented high fever (> • c), plus cough or breathing difficulty, and has been in contact with a person believed to have had sars, or has a history of travel to or stay in a geographic area where documented transmission of the illness has occurred, during the days prior to onset of symptoms ("suspect case"). a suspect case with infiltrates consistent with pneumonia or respiratory distress syndrome (rds) by chest x-ray is reclassified as a probable case. the revised case definition as of may , (see: http://www.who.int/csr/sars/casedefinition/en/) for the first time includes virus-specific laboratory results: a suspect case that tests positive for sars-cov in one or more assays should also be reclassified as probable. while recommendations have been issued for the use of laboratory methods for sars-cov (see: http://www.who. int/csr/sars/labmethods/en/), there are, however, at present no defined criteria for negative sars-cov test results to reject a diagnosis of sars. given the rather low shedding of sars-cov from the upper respiratory tract (drosten et al., a) , and the insufficient sensitivity of presently available laboratory methods, premature exclusion on the basis of negative test results may lead to tragic consequences. positive laboratory test results for other agents able to cause atypical pneumonia may serve as exclusion criteria; according to the case definition, a case should be excluded if an alternative diagnosis can fully explain the illness. nevertheless, the possibility of dual infection must not be ruled out completely. the required epidemiological linkage has repeatedly proven to be problematic. until an area is recognised as being affected, only imported cases fulfil the criteria for sars but not those who became infected locally through contact with unrecognised cases. thus, precious time may be lost until cases are recognised and appropriate measures taken. a thorough analysis showed that the existing who criteria lack sensitivity in the pre-hospital setting (rainer et al., ) . this again may be problematic as it may delay appropriate management of sars cases. the human coronaviruses known prior to march are difficult to propagate in cell cultures. their disease associations-generally mild respiratory illness ("common cold"), enteric and rarely possibly neurological disease-led to their widespread neglect in medical virology; only few groups worked on various scientific aspects, and very few laboratories offered routine diagnostic tests, mainly by pcr. sars-cov, on the other hand, is readily propagated in vitro and may also be detected by pcr and indirectly through antibody testing. nevertheless, and despite considerable progress in this field, much remains to be done until laboratory tests become a useful tool for the management of sars cases (world health organization multicentre collaborative network for severe acute respiratory syndrome diagnosis, ) . the presence of the infectious virus can be detected by inoculating suitable cell cultures (e.g., vero cells) with patient specimens (such as respiratory secretions, blood or stool) and propagating the virus in vitro. once isolated, the virus must be identified as sars-cov using further tests. according to international consensus, such work has to be performed under biosafety level (bsl) three conditions. sars-cov-specific rna can be amplified from various clinical specimens, especially in respiratory secretions and in stool, by pcr. high concentrations of viral rna of up to million molecules per millilitre were found in sputum. viral rna was also detected, albeit at extremely low concentrations, in plasma during the acute phase and in faeces during the late convalescent phase, suggesting that virus may be shed in faeces for prolonged periods of time (drosten et al., a) . a commercial real-time rt-pcr test kit containing primers and positive and negative controls developed by the bernhard nocht institute (http://www.bnihamburg.de/) is available (http://www.artus-biotech.de). an inactivated standard preparation is also available for diagnostic purposes through the european network for imported viral infections (enivd; http://www.enivd.de). enivd is also preparing for an international external quality assessment scheme for sars-cov assays. the existing pcr tests cannot rule out, with certainty, the presence of sars-cov in patients . on the other hand, contamination of samples in laboratories performing pcr may lead to false-positive results, unless appropriate precautions are taken. various methods were developed for the detection of antibodies produced in response to infection with sars-cov by probably virtually all patients. the first type of antibody test to be employed was the immunofluorescence assay (ifa). using cells infected with the patient's own virus isolate and an antihuman igg:fitc conjugate, we were able to demonstrate specific seroconversion in the two frankfurt sars patients (drosten et al., a; fig. ). an enzyme-linked immunosorbent assay (elisa) was developed that detects antibodies in the serum of sars patients and reliably yields positive results at around day after the onset of illness (world health organization multicentre collaborative network for severe acute respiratory syndrome diagnosis, ). the neutralisation test (nt) assesses and quantifies, by means of titration, the ability of patient sera to neutralise the infectivity of sars-cov on cell culture; the nt titre may therefore be correlated to clinical immunity although this has yet to be demonstrated. however, nt is limited to institutions with bsl- facilities. the only antibody test commercially available so far is an ifa which yields a positive result from about day after the onset of illness (euroimmun, lübeck, germany). as the diagnosis of sars is based entirely on a set of clinical and epidemiological criteria so far, reported case numbers are likely to include a substantial number of non-sars patients. therefore, recovered patients should be tested systematically for specific sars-cov antibody reactivity to confirm their diagnoses in retrospect and thus allow a better understanding of epidemiological and other features . although electron microscopy has an important role in the rapid diagnosis of infectious agents in emergent situations (hazelton and gelderblom, ) , it has provided only circumstantial evidence in the case of sars. when a virus-like agent was first visualised in clinical material from sars cases by electron microscopy, its classification was ambiguous; it later turned out to be human metapneumovirus (poutanen et al., ) . even in cases in whom coronavirus-like particles were detected, these could not be distinguished from the 'classic' human coronaviruses. no specific treatment recommendations can be made at this time. primary measures include isolation and the implementation of stringent infection control measures to effectively prevent further transmissions. empiric therapy should include coverage for organisms associated with any community-acquired pneumonia of unclear aetiology, including agents with activity against both typical and atypical respiratory pathogens. treatment choices may be influenced by severity of the illness. oxygen supplementation is often necessary, and severe cases seem to do better if intensive care including artificial respiration is commenced early (so et al., ) . efforts are underway at various institutions to assess potential anti-sars-cov agents in vitro. ribavirin, a "broad spectrum" agent active against various rna viruses has also been used clinically in sars patients (koren et al., ) , but seems to lack an in vitro effect (cinatl et al., a) . corticosteroids were widely used in sars patients, particularly in china. the rationale for their administration is the observation that tissue changes suggest that part of the lung damage is due to cytokines induced by the virus (peiris et al., a) . some clinical reports also underline their usefulness (zhao et al., ) . other therapies are being explored, such as convalescent plasma (wong et al., b) or normal human immunoglobulin which may be beneficial through an immunomodulatory effect or through acting against agents causing secondary infections. preliminary clinical data suggest that protease inhibitors used for anti-hiv therapy, lopinavir and nelfinavir (yamamoto n, personal communication), might have some efficacy, both as initial therapy and in the rescue setting. hong kong researchers reported at the who global conference on sars in kuala lumpur in june that sars patients treated with kaletra (lopinavir with low-dose ritonavir) plus ribavirin experienced a % reduction in death rate. while efforts are underway to develop more targeted anti-sars-cov approaches, broad screening of available substances in vitro has led to some potentially important clues. recently glycyrrhizin, a compound found in liquorice roots (glycyrrhiza glabra l.), was reported to have good in vitro activity against sars-cov (cinatl et al., a) . the mechanism of glycyrrhizin's activity against sars-cov is unclear. glycyrrhizin has previously been used to treat patients with hiv- and chronic hepatitis c virus . interestingly, this compound may be contained in some of the herbal preparations widely used in sars patients in china as part of traditional chinese medicine (lin et al., ) . furthermore, interferons inhibit sars-cov in vitro. in a recent study (cinatl et al., b) , interferon ß was more potent than interferon ␣ or ␥. therefore, it could become a drug of choice in future, alone or in combination with other antiviral drugs. the rapid success in identifying the causative agent of sars results from a collaborative effort-rather than a competitive approach-by high-level laboratory investigators making use of all available techniques, from cell culture through electron microscopy (hazelton and gelderblom, ) to molecular techniques, in order to identify a novel agent. hopefully this approach, coordinated by who, will serve as a model for future instances of emerging infections that will undoubtedly take place (ludwig et al., ) . despite the exemplary efforts that led to the identification of the causative novel coronavirus and allowed enormous knowledge about it to be accumulated within only a few months, it is maybe surprising that this success in terminating the outbreak has to be attributed to "old-fashioned" measures such as rapid and strict isolation of suspect cases and thorough contact tracing (world health organization, c); one is left wondering whether the same might also have been achieved without knowledge of the aetiology. thanks to an internationally well-coordinated and in most cases timely and determined response no new cases of sars have been notified since june . several countries reported sars cases imported from areas reporting outbreaks but did not experience secondary transmission; likewise, vietnam was the first country to demonstrate that-through a combination of early detection and public alert followed by decisive public health action and often heroic efforts by individuals-further transmission could be curtailed (reilley et al., ) . the absence of new clinical cases worldwide suggests that sars-cov no longer circulates within the human population; however, the possibility of clinically "silent" infections or of long-term virus carriers cannot be ruled out completely. furthermore, the origin of the agent remains obscure; sars-cov or a closely related virus persisting in a hitherto unidentified animal reservoir may yet again cross the species barrier and lead to human outbreaks. numerous questions relating to the epidemiology of sars have yet to be answered (normile and enserink, ; breiman et al., ) . at the time of writing (october ) it is completely uncertain whether sars will ever reappear. it is unclear whether seasonal recurrences may occur. in southern china, unlike europe and north america, the annual influenza peak incidence is from march to july (huang et al., ); thus, it shows a similar epidemic curve as the sars outbreak in (enserink, b) . the advent of the next 'flu season will pose considerable problems, given the lack of reliable laboratory methods for the early diagnosis of sars. the case definitions, too, will need to be adjusted to a world without sars; in theory, new cases are "impossible" as the criterion of an epidemiological link cannot be fulfilled. precious time may therefore be lost before a reappearance is detected. vigilance for sars must clearly be maintained (see: alert, verification and public health management of sars in the post-outbreak period- august -rationale for continued vigilance for sars; http://www.who.int/csr/sars/postoutbreak/ en/). for this purpose, who has defined three geographical zones according to their presumed risk for a sars recurrence: a potential zone of re-emergence, comprising guangdong and other areas where animal-to-human of sars-cov might occur; nodal areas, comprising hong kong, vietnam, singapore, canada, and taiwan, with sustained local transmission in spring or entry of numerous persons from the potential zone of re-emergence; and low risk areas. sars-related vigilance should be staged according to the zone in which a particular area is situated; for low risk areas, surveillance should be for clusters of "alert" cases among health care workers, other hospital staff, patients and visitors in the same health care unit. a sars alert is defined as two or more health care workers or hospital-acquired illness in at least three individuals (health care workers and/or other hospital staff and/or patients and/or visitors) in the same unit fulfilling the clinical case definition of sars and with onset of illness in the same -day period. in the other zones, this should be supplemented by enhanced surveillance, plus special studies for sars-cov infections in animal and human populations in the potential zone of re-emergence. besides improving existing detection assays-for instance, pcr methods based on the amplification of the nucleoprotein gene may be intrinsically more sensitive, due to the coronaviral transcription strategy , and thus be valuable for early diagnosis-further laboratory research needs to include detailed physico-chemical analysis of sars-cov proteins to allow the development of novel compounds based on targeted drug design (anand et al., ) . although an effective vaccine cannot be expected to be available soon, the relative ease with which sars-cov can be propagated in vitro is clearly helpful. a suitable animal model for sars may be available in the form of cynomolgus macaques (macaca fascicularis) . while the availability of vaccines against animal coronaviruses, such as avian infectious bronchitis virus, transmissible gastroenteritis coronavirus of pigs, and feline infectious peritonitis virus, is encouraging, the obvious lack of protective immunity in humans after infection with hcov oc and e is not. there is also currently no commercial veterinary vaccine to prevent respiratory coronavirus infections, except for infectious bronchitis virus infections in chickens. further research is also urgently needed to determine whether immune pathogenesis plays a rôle in sars or whether immune enhancement may occur, the chances of developing an effective and safe vaccine therefore remain uncertain. it is to be hoped that after such an encouraging start in an atmosphere of open collaboration and mutual trust, progress in sars-cov research will not be impeded by patent matters (gold, ) . coronavirus main proteinase ( clpro) structure: basis for design of anti-sars drugs clinical features and short-term outcomes of patients with sars in the greater toronto area role of china in the quest to define 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investigate the cause of severe acute respiratory syndrome description and clinical treatment of an early outbreak of severe acute respiratory syndrome (sars) in guangzhou, pr china key: cord- -mettlmhz authors: ortiz-prado, esteban; simbaña-rivera, katherine; gómez-barreno, lenin; rubio-neira, mario; guaman, linda p.; kyriakidis, nikolaos c; muslin, claire; jaramillo, ana maría gómez; barba-ostria, carlos; cevallos-robalino, doménica; sanches-sanmiguel, hugo; unigarro, luis; zalakeviciute, rasa; gadian, naomi; lópez-cortés, andrés title: clinical, molecular and epidemiological characterization of the sars-cov virus and the coronavirus disease (covid- ), a comprehensive literature review date: - - journal: diagn microbiol infect dis doi: . /j.diagmicrobio. . sha: doc_id: cord_uid: mettlmhz abstract coronaviruses are an extensive family of viruses that can cause disease in both animals and humans. the current classification of coronaviruses recognizes species in subgenera that belong to the family coronaviridae. from those, at least seven coronaviruses are known to cause respiratory infections in humans. four of these viruses can cause common cold-like symptoms. those that infect animals can evolve and become infectious to humans. three recent examples of these viral jumps include sars cov, mers-cov and sars cov- virus. they are responsible for causing severe acute respiratory syndrome (sars), middle east respiratory syndrome (mers) and the most recently discovered coronavirus disease during (covid- ). covid- , a respiratory disease caused by the sars-cov- virus, was declared a pandemic by the world health organization (who) on march . the rapid spread of the disease has taken the scientific and medical community by surprise. latest figures from th may show more than million people had been infected with the virus, causing more than , deaths in over countries worldwide. the large amount of information received daily relating to covid- is so abundant and dynamic that medical staff, health authorities, academics and the media are not able to keep up with this new pandemic. in order to offer a clear insight of the extensive literature available, we have conducted a comprehensive literature review of the sars cov- virus and the coronavirus diseases (covid- ). the viral membrane contains the spike (s) glycoprotein that forms the peplomers on the virion surface, giving the virus its 'corona'or crown-like morphology in the electron microscope. the membrane (m) glycoprotein and the envelope (e) protein provide the ring structure. within the virion interior lies a helical nucleocapsid comprised of the nucleocapsid (n) protein complexed with a single positive-strand rna genome of about kb in length [ ] . the first genome of sars-cov- named wuhan-hu- (ncbi reference sequence nc_ ) was isolated and sequenced in china in january [ , ] . the sars-cov- genome has similarities to other viruses: approximately % similarity to the bat coronavirus batcov rath ; an estimated % similarity with sars-cov [ ] , and an estimated % identity with mers-cov [ , ] . sars-cov- has a positive-sense single-stranded rna genome. it is approximately , bases in length and comprises of a ′ terminal cap structure and a ′ poly a tail. according to wu et al. [ ] , this novel coronavirus (ivdc-hb- / strain) has open reading frames (orfs) encoding proteins. the ' terminus of the genome contains the orf ab and orf a genes. orf ab is the largest gene and encodes the pp ab protein that contains non-structural proteins named nsps (nsp -nsp and nsp -nsp ). orf a encodes the pp a protein and also has nsps (nsp -nsp ) [ ] . the ' terminus of the genome contains four structural proteins: spike (s) glycoprotein; envelope (e) protein; membrane (m) glycoprotein and nucleocapsid (n) phosphoprotein. it also contains accessory proteins ( a, b, p , a, b, b, b and orf ) [ ] (figure b ). the global scientific community from countries have united to study this novel coronavirus by sequencing and submitting , sars-cov- genomes to the global initiative on sharing all influenza data (gisaid) (https://www.gisaid.org/) between december and april [ , ] . sars-cov- has accumulated mutations in its rna genome as the outbreak progresses. as an intracellular obligate microorganism, the coronavirus exploits the host cell machinery for its own replication and spread. since virus-host interactions form the basis of diseases, knowledge about their interplay is of great importance, particularly when identifying key targets for antivirals. j o u r n a l p r e -p r o o f ace is a type i membrane protein that participates in the maturation of angiotensin, a peptide hormone that controls vasoconstriction and blood pressure [ ] . in the respiratory tract, ace is widely expressed on the epithelial cells of alveoli, trachea, bronchi, bronchial serous glands [ ] , and alveolar monocytes and macrophages [ ] . xu et al. reported the [ ] rna-seq profiling data of organs with para-carcinoma normal tissues from the cancer genome atlas (tcga; https://www.cancer.gov/tcga) and organs with normal tissue from fantom cage (https://fantom.gsc.riken.jp/). these were used to validate the expression of the human cell receptor ace in the virus and may indicate the potential infection routes of sars-cov- [ ] . interestingly, the ace receptor is expressed more in oral cavity than lung. this potentially could indicate that susceptibility and infectivity of sars-cov- is greater from oral mucosa surfaces [ ] . following the binding of the rbd in the s subunit to the receptor ace , sars-cov- s protein is cleaved by the cell surface-associated transmembrane protease serine tmprss , which activates s domain for membrane fusion between the viral and cell membrane [ ] . a functional polybasic (furin) cleavage site was found at the s -s boundary through the insertion of nucleotides [ , , ] . the s , t and s residues of o-linked glycans flank the cleavage site and are unique in sars-cov- [ ] . in addition to the s glycoprotein -ace receptor complex, wang following the release and uncoating of viral rna to the cytoplasm, coronavirus replication starts with the translation of orf a and orf b into polyproteins pp a and pp ab via a frameshifting mechanism (figure ) [ ] . subsequently, polyproteins pp a and pp ab are processed by internal viral proteases, including the main protease m pro , a potential drug target whose crystal structure was recently determined for sars-cov- [ ] . polyprotein cleavage yields mature replicase proteins, which assemble into a replicationtranscription complex that engages in negative-strand rna synthesis. both full-length and multiple subgenomic negative-strand rnas are produced. the former serves as template for new full-length genomic rnas and the latter template the synthesis of the subgenomic mrnas required to express the structural and accessory protein genes residing in the ′proximal quarter of the genome [ ] . coronavirus rna replication occurs on a virusinduced reticulovesicular network of modified endoplasmic reticulum (er) membranes [ ] . the assembly of virions is quickly ensued with the accumulation of new genomic rna and structural components. the n protein complexes with genome rna, forming helical structures. then, the transmembrane m protein, localized to the intracellular membranes of the er -golgi intermediate compartment (ergic) , interacts with the other viral structural proteins (s, e and n proteins) to allow the budding of virions [ , ] . following assembly and budding, virions are transported in vesicles and eventually released by exocytosis. normal immune responses against the majority of viruses involve a rapid containment phase mediated by innate immunity components and -if necessary-a delayed, yet more sophisticated adaptive immunity phase that should be able to eradicate the pathogen andhopefully-generate long-lasting immunological memory. the former includes antiviral type i interferons (ifns), macrophage and neutrophil activation that leads to proinflammatory cytokine production and nk cells on the other hand, anti-viral adaptive immune responses involve a virus-tailored coordinated attack by antigen specific cd + cytotoxic t cells (ctls), the th subset of cd + t helper cells that orchestrates the j o u r n a l p r e -p r o o f immune response against viruses and other intracellular pathogens, specific antibody producing plasma cells, and finally the production of memory t and b cell subsets. immune responses following sars-cov- infection can be a double-edged sword. a rapid and robust type i ifn orchestrated response can lead to virus clearance and -given that antiviral lymphocytes are activated and expanded-immune memory. conversely, a late activation of innate immunity, possibly owing to is usually associated with severe pathology that can lead to pneumonia, ards, septic shock, multi-organ failure and, eventually, death. in this line, a delayed type i ifn response and inefficient sars-cov- clearance by alveolar macrophages can promote excessive viral replication that can lead to severe pathology accompanied by increased viral shedding and, thus, viral transmissibility. accordingly, in patients whose immune system is weakened or otherwise dysregulated, such as older men with comorbidities, severe covid- is clearly more likely to occur [ ] [ ] [ ] . a recent study has demonstrated that the average duration of sars-cov- viral shedding was days after covid- onset, raising a debate as to the optimal time of patient isolation [ ] . however, in terms of transmission viral shedding seems to be more relevant in the early phases of the infection as it can precede covid- symptoms by - days whilst up to % of infections are associated with viral shedding by asymptomatic cases [ ] [ ] [ ] therefore, individuals that mount efficient containment-phase immune responses accompanied by decreased inflammatory responses will not experience infection-or immune response-mediated overt manifestations, but may be important silent spreaders of sars-cov- . type i ifns are mainly produced by plasmacytoid dendritic cells (pdcs) and have a plethora of antiviral effects such as blocking cell entry and trafficking of viral particles, inducing rnase and dnase expression to degrade virus genetic material, enhancing presentation of viral antigens by mhc-i, inhibiting protein synthesis, inducing apoptosis of j o u r n a l p r e -p r o o f infected cells and activating anti-viral subsets such as macrophages and cytotoxic nk cells and t lymphocytes [ ] . pathogen recognition receptors like cytosolic rig-i and mda- [ , ] or endosomal toll like receptors (tlrs) and that recognize viral rna [ ] are responsible for the activation of signaling cascades that activate the transcription factors nf-kb, interferon regulatory factor (irf) and irf that translocate to the nucleus and induce proinflammatory cytokines and type i interferon (ifn) production. in turn, type i ifns activate the downstream jak-stat signal pathway resulting in expression of ifnstimulated genes (isgs) [ , ] . our experience from sars-cov and mers-cov infection has shown that delayed type i ifn production and excessive recruitment and activation of infiltrating proinflammatory cells (neutrophils and monocytes-macrophages) are possible mediators of lung dysfunction and bad prognostic factors for the outcome of the infection. delayed type i ifn production allows for highly efficient viral replication that, in turn, results in recruitment of hyperinflammatory neutrophils and monocytes. therefore, the pathogen recognition receptors (prrs) of these proinflammatory cells recognize high numbers of their ligands and subsequently secrete excessive amounts of proinflammatory cytokines that lead to septic shock, lung pathology, pneumonia or acute respiratory distress syndrome. it has been shown that in severe cases both sars-cov and mers-cov fruitfully employ an immune evasion mechanism whereby early type i ifn responses to viral infection are dampened [ ] . this can be achieved by blocking signaling both upstream, as well as downstream of type i ifn expression. sars-cov can inhibit irf nuclear translocation, whereas mers-cov can impede histone modification [ ] . additionally, both viruses can inhibit ifn signaling by decreasing stat phosphorylation [ ] . due to the many sequence similarities of sars-cov- with sars-cov and mers-cov it would be enticing to speculate that similar mechanisms are also present, however further studies are needed to shed light to this hypothesis. hyperactivated neutrophils and monocytes-macrophages are the usual source of the cytokine storm. in this aspect, absolute neutrophil counts and neutrophil to lymphocyte j o u r n a l p r e -p r o o f ratio (nlr) were strongly associated with disease severity in a large cohort of covid- patients and were proposed as markers of adverse disease prognosis [ ] . interestingly, the increased amounts of proinflammatory cytokines in serum associated with pulmonary inflammation and extensive lung damage described both in sars [ ] and mers diseases [ ] were also reported in the early study of patients with covid- in wuhan [ ] . evidence shows that the leading cause of covid- mortality is respiratory failure caused by acute respiratory distress syndrome (ards). there is an association with a cytokine storm mediated by high-levels of proinflammatory cytokines including il- , il- , il- , g-csf, ip- , mcp- , mip- a and tnf-α. ards was associated with increased fatality and subsequent studies confirmed il- and c-reactive protein are significantly upregulated in patients that died compared to convalescent patients [ ] moreover, a recent study of patients in wuhan identified that severe cases showed significantly higher cytokines and chemokines such as tumor necrosis factor-α (tnf-α), il- , il- , il- and il- expressed [ ] . in accordance with these findings, therapeutic strategies are being tested. a phase randomized controlled trial of il- blockade (anakinra) in sepsis has shown significant survival benefit in patients with hyperinflammation, without apparent increased adverse events [ ] . currently, a multicenter, randomized controlled trial of tocilizumab (il- receptor blockade, licensed for cytokine release syndrome), is being trialed in patients with covid- pneumonia presenting with high levels of il- in china (chictr ) [ ] . moreover, several clinical trials are exploring if the well-established antiviral [ ] and anti-inflammatory effects of hydroxychloroquine will be effective in treating patients with covid- as has previously been suggested for sars-cov infection [ ] . this has also been demonstrated in vitro for sars-cov- [ ] . in contrast, janus kinase (jak) inhibition has been proposed as a potential treatment in order to reduce both inflammation and cellular viral entry in covid- [ ] . thus, it comes as no surprise that in a recent correspondence, lancet authors have identified the following potential therapeutic options for cytokine storm syndrome including ards the use of corticosteroids, selective cytokine blockade (eg, anakinra or tocilizumab) and jak inhibition [ ] . j o u r n a l p r e -p r o o f virus presentation to the different t cell subsets stands on the crossroads between innate and adaptive immune responses. studies on sars-cov and mers-cov [ ] presentation have identified several susceptibility and protection conferring hla alleles. the dearth of similar data regarding sars-cov- antigen presentation to t cells and possible virus evasion mechanisms of this process suggests it is a virgin investigation field to be explored. apart from the sustained inflammation and cytokine storm, lymphopenia has been implicated as a major risk factor for ards and mortality in the context of covid- [ ] . similar findings were described for sars-cov infected patients who had considerable decreases of cd + t and cd + t cells [ ] . however, in convalescent patients specific tcell memory responses to sars-cov were still found six years post infection [ ] . showed no reactivity with viral antigens. however, the small number of sera used (n= ) implies that further investigation is needed to corroborate these results [ ] . nonetheless, since we are currently in early stages of sars-cov- pandemic more studies need to be carried out to shed light on antibody persistence (both igm and igg) and protective effects. recently, macaques re-challenged with sars-cov- after a primary infection did not show signs of re-infection, suggesting that protective immunity and memory responses were fruitfully mounted. this finding can also impact vaccine production strategies [ ] . importantly, covid- convalescent sera was shown to hold promise as a passive immune therapy alternative to facilitate disease containment [ ] . to the best of our knowledge, at j o u r n a l p r e -p r o o f least one pharmaceutical company, takeda, is preparing to purify antibody preparations from covid- convalescent sera against sars-cov- [ ] . a recently published case report of a patient with mild-to-moderate covid- revealed the presence of an increased activated cd + t cells and cd + t cells, antibody-secreting cells (ascs), follicular helper t cells (tfh cells), and anti-sars-cov- igm and igg antibodies, suggesting that both cellular and humoral responses are important in containing the virus and inhibiting severe pathology [ ] . antibody dependent enhancement (ade) is a mechanism whereby non-protective antibodies produced during an infection with an agent either mediate increased uptake of this agent into target cells or cross-recognize a different pathogen and facilitate its entrance to target cells [ ] . evidence emerging over the past two decades suggests that antibodies against different coronavirus can cross-react to some extent and mediate ade [ ] . ade in the context of sars-cov was thought to be mediated by antibodies produced against e-cov [ ] and was contemplated as contributing to high mortality rates in china [ ] . the described mechanism suggests that low affinity or low title anti-spike protein antibodies rather than neutralizing the virus result in fc receptor mediated infection of immune cells, further aggravating the dysregulation of anti-sars-cov immune responses [ ] . indeed, in vitro as well as in vivo experimental models have shown that ade hinders the ability to manage inflammation in the lung and elsewhere. this may lead to ards and other hyperinflammation-induced clinical manifestations also observed in several of the documented cases of severe covid- [ , ] . while the molecular and immunological host response to sars-cov- infection has not yet been fully elucidated to confirm ade is occurring, anti-sars-cov- antibodies have been shown to partially cross-react with sars-cov, suggesting enhancement is a possibility. with this in mind, ade in populations previously exposed to other coronavirus can partially explain the geographic discrepancies observed in covid- pathogenesis and severity. finally, ade can have several implications in vaccine development as low-affinity or low-titer antibody producing vaccines can increment susceptibility rather than confer protection against the pathogen as has previously been described for a dengue vaccine [ ] [ ] [ ] . j o u r n a l p r e -p r o o f detection methods based on nucleic amplification are often used in the case of sars-cov, mers-cov and other viruses, because have high sensitivity and specificity, particularly in the acute phase of infection [ ] . case identification and surveillance of covid- spread although rt-qpcr assay is considered the gold-standard method to detect viruses such as sars-cov and mers-cov [ , ] , currently available rt-qpcr assays targeting sars-cov- have important considerations. firstly, due to the genome similarity of j o u r n a l p r e -p r o o f sars-cov- to sars-cov ( % of nucleotide identity [ ] ), some of the primer-probe sets described by different groups and listed in the who coronavirus disease (covid- ) technical guidance [ ] , have cross-reaction with sars-cov and other bat-associated sars-related viruses, therefore, it is important to run confirmatory tests. loop-mediated isothermal amplification (lamp) is a one-step isothermal amplification reaction that couples amplification of a target sequence with four to six primers, to ensure high sensitivity and specificity, under isothermal conditions ( - °c), using a polymerase with high strand displacement activity [ ] . in the case of an rna sample, lamp, is preceded by the reverse transcription of the sample rna. rt-lamp has been used before for the detection of various pathogens [ ] . including sars-cov- and other respiratory viruses [ , ] . recently, it received emergency use authorization (eua) from the u.s. j o u r n a l p r e -p r o o f serological tests also, called immunoassays, are rapid and simple alternatives for screening of individuals that have been exposed to sars-cov- based on the qualitative or quantitative detection of sars-cov- antigens and/or anti-sars-cov- antibodies. there are several types of serological tests available, including elisa (enzyme-linked immunosorbent assay), iift (indirect immunofluorescence test), lateral flow immunoassays and neutralization tests. immunoassays assays are very useful because they allow to study the immune response(s) to sars-cov- in a qualitative and quantitative manner. in addition, help to determine the precise rate of the infection [ , ] , and to determine more precisely the fatality rate of the infection [ ] . several sars-cov- targeted serological tests are commercially available or in development [ ] . a recently developed kit, reported a sensitivity of . % and specificity of . % [ ] using sars-cov- igg-igm combined antibody rapid (within minutes) test [ ] . despite their simple and fast readout and their potential for being used outside laboratory environments (bedside, small clinics, airports, train stations, etc.), serological tests have a critical disadvantage; given the fact that antibodies specifically targeting the virus would normally appear after days or longer [ ] after the illness onset [ ] , tests based on this principle have a lag period of approximately to days post-infection. during this lag period, infected and non-infected individuals will both result in a negative output. in addition, it is important to highlight that because serological tests depend on the ability to produce antibodies, intrinsic immunological differences and/or responses between individuals, can significantly affect the outcome of these tests. recently, some commercially available immunoassays received ce mark for professional use [ , ] , and therefore are registered as in vitro diagnostic devices. currently, there are a plethora of antibody tests for covid- with variable performance (sensitivity varying from to %, specificity from to %, reviewed in (foundation for innovative new diagnostics) [ ] . different manufacturers of serological assays declare that their assays have no cross reactivity to other human coronaviruses and other respiratory viruses. however, despite the data provided by manufacturers, recent studies highlighted that several of the commercially available tests have sensitivity and/or specificity issues that should be considered for using and analyzing results of many of these tests [ ] [ ] [ ] [ ] [ ] . j o u r n a l p r e -p r o o f as mentioned before, immunoassays -particularly tests detecting anti-sars-cov- igm and/or igg-indicate that the person has been exposed to the virus. in the case of other viral infections, having antibodies targeting a pathogen has often been considered an indication of having immunity against that pathogen [ ] . based on this idea, some governments have suggested using serological tests, to determine who has developed immunity against sars-cov- and provide positive individuals a "risk-free certificate" or "immunity passports", which would enable them to travel or to return to work, assuming that they are protected against re-infection [ ]. however, based on the limited knowledge of how immunity to this virus works [ ] , there is not enough evidence to declare a person immune, or to confirm that people who have anti-sars-cov- antibodies are protected from a re-infection. even though covid- can be diagnosed using qpcr as the gold standard, inadequate access to reagents and equipment has slowed disease detection even in developed countries such as the us. several low cost and rapid tests using different approaches have been described. unlocking) technique for the detection of covid- and the detectr (developed by mammoth biosciences) prototype rapid detection diagnosis kit using crispr to detect the sars-cov- in human samples have been described [ ] . the use of rna aptamers, have recently emerged as a powerful background-free technology for live-cell rna imaging due to their fluorogenic properties upon ligand binding, a technology that could be of use to diagnose sars-cov- infection [ ] . finally the use of next generation sequence (explify®) might be used to detect and identify bacterial, viral, fungal, and parasitic pathogens by their unique genome sequences [ ] . in covid- symptomatic infection, the clinical presentation can range from mild to ventilation assistance [ ] . the spectrum of symptoms of covid infection are characteristic of a mild disease in most of the cases, however, it is important to point that the progression could lead to a severe respiratory distress. asymptomatic infection (while incubation occurs) was described both in the first cases in wuhan and in other cohorts. a group of isolated patients were screened for sars-cov- , where % ( cases) were positive for the test, and half of these cases had no symptoms. on the other hand, there are reports of cases without overt symptoms in which there were ground glass images in the chest tomography in up to % of patients [ ] . of the asymptomatic cases studied in wuhan city, the . % of people exposed developed specific symptoms in . days, and the remaining . % were symptomatic in the following . days (ci, . to . days). the median estimated incubation period was . days ( % ci, . to . days) [ ] . some patients with initially mild symptoms had symptom progression over the course of one week [ ] . the descriptive studies available so far have concluded that the majority of cases are mild infections (more than % of cases); with up to % of patients being sever j o u r n a l p r e -p r o o f in most cohorts, and less than % have been considered as critical cases with high vital risk [ ] . in a study describing patients with covid- pneumonia in wuhan, the most common clinical characteristics at the onset of the disease were described. this is consistent with other international cohorts (table ) [ ] . it is important to note that fever is not always present and up to % of patients could had a low grade temperature between . to degrees celsius or normal temperature [ ] . if these patients required hospitalization, % developed a fever during the course of the illness. rarer accompanying symptoms included headache without warning signs, odynophagia and rhinorrhea. gastrointestinal symptoms such as nausea and watery diarrhea were relatively rare [ ] . dyspnea develops after a median of to days from the onset of symptoms. it is important to notice that, if dyspnea is an important clinical finding, not all the patients with this j o u r n a l p r e -p r o o f symptom will develop severe respiratory distress or require oxygen supplementation [ ] . according to world health organization (who) guidelines, covid- infection can present as pneumonia without signs of severity, and could be managed in the outpatient setting. this is applicable to those patients who do not need supplemental oxygen [ ] . as previously mentioned, the most serious manifestation of covid infection is pneumonia, characterized by cough, dyspnea, and infiltrates on chest images; the latter is indistinguishable from other viral lung infections. acute respiratory distress syndrome (ards) is a major complication of covid pneumonia in patients with severe disease. this develops in % after a median of eight days. mechanical ventilation is implemented in . % of cases [ ] . in different case reports, the need for supplemental oxygen via the nasal cannula was required in approximately % of hospitalized patients. % required non-invasive mechanical ventilation, and less than % required invasive mechanical ventilation with or without extracorporeal membrane oxygenation (ecmo) [ ] . it is important to mention that the proportion of severe cases is highly dependent on the study population and may be related to the epidemiological behavior of the infection in each country. additionally, the number of people tested will influence the denominator. in italy, the average age of people infected with covid- is between and years, and % of those hospitalized require admission to the intensive care unit (icu) [ ]. the who recommendations had established that severe covid- disease could be defined by the following parameters in table [ ] . [ ] . among the established risk factors for the development of ards is age greater than years, diabetes mellitus and hypertension, in at least % of patients [ ] . it should be clarified that, although advanced age is identified as a risk factor for a severe infection, those of any age may suffer from severe illness from covid- . the descriptions made so far of the patients from china have determined that almost % of the patients were between the ages of and years (cohort of , cases) [ ] . in other population settings, such as in the united states, more than % of confirmed the clinical characteristics of symptomatic cases and their severity has been described. in addition to the symptoms reported by the patients, the findings on physical examination may be absent during mild coivd- infection. those with moderate to severe covid- infection have various signs during pulmonary auscultation, however the most common findings include: wet rales; global decrease in respiratory sounds and increased thrill [ ] . early recognition is essential to classify cases as potential cases and initiate one of the most important measures to contain the pandemic, isolation. . anyone who has resided or been traveling in areas where widespread community transmission has been reported. . any patient who has had potential exposure through attending events or has spent time in specific settings where cases of covid- have been reported. the scenarios described respond to the context of a high suspicion of covid- infection. the world health authorities (cdc, who) continually update these contexts. there have been multiple case definitions and clarifications regarding when to perform a covid- test: • they have pointed out the importance of fever, cough and dyspnea as sentinel symptoms, since these should form part of the clinical judgment that guides doctors. this allows to expand the group of suspicious patients. • in cases of severe respiratory distress of undetermined etiology and that do not meet the previously indicated criteria, a screening for covid- would be indicated. • in areas of limited resources, the suggestion is to prioritize cases that require hospital care, and in this way guide the epidemiological fence to order isolation and protect the most vulnerable people (chronically ill and over years of age), as well as test those with the greatest possibility of exposure (travelers and health personnel). currently, there is no laboratory data profile that is framed in covid infection. from a cohort of patients confirmed with covid , these findings were classified as mild, moderate and severe disease [ ] . high levels of d-dimer and more severe lymphopenia have been associated with mortality due to a prothrombotic state that determines multi-organ failure. in general, leukopenia and / or leukocytosis can be found in the interpretation of blood biometry, however, the most widely described finding is lymphopenia [ ] . it should be considered that in the context of viral pneumonia biomarkers such as procalcitonin and pcr are not useful, as often these biomarkers are in the normal range for patients with covid- . among other findings, descriptive studies have reported considerable elevations of lactate dehydrogenase and ferritin as well as alteration in aminotransferases; although elevation ranges for these parameters have not been established [ ] . about the imaging findings, covid viral pneumonia has similar features on imaging to other viral infections. although computed tomography (ct) is the test of choice, it is not useful for a definitive diagnosis due to the wide variety of images that can be found in patients with covid infection. this statement is derived from a large cohort of more than wuhan patients, where rt-pcr confirmation of covid and chest ct images of these patients were correspondingly analyzed. ct images were determined to have a sensitivity of %; however, the specificity was only % [ ] . in general, the majority of descriptive studies concur that the finding of ground glass opacifications is most common. it is typically basal and bilateral, and rarely associated with underlying consolidation. a multicenter chinese study that retrospectively reviewed the ct scans of patients found that % had typical ground-glass images and up to % had this finding along with consolidations. these findings were more frequent in the most j o u r n a l p r e -p r o o f severe and older age groups of patients [ ] . pleural effusion ( %), and lymphadenopathy ( . %) [ ] . the emergence and outbreak of sars-cov- , the causative agent of covid- , has rapidly become a global concern that highlights the need for fast, sensitive, and specific tools to monitor the spread of this infectious agent. diagnostic protocols to detect sars-cov- using real-time quantitative polymerase chain reaction (rt-qpcr) were listed on the world health organization (who) website as guidance, however, various institutions and governments have chosen to establish their own protocols that might not be publicly available or listed by who. there are important challenges associated with close surveillance of the current sars-cov- outbreak. firstly, the rapid increase of cases has overwhelmed diagnostic testing capacity in many countries, highlighting the need for a high-throughput, scalable pipelines for sample processing [ , ] . secondly, given that sars-cov- is closely related to other coronaviruses [ ] , some of the currently available nucleic acid detection assays can result in false positives [ ] . thirdly, critical concern for molecular detection is the low sensitivity reported for rt-qpcr assays [ ] and serological tests [ ] , particularly in the early stages of infection. additionally, most of the available rt-qpcr assays require sample processing and equipment only available in diagnostic and/or research laboratories. it is important to mention that coinfection is a possibility, as some reports from italy and in spain, less than % of cases in a cohort correspond to plwh, which have had a satisfactory evolution and less than half required an intensive care unit [ ] . in the us, of the , hospitalized patients in the new york area, only patients had hiv, while in san francisco, data was published on , people who had diagnosed with sars-cov- infection, of which less than % had hiv and none of them developed severe covid- [ ] . despite the existence of in vitro studies on the efficacy of the use of lopinavir / ritonavir, it is currently known that its effect in cases of moderate and severe covid- is null, and therefore at the moment no recommendation can be given nor how treatment, much less as prophylaxis [ ] . this clarification is important given that there is a belief that plwhs could be protected if they take antiretroviral therapy. therefore, current recommendations for plwhs are to maintain antiretroviral therapy with the goal of controlling hiv as well as following the same standards of care as the general population to avoid acquiring a sars-cov- infection [ ] . regarding sars-ncov infection in pregnant women, there is currently limited evidence about the effect of the virus on the mother or fetus. however, due to the physiological changes typical of pregnancy, especially on the immune system (immunosuppression) and the cardiopulmonary system, pregnant women are thought to be more susceptible to developing severe symptoms when they acquire the viral respiratory disease. in , when influenza a h n infection occurred, only % of the infected population were pregnant, yet they accounted for % of infection-related deaths [ ] . pregnancy ( %) [ ] . in another study of pregnant patients infected with mers-cov, presented adverse results ( %), neonates were admitted to the neonatal intensive care unit ( %) and three of them died ( %) [ ] . however, it is important to note the small sample size which could increase the risk of bias and low power of the study. with information obtained so far from the wuhan sars-cov- outbreak, the infection appears to be less severe for pregnant women, compared to previous sars-cov and mers-cov outbreaks [ ] . however, it is important to take into account that the data from covid- infection should be monitored with a doppler ultrasound every two weeks, due to the risk of developing intrauterine growth restriction [ , ] . the time of termination of the pregnancy, as well as the method, also depend on several factors, including gestational age, maternal condition in relation to sars-cov- infection, presence of maternal comorbidities, and fetal condition. decisions must be made collaboratively during multidisciplinary team discussions, with individualized management plans established for each patient [ ] . a diagnosis of covid- alone is not an indication for the termination of pregnancy, rather it should be made in combination with consideration of morbidity and mortality of both the fetus and mother. after delivery, the use of corticosteroids is recommended for antenatal fetal lung maturation, with betamethasone or dexamethasone [ ] ; taking special care in critically nursing patients, as this may worsen their condition, and may delay delivery, which is necessary for the management of these patients [ , ] . the symptoms children present with are similar to adults, as is the incubation period ranging from to days (mean of . ). a cough is the most frequent presenting symptom ( %) followed by fever ( %). there is a higher occurrence of gastrointestinal symptoms including diarrhea ( %), nausea, vomiting ( %) and abdominal pain. these gastrointestinal symptoms are usually more variable in children than adults and are sometimes the only clinical manifestation in associations with fevers. [ , ] . the clinical progression and disease severity in pediatric patients is markedly different from that of adults. over % of affected children are asymptomatic or have mild to moderate disease [ ] . the majority of serious cases in children are related to those with significant comorbidities such as heart disease, immunosuppression, etc. to date of this review, only a few cases of children without underlying comorbidities have died as a result of covid have been reported. this difference of severity of illness between adults and j o u r n a l p r e -p r o o f children has not been clarified, however, several theories have been postulated. these include that children express more ace receptors in their lungs which confer some protection to severe injuries such as those caused by rsv and which would decrease dramatically with age [ , ] . immunological factors may also influence outcomes, as in childhood we are most exposed to frequent challenges including recent seasonal viruses such as rsv in the winter months. most likely, it is multifactorial and depends on factors from both the host and the virus itself [ ] . abnormal radiological (ct) findings are found in asymptomatic children and consist of bilateral lung lesions ( %). elevated crp (creactive protein), procalcitonin pct ( %), and liver enzymes are present in most affected children, unlike adults in whom pct is not a reliable marker. virus elimination via the stool even after the negativity in the nasopharyngeal mucosa and the disappearance of symptoms makes them a potential source of contagion through the fecal-oral route [ ] . patients with cancer are generally more susceptible to infections than healthy people, because they have a state of systemic immunosuppression that is exacerbated during chemotherapy or radiotherapy [ ] . in china, according to national surveillance data, coronavirus infection occurs in . % of patients with malignant tumors. this is a much higher proportion than the general incidence of . % [ ] . p< . ) even after adjusting for age [ ] . further research, completed in a tertiary hospital in wuhuan -china similarly found that % of patients with cancer and sars-cov- infection died, most of them over years of age [ ] . due to these findings, it has been proposed by many international entities that during the pandemic, for prevention, an individualized plan based on the patient's specific conditions is required, with the aim to minimize the number of visits to health institutions.  for early-stage patients with need of post-surgical adjuvant chemotherapy, especially those whose clinical, pathologic, and molecular biologic staging suggest a better prognosis, the start time of adjuvant chemotherapy may be delayed up to days after surgery without affecting the overall effect of treatment [ ] .  for patients with advanced cancer, the main approach should be to minimize hospitalization in covid- positive installations. replacing the existing intravenous treatment regimen with oral chemotherapy during this special period may be considered, to ensure that treatment is not interrupted for a long time during the pandemic [ ] . however, if there is a suspicion of covid- infection in this population group, the same updated diagnostic guidelines and the corresponding management should be followed depending on their severity of illness. moreover, an individualized follow-up plan should be outlined due to higher likelihood of complications in this group of population [ ] . it should be noted that patients attending out-patient appointments for cancer have higher levels of anxiety, depression and other mental health problems than the general population. studies have shown that approximately % of malignant tumor survivors have a moderate to severe fear of tumor recurrence [ ] . for this reason, psychologist surveillance of outpatients in quarantine or during hospitalization should be considered. reported complications derived from covid- describe a severe disease that requires management in an intensive care unit (icu) in approximately % of proven infections. j o u r n a l p r e -p r o o f appear to be most susceptible to the life-threatening complications. the risk of patient-topatient transmission in the icu is currently unknown, therefore adherence to infection control precautions is paramount [ , ] . progressive deterioration of respiratory function is undoubtedly the most common and lifethreatening complication of the infection. the prevalence of hypoxic respiratory failure in covid- patients is %, and it can progress to acute respiratory distress syndrome (ards), with the need of mechanical ventilation support at . days on average. one study found that between and % of hospitalized patients require admission to the icu due to respiratory deterioration [ ] . as respiratory complications are the most common cause of severe deterioration, early identification of them will undoubtedly help in timely support. support provided should be adapted to take into account risk factors such as advanced age, neutrophilia and organic dysfunction for the development of ards. the diagnostic support of pulmonary tomography is undoubtedly a valid tool; images in patients with different clinical types of covid- have characteristic manifestations, but it can become an operational problem due to the difficulty in performing imaging on critically ill patients. on the contrary, lung ultrasound at the bed-side could provide an alternative to radiographs and tomography during the diagnosis of covid- [ , ] . since more than % of hospitalized patients will require supplemental oxygen, it is recommended that oxygen should be started when pulse oximetry values fall below %. an upper-limit of % saturation should be established, since higher values have been shown to be harmful [ , ] . hemodynamic deterioration has a variability of presentation, this depends on the study population and the definition [ ] , the presence of shock in the intensive care unit may be present between to % [ , ] . cardiomyopathy related to viral infection is one of j o u r n a l p r e -p r o o f the main causes of hemodynamic detriment, occurring in up to % of patients with covid- [ ] . hemodynamic failure is one of the main causes of death in these patients, with percentages of up to %, inconclusive risk factors are associated to date such as diabetes, hypertension, lymphopenia, and elevation of d-dimer [ ] . acute kidney injury (aki) is present in up to % of critically ill patients, podocytes and proximal tubule cells are potential host cells for sars-cov- , caused by the virus induced cytopathic effect. the diagnosis is based on markers of early kidney injury and urinary output [ ] . initial management of shock is based on fluid resuscitation, based on the application of dynamic parameters to predict response to fluids, such as variation in stroke volume (svv), variation in pulse pressure volume (ppv) and change in stroke volume with passive leg elevation or fluid challenge above static parameters [ ] . variables such as skin temperature, capillary refill time and/or serum lactate measurement are currently valid tools to assess shock. the volume of liquids used in resuscitation should be restricted and administered in relation to dynamic assessment. a liberal water resuscitation strategy is not recommended, rather a balance of crystalloids over colloids as resuscitation liquids should be encouraged and avoiding the use of hydroxyethyl starches, albumin, dextrans or gelatins [ , ] . indirect evidence suggests that the target mean arterial pressure (tam) for patients with septic shock is mmhg using vasoactive support [ ] . the recommendation of norepinephrine use as the first agent is maintained. if norepinephrine is unavailable, vasopressin or epinephrine could be used, avoiding the use of dopamine as the initial vasopressor due to the potential development of arrhythmias [ , ] . in patients with covid- and shock with evidence of cardiac dysfunction and persistent hypoperfusion despite fluid resuscitation and norepinephrine use, dobutamine as inotropic is recommended. given the development of refractory septic shock, the suggestion of the use of hydrocortisone in continuous infusion is maintained, as indirect evidence, this in favor of reducing the length of stay in the icu and the resolution time of the shock [ ] . according to the investigative mission of the who in china, the case-fatality rate ranged other reports from china have coincided with this clinical risk profile, for example, a study that included confirmed cases, patients who had ards had as main underlying diseases: diabetes and high blood pressure. of these cases, patients died [ ] . according to who, the recovery time is estimated to be two weeks for patients with mild infections and three to six weeks for those with serious illnesses. on the other hand, cdc established that people who had symptoms in the mild to moderate spectrum and maintained home isolation have a resolution of days after the fever decrease, and there was a substantial improvement in respiratory symptoms, even without use of medications. isolation may be limited to days from resolution of symptoms, however, it must be adapted to the population circumstances of the epidemic [ ] . the evolution of the epidemiological curve in covid- outbreak makes consider containment strategies in china primarily, and other countries based on nonpharmaceutical interventions (npis). according to the who, the most effective measure is hands washing [ ] . combination as public health measures reduced contact rates in the population and therefore reduce virus transmission (table ) [ ] . table non -pharmacological measures. increasing the level of hand cleanliness to % in places with a high concentration of people, like all airports in the world would have a reduction of % in the impact of a potential disease spreading [ ] . the epidemiological evolution of the covid- pandemic through phases has required the application of specific measures according to the time or phase in which the virus is found in each country. the evolution in the non-pharmacological measures has been as variable as the pharmacological ones, in such a way, since january to march, it was ensured that the use of face masks was limited only to people who had contact with epidemiological foci, not to healthy people [ ] . this concept was also reinforced by cdc, in order to optimize the use of masks for health workforce. definitely the course of the pandemic was changing rapidly, which also demanded the change from containment measures to mitigation. the recommendations in the current context remain regarding the use of a facial mask in the community, but its optimization is important for health professionals. the use of the mask is not a substitute for handwashing and social distancing measures, as these ones together allow avoiding viral particles in aerosols or drops, as a low cost and accessible measure for general population [ , ] . there is still non-specific information for the recommendation of masks, in general, having in several studies claims that surgical or cotton cloth masks do not prevent the spread of the sars-cov- virus [ ] . the evidence about the transmission of the virus in the asymptomatic period also changed the containment measures, suggesting the community use of masks. it is from this that the recommendations for the rational use of masks arise since in some j o u r n a l p r e -p r o o f countries the massive use of n masks was reported, masks indicated for the use of medical personnel [ ] . regarding to this non pharmaceutical recommendations, the studies suggest to priories the resources on vulnerable population, in endemic areas, older people, adult with comorbidities and health workforce. studies are still needed on the duration of the protective effect of the masks and above all the possibility of their reuse for resource optimization. meanwhile the most important recommendation continues to be its use in addition to hand hygiene and social distancing [ ] . therapeutic j o u r n a l p r e -p r o o f this effect is reinforced by azithromycin. there were the best results in terms of viral load reduction, even though is mentioned some limitations in the study like small sample size, a short long-term outcome follow-up, and dropout of six patients from the study [ ] . concerning to mortality rates, a study was conducted in new york with patients ( . - . ). thus, it concludes that the use of hcq is not associated with either a decreased or increased clinical impairment, intubation or death [ ] results reinforced by other study in hospitalized patients with covid- diagnosis in new york city, whom received treatment with hydroxychloroquine, azithromycin, or both drugs was not associated with significantly reduction in mortality [ ] . relating to safety of this drug, in a study carried out in patients in which the theoretical complications of the use of hcq and its combination with macrolides (azithromycin) were assessed by serial electrocardiograms, the following results were obtained. in % of patients ( ) received chloroquine, % ( ) received hcq, and % ( ) [ ] . supportive therapies in immune regulation, together with the use of antivirals, are important to take into account, especially in those patients in a serious and critical state, in which they could improve the clinical response and perhaps avoid residual lung injuries. the convalescent plasma is extracted from recovered individuals from an infection, being an antibody transfer medium to provide passive immunity (neutralizing antibodies and globulin). the goal is to provide a rapid immune response until the patient can develop their own active immune response in the hope that there will be clinical improvement [ ] . improvement in most individuals, as well as viral suppression days after treatment [ ] . in the same country, at the shenzhen hospital, cases of patients with severe covid- were reported who met criteria for acute respiratory distress syndrome (ards), who were administered convalescent plasma (titration greater than : and neutralizing antibodies greater than ). it was found that clinical recovery occurred approximately days after the transfusion ( patients) and of the patients were discharged days after admission. it is important to mention that this group of patients also received antivirals, methylprednisolone and all the necessary support measures in intensive care [ ] . other drugs like ivermectin, nitazoxanide, and others have been studied in the context of covid- treatment, but the results are inconsistent. all of the clinical trials evidence, supporting or against the use of mentioned drugs are detailed in (supplementary table ). this review summarized some drug repurposing agents previously known to has efficacy against other virus like sars-cov, mers-cov, influenza. actually, exist some new drugs with high potential action on targets for covid- therapeutics. it is important to notice that there is no specific treatment for the coronavirus approach. in context of the scientific evidence exposed and the particular clinical features of each patient, the reader will be able to make the best clinical and therapeutic decisions. when it comes to vaccine design and manufacturing, the main objectives are to ensure its safety, its efficacy in activating specific adaptive immune responses and the production ofideally-long term memory. thus, eliciting protective immune responses including neutralization antibodies and/or ctl generation is of paramount importance. huge challenges need to be tackled in order to minimize the long and cumbersome process of vaccine generation. among them, candidate antigen targets need to be identified, immunization routes and delivery systems investigated, animal models set, adjuvants optimized, scalability and production facility considered, target population selected, and vaccine safety and long-term efficiency evaluated. currently there are no approved vaccines against any human coronavirus, suggesting that their generation is quite novel. several candidate vaccines against sars-cov had shown promise reaching phase i or phase ii clinical trials [ , ] , but the rapid containment of sars-cov expansion rendered them redundant, did not allow for a test population for phase iii trials and, therefore, put their further assessment to a halt. ctl memory could last up to years after infection [ ] . these data suggest that vaccine strategies employing viral structural proteins that can elicit effective, long-term memory t cell responses could yield fruitful results. on the other hand, the s spike protein region containing the ace receptor binding domain (rdb) is the obvious option when neutralizing antibody responses are considered [ ] [ ] [ ] . indeed, a candidate sars vaccine antigen consisting of the rbd of sars-cov spike protein was created and found it could elicit robust neutralizing antibody responses and long-term protection in vaccinated animals [ ] . the fact that covid- convalescent sera shows potential as a therapeutic approach [ ] aligns with the theory that efficient b cell responses are mounted and lead to production of protective antibodies. two different groups, using an immunoinformatic approach mapped several ctl and b cell epitopes on different proteins of the virus [ , ] . moreover, various ctl epitopes were found to be binding mhc class i peptide-binding grooves via multiple contacts, illustrating their probable capacity to elicit immune responses [ , ] . consequently, these identified b and t cell epitopes could be potential targets for therapeutic vaccines. however, important safety considerations should be taken into account before releasing a new vaccine in the market. previous studies on macaque models have shown that a vaccineinduced anti-spike protein antibody at the acute stage of sars-cov infection can provoke severe acute lung injury [ ] . similar observations of sars-cov vaccine-induced pulmonary injury have also been described in multiple several murine and monkey animal models [ ] . an additional factor that needs to be checked in phase ii and iii trials is that the vaccine does not cause ade of the pathogen, as has previously been described. such concerns have risen in the context of a dengue vaccine [ ] . the pharmaceutical companies that are currently on a race to produce a vaccine for covid- along with the vaccine developing strategies they are using are summarized in table and figure . as can be easily deduced from table hospitalization and admission to already heavily charged icus due to these pathologies that could prove critical for weaker health systems that would struggle to carry the burden of combined outbreaks. moreover, vaccinating health care workers is crucial for reducing the risk of absence due to disease, thereby strengthening the healthcare workforce and minimizing the risk to infect covid- hospitalized patients with additional pneumoniacausing pathogens. lastly, covid- patients vaccinated for influenza and streptococcus pneumoniae allow their immune system to focus on one pathogen and, therefore, give it a better fighting chance against sars-cov- infection [ ] . high risk groups prioritized for vaccination for these two pathogens include pregnant women, persons with immunocompromised immune systems (either due to congenital or acquired immunodeficiencies), children, adults ≥ years and health care professionals. j o u r n a l p r e -p r o o f heat or chemical treatment inactivation. f) attenuated live pathogen vaccine strategies consist in administering a live pathogen that due to cell culture passaging has lost its virulence. they usually elicit robust and long-term memory immune responses without the need to administer an adjuvant. g) in dna vaccines the dna codifying a highly immunogenic antigen is administered and captured by professional antigen presenting cells (apcs) leading to antigen production and presentation by these cells. h) moderna's vaccine candidate already in phase i clinical trials uses an mrna vaccine approach whereby the genetic information codifying for the s protein of sars-cov- is delivered in lnps to enhance absorption by apcs. once uptaken by apcs the mrna induces the expression of s antigen that is subsequently mounted on and presented by mhc molecules to elicit adaptive immune response. numerous studies confirm that climate has an impact on virus (i.e., influenza, coronavirus, etc.) spread through manipulating the conditions of i) its diffusion, ii) the virus survival outside the host, and iii) the immunity of host population [ ] . meteorological conditions, such as temperature, humidity, wind speed and direction, atmospheric pressure, solar radiation (including ultraviolet (uv) spectrum) and precipitation amount and intensity depend on the latitude and the elevation of the location, thus creating distinct climatic zones in the planet. while in some regions, such as temperate climate zones, human influenza peaks have clear seasonal cycles, in others it is not as predictable [ ] [ ] [ ] [ ] [ ] . an array of studies, investigating the relationship between climatic factors and the activity of influenza all over the world, concluded that at the high latitudes of the world the peaks of influenza correlate with cold and dry weather conditions (i.e., winter season), while around the equatorial zone, it is more common during the months of high humidity and precipitation [ ] [ ] [ ] [ ] [ ] [ ] [ ] . essentially, it depends on explicit threshold conditions based on monthly averages of specific humidity and temperature. when specific humidity drops below - g/kg and temperature drops below - °c, the peak of influenza is stimulated during the cold-dry season, however, for tropical and subtropical (always humid and warm) regions, it is likely to 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fengyu; li, feng; deng, xilong title: prognostic factors for covid- pneumonia progression to severe symptoms based on earlier clinical features: a retrospective analysis date: - - journal: front med (lausanne) doi: . /fmed. . sha: doc_id: cord_uid: q p kg approximately – % of covid- patients will develop severe pneumonia, and about % of these will die if not properly managed. earlier discrimination of potentially severe patients basing on routine clinical and laboratory changes and commencement of prophylactical management will not only save lives but also mitigate the otherwise overwhelming healthcare burden. in this retrospective investigation, the clinical and laboratory features were collected from covid- patients who were classified into mild ( cases) or severe ( cases) groups according to their clinical outcomes after – days post-admission. the subsequent analysis with single-factor and multivariate logistic regression methods indicated that factors on admission differed significantly between mild and severe groups but that only comorbidity with underlying diseases, increased respiratory rate (> /min), elevated c-reactive protein (crp > mg/l), and lactate dehydrogenase (ldh > u/l) were independently associated with the later disease development. finally, we evaluated their prognostic values with receiver operating characteristic curve (roc) analysis and found that the above four factors could not confidently predict the occurrence of severe pneumonia individually, though a combination of fast respiratory rate and elevated ldh significantly increased the predictive confidence (auc = . , sensitivity = . , and specificity = . ). a combination consisting of three or four factors could further increase the prognostic value. additionally, measurable serum viral rna post-admission independently predicted the severe illness occurrence. in conclusion, a combination of general clinical characteristics and laboratory tests could provide a highly confident prognostic value for identifying potentially severe covid- pneumonia patients. approximately - % of covid- patients will develop severe pneumonia, and about % of these will die if not properly managed. earlier discrimination of potentially severe patients basing on routine clinical and laboratory changes and commencement of prophylactical management will not only save lives but also mitigate the otherwise overwhelming healthcare burden. in this retrospective investigation, the clinical and laboratory features were collected from covid- patients who were classified into mild ( cases) or severe ( cases) groups according to their clinical outcomes after - days post-admission. the subsequent analysis with single-factor and multivariate logistic regression methods indicated that factors on admission differed significantly between mild and severe groups but that only comorbidity with underlying diseases, increased respiratory rate (> /min), elevated c-reactive protein (crp > mg/l), and lactate dehydrogenase (ldh > u/l) were independently associated with the later disease development. finally, we evaluated their prognostic values with receiver operating characteristic curve (roc) analysis and found that the above four factors could not confidently predict the occurrence of severe pneumonia individually, though a combination of fast respiratory rate and elevated ldh significantly increased the predictive confidence (auc = . , sensitivity = . , and specificity = . ). a combination consisting of three or four factors could further increase the prognostic value. additionally, measurable serum viral rna post-admission independently predicted the severe illness occurrence. in conclusion, a combination of general clinical characteristics and laboratory tests could provide a highly confident prognostic value for identifying potentially severe covid- pneumonia patients. keywords: covid- , sars-cov- , risk factor, clinical manifestation, prognostic factor background the novel coronavirus (sars-cov- ) has seemed to sweep across the globe ever since its first successful jump from bat to human being through a still unknown intermediate(s) in approximately late nov ; it still shows a tendency toward significant surges in incidence worldwide ( ) ( ) ( ) . the sars-cov- virus seems more contagious than its sibling virus, severe acute respiratory syndrome (sars) virus, which broke out in ; as of march , over , individuals have contracted covid- pneumonia within months, which was about times that of the total sars cases ( , in months) ( ). the surging increase in covid- patients within a short time window will severely impact the limited medical resources, including physicians, nurses, protective suits, masks, and goggles. data from the chinese mainland showed that the majority of total infected patients will recover under simple supervision management, such as quarantine in a compartment hospital isolation ward, but that the overall case fatality rate was . % ( ) . for the clinical treatment of covid- patients under shortage of enough medical supplies, the critical issues and priorities are to treat the severe covid- patients [about % of the whole population ( ) ] and to save their lives with preventive and intensive medical care. however, the clinical presentation of covid- patients differs substantially, and this can include asymptomatic infection, mild upper respiratory tract illness, and severe viral pneumonia ( , ( ) ( ) ( ) . the most crucial issue is therefore to identify these patients and prioritize their treatment strategy by applying prophylactic medical treatment and management before they progress to the severe stage. as we know, respiratory function worsens in the severe stage. in clinical practice, saturated oxygen (< % in rest state), reparatory rate (> times/min), and deteriorated chest radiology imaging (x-ray and ct more high resolution) provide references to confirm their severity ( , , ) . because of the hypoxia stress, most patients will experience an over-reactivated immune storm, including elevated expression levels of some specific immunological cytokines and changes in certain types of immune cell counts ( , ) . biopsy analysis also showed that the lung bilateral diffuse alveolar damage with cellular fibromyxoid exudates ( ) . however, ct imaging and immunology detection is not only expensive but also largely unavailable an unable to cope with the significant rise in suspected cases, particularly in those hospitals that are not well-equipped. can some routine clinical characteristics or/and laboratory measurements (or their combination) predict the occurrence of severe cases? in this study, we retrospectively analyzed the clinical characteristics of those patients who progressed to severe pneumonia later and found that five simple clinical features and laboratory detection at an earlier time point could serve as prognostic factors facilitating discrimination of severe cases in advance. covid- diagnosis was determined according to the criteria in the new coronavirus pneumonia diagnosis and treatment plan (trial version ) issued by the national health and health commission ( ). all covid- patients admitted to guangzhou eighth people's hospital from january to february , , were included in this study. this study complied with the medical ethics of guangzhou eighth people's hospital. we obtained written consent from the patients. for this analysis, inclusion criteria were the following: ( ) diagnosed as mild or ordinary on admission and ( ) length of hospitalization > days and overall duration of the disease > days. qualified patients were then classified into a mild symptom group and a severe symptom group based on the clinical manifestation. the severe symptom diagnosis was determined according to the following criteria: ( ) respiratory distress, rr ≥ times/min in the resting state; ( ) oxygen saturation ≤ % in the resting state; and ( ) arterial blood oxygen partial pressure (pao )/oxygen concentration (fio ) ≤ mmhg). the rest of the patients were in the mild group. patient general information, including gender, age, underlying diseases, epidemic history, etc., and their clinical data including symptoms, signs, clinical classification (course duration > days), laboratory test results, and sars-cov- viral test results were obtained with standardized data collection forms from electronic medical records. quantitative data was firstly tested to be normality distribution with the kolmogorov-smirnov method. then, for normalized distributed data, t-test and tamhane t methods were used for variance of even and uneven data, respectively. for nonnormal data, which was expressed as the median (quartile) [m (p , p )], the mann-whitney u-test was employed. the chi-square test (or fisher exact probability method) was utilized for analyzing qualitative data. logistic regression analysis and the receiver operating characteristic curve (roc) analysis was employed to analyze the independent risk factors. the difference was statistically significant at p < . . all analysis was performed using spss software (version . ). a total of covid- cases (about % of total cases in guangzhou, china) were admitted to guangzhou eighth people's hospital for treatment from january to february , ( figure ). according to the inclusion criteria, cases were excluded because cases were already in the severe symptom stage, cases had a short hospitalization time of < days, and patients had other defects, such as being short of a complete set of detection. finally, cases, including males and females, were qualified to be included for further investigation, and all their disease courses were over days, with a maximum of days. based on the severity of disease at days post-admission, patients fell in the mild group ( general cases and mild cases) and patients in the severe group ( severe cases and seven critical cases). all included patients were aged between . and years (averaged . ± . years) ( table ) . among them, cases had at least one underlying disease, including cases with hypertension, eight cases with diabetes, five cases with coronary heart disease, two cases with chronic obstructive pulmonary disease, two cases with chronic kidney disease, two cases with chronic liver disease, and two cases with sleep apnea syndrome. five individuals with two or more basic disorders and cases with obesity (bmi > ). epidemiologically, cases had a history of traveling to or living in the hubei epidemic area before disease onset. interestingly, we observed that seven patients developed serum sars-cov- viral rna positive after admission but ahead of diagnosis to be a severe symptom. the single-factor analysis was applied for each factor between the mild group and the severe group ( table ) . more patients in the severe group were old, obese (bmi > ), and had underlying diseases, particularly hypertension and diabetes (p < . ), compared with the mild group. among the general factors, no significant difference could be seen with regards to gender, history of traveling to or living in an epidemic region, coughing, sneezing, muscle joint pain, headache, fatigue, and gastrointestinal symptoms between these two groups (p > . ). however, more patients in the severe group exhibited high fever, chest tightness, and shortness of breath (fast respiratory rate) (p < . ). the serum concentration of c-reactive protein, procalcitonin, d-dimer, albumin, and lactate dehydrogenase (ldh) increased significantly in the severe group (p < . ). compared to the mild group, patients in the severe group had lower absolute lymphocyte counts, higher eosinophil counts (p < . ), and similar levels of other parameters, including white blood cells, neutrophils, platelets, hemoglobin, prothrombin time, activated partial thromboplastin time, blood lactic acid, blood creatinine, and creatine kinase. interestingly, the levels of glutamate aminotransferase (alt) and aspartate aminotransferase (ast) significantly increased for severe patients (p < . ). however, the median values of alt and ast were still within the normal range, indicating that most of the severe covid- patients had no significant liver damage. importantly, all seven patients with the presence of sars-cov- viral rna in blood during the hospitalization, but before being in the severe stage, finally progressed to the severe stage; they included two severe cases and five critical cases (p < . ). next, all categorical variables were converted into covariates, including age, presence of underlying diseases (yes or no), hypertension (yes or no), diabetes (yes or no), obesity (yes or no), temperature (< . , . - . , > . • c), fast respiratory rate (yes or no), elevated c-reactive protein (> mg/l), decreased lymphocyte count (< . * e /l) and eosinophil count (< . * e /l), elevated procalcitonin (> . ng/l), elevated d-dimer (>= . µg/l), decreased albumin (< g/l), and elevated lactate dehydrogenase (ldh, > u/l), and were then subjected to single-factor logistic regression together with multiple independent variables. those variables with statistical significance were chosen for subsequent binary logistic regression analysis to test the model coefficients, goodness-of-fit, and multicollinearity. four factors identified to be significantly relevant to the severity of covid- were underlying diseases (x ), fast respiratory rate (> times/min) (x ), elevated creactive protein level (crp > mg/l) (x ), and elevated lactate dehydrogenase level (ldh > u/l) (x ) ( table ) . finally, the multifactor logistic regression equation was obtained: p = − . + . x + . x + . x + . x . the β values and odds ratios (or) for each factor are shown in table . the result indicated that elevated ldh ranks as having the highest correlation to severe symptom development (or = . ), followed by the fast respiratory rate (or = . ), underlying diseases (or = . ), and elevated crp (or = . ). to better evaluate the prediction capacity of each of the independent risk factors, we plotted their receiver operating characteristic curve (roc) for the development of severe covid- pneumonia and calculated the area under the roc curve (auc value), sensitivity, specificity, cut-off value, youden index, and p-value ( table ) for all of them. according to the general standard that auc values between . and . mean a medium level of diagnostic values and auc values over . mean a high level of diagnostic values, we observed that all the factors (auc < . ) failed to provide a high prognostic value when used alone. a two-factor combination test then showed that the combination of fast respiratory rate and elevated ldh could provide a highly confident prediction (auc = . , sensitivity = . , and specificity = . ) ( table ). the auc values of elevated ldh plus underlying diseases or plus elevated crp were both over . , but their sensitivity or specificity was lower than . . then, triple factor combination significantly increased the prognostic efficacy, and all combinations had increased sensitivity and specificity (table ) . finally, we calculated the prognostic value of the combination of all four factors and found that the auc value was significantly increased to . ( % ci . - . ), the sensitivity to . , and the specificity to . ( table ) . our study showed that underlying disease, fast respiratory rate (> times/min), elevated serum c-reactive protein level (crp, > mg/l), and elevated lactate dehydrogenase level (ldh, > u/l) were four independent risk factors for predicting the progression of some covid- patients from mild to severe conditions. firstly, elevated lactate dehydrogenase levels ranked as number (or = . ) and fast respiratory rate as number (or = . ) among the four factors ( table ) . interestingly, an elevated lactate dehydrogenase level was associated with severe sars infection ( ) , which broke out in , but was absent in the severe mers infection ( ) , which is still circulating. when used individually, all four factors have a moderate prediction value for their low specificity and sensitivity (auc values < . ) ( table ) . secondly, we found that the combination of two factors, fast respiratory rate plus elevated ldh, could provide a high prognostic value for severe symptom development (auc = . , sensitivity = . , and specificity = . ). combinations of triple factors could significantly increase the prognostic value (auc > . ). finally, a combination of all four factors, provide an excellent prognostic efficacy, achieving auc = . ( % ci . - . ) with high sensitivity ( . ) and specificity ( . ). our hospital has treated over % of covid- patients in guangzhou city-− cases as of february , -including severe cases but only one death case. all the patients except two patients recovered as of march . a retrospective analysis of all the cases revealed that the extremely low fatality rate in our hospital, one of cases ( . %)-significantly lower than the overall fatality rate ( . %) in china ( ), was largely attributed to the effect of an expert panel, consisting of physicians from multiple disciplines, including infectious diseases, respiratory diseases, and intensive care unit (icu), and radiology. patients newly admitted were reviewed by the panel, and patients who meet several of the following criteria were transferred immediately to the icu isolation ward for close supervision, including, ( ) the illness onset has entered - days; ( ) over years old; ( ) obesity, pregnant women, children; ( ) with underlying diseases, especially hypertension, diabetes, copd; ( ) fast respiratory rate; ( ) obvious decline in spirit and appetite; ( ) significant reduction and/or progressive decline of peripheral blood lymphocytes; ( ) decrease in albumin; ( ) elevated creactive protein; ( ) elevated lactate dehydrogenase; and ( ) quickly deteriorated or with two or more lesions in lungs revealed by chest imaging. once they progressed to the severe stage, they received treatment immediately. the above four prognostic factors, as routine and affordable clinical characteristics, were included in these criteria, and their immediate and preventive therapies were facilitated retrospectively. all seven patients who were detected to be serum viral rna positive developed severe symptoms very soon, which further confirmed our previous observation that detectable -ncov viral rna in blood is a reliable indicator for further clinical severity ( ) . however, as the viral rna positive rate was low high (seven of cases, . %) in this study and other reports ( ) and viral rna detection is expensive, we do not recommend the continuous detection of viral rna. in this regard, we suggest reserving the precious reagent for confirming virus infection. in conclusion, our study indicated that underlying disease, a fast respiratory rate, elevated serum c-reactive protein level, and elevated lactate dehydrogenase level significantly correlated to the development of severe covid- pneumonia; additionally, elevated lactate dehydrogenase and a fast respiratory rate (possibly plus one or two more other factors) can serve as prognostic factors for the discriminating potential severe cases among the mild covid- patients. our study provided convenient, reliable, and affordable references for both patients and physicians to make a highly confident decision to commence management and treatment safely. with our successful experience of treating covid- patients, we retrospectively found that routine clinical features could reliably predict severe pneumonia development and could thus provide quick and affordable references for physicians to save patients with otherwise fatal covid- using their limited medical resource. the original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding authors. ethical review and approval was not required for the study on human participants in accordance with the local legislation and institutional requirements. the patients/participants provided their written informed consent to participate in this study. hh, fl, and xd conceived the study and wrote the manuscript. hh and sc collected data and performed the data analysis. hh, sc, yul, yol, yf, and xd participated in the clinical treatment. ll, cl, and xt supervised the clinical treatment. fh analyzed the results. all authors read the manuscript and approved the final version. all authors contributed to the article and approved the submitted version. world health organization. coronavirus disease (covid- ) situation reports epidemiological and clinical characteristics of cases of novel coronavirus pneumonia in wuhan, china: a descriptive study early transmission dynamics in wuhan, china, of novel coronavirus-infected pneumonia recombination and insertion in the evolution of -ncov. biorxiv vital surveillances: the epidemiological characteristics of an outbreak of novel coronavirus diseases (covid- ) -china clinical features of patients infected with novel coronavirus in wuhan clinical characteristics of hospitalized patients with novel coronavirus-infected pneumonia in wuhan clinical characteristics of coronavirus disease in china imaging and clinical features of patients with novel coronavirus sars-cov- initial ct findings and temporal changes in patients with the novel coronavirus pneumonia ( -ncov): a study of patients in wuhan, china aberrant pathogenic gm-csf+ t cells and inflammatory cd +cd + monocytes in severe pulmonary syndrome patients of a new coronavirus notice on issuing a new coronary virus pneumonia diagnosis and treatment plan (trial version clinical and laboratory features of severe acute respiratory syndrome vis-a-vis onset of fever predictive factors for pneumonia development and progression to respiratory failure in mers-cov infected patients detectable -ncov viral rna in blood is a strong indicator for the further clinical severity detection of sars-cov- in different types of clinical specimens prognostic factors for covid- pneumonia progression to severe symptom based on the earlier clinical features: a retrospective analysis. medrxiv the authors would like to thank all nurses in the treatment team for taking care of the patients and doctors in the expert panel for their treatment guidance. this manuscript has been released as a preprint at medrxiv ( ) . the contents of this article are solely the responsibility of the authors and do not necessarily represent the views of any organization. the authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.copyright © huang, cai, li, li, fan, li, lei, tang, hu, li and deng. this is an open-access article distributed under the terms of the creative commons attribution license (cc by). the use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. no use, distribution or reproduction is permitted which does not comply with these terms. key: cord- - qh pblc authors: quah, jessica; jiang, boran; tan, poh choo; siau, chuin; tan, thean yen title: impact of microbial aetiology on mortality in severe community-acquired pneumonia date: - - journal: bmc infect dis doi: . /s - - - sha: doc_id: cord_uid: qh pblc background: the impact of different classes of microbial pathogens on mortality in severe community-acquired pneumonia is not well elucidated. previous studies have shown significant variation in the incidence of viral, bacterial and mixed infections, with conflicting risk associations for mortality. we aimed to determine the risk association of microbial aetiologies with hospital mortality in severe cap, utilising a diagnostic strategy incorporating molecular testing. our primary hypothesis was that respiratory viruses were important causative pathogens in severe cap and was associated with increased mortality when present with bacterial pathogens in mixed viral-bacterial co-infections. methods: a retrospective cohort study from january to july was conducted in a tertiary hospital medical intensive care unit in eastern singapore, which has a tropical climate. all patients diagnosed with severe community-acquired pneumonia were included. results: a total of patients were in the study. microbial pathogens were identified in ( . %) patients. mixed viral-bacterial co-infections occurred in ( . %) of patients. isolated viral infections were present in patients ( . %); isolated bacterial infections were detected in patients ( . %). hospital mortality occurred in ( . %) patients. the most common bacteria isolated was streptococcus pneumoniae and the most common virus isolated was influenza a. univariate and multivariate logistic regression showed that serum procalcitonin, apache ii severity score and mixed viral-bacterial infection were associated with increased risk of hospital mortality. mixed viral-bacterial co-infections were associated with an adjusted odds ratio of . ( % ci . – . , p = . ) for hospital mortality. conclusions: respiratory viruses are common organisms isolated in severe community-acquired pneumonia. mixed viral-bacterial infections may be associated with an increased risk of mortality. the microbial aetiology of severe community-acquired pneumonia (cap) remains varied throughout the world, with influences due to seasonal climate change, outreach of vaccination programmes and pathogen endemicity [ , ] . two decades ago, it was thought that viruses played a minor role in the pathogenesis of severe cap, notwithstanding influenza epidemics [ ] . recent literature contradicts this and suggests that viruses are frequently found in severe cap [ , ] . advances in molecular techniques have improved the sensitivity, accuracy and turnaround time of microbial diagnostic tests [ , ] . the availability of highly multiplexed commercial tests kits for common viral and bacterial pathogens has enabled these tests to be performed in large numbers of patients simultaneously, and across a variety of clinical settings [ , ] . multiple respiratory viruses may be present concurrently, or co-exist with bacterial pathogens to cause disease [ ] [ ] [ ] . the reported incidence of viruses in severe cap resulting in respiratory failure ranges . % to % [ , ] . this variation in the detection rate of viruses reflects potential limited availability of test assays and heterogeneity of physician practices in viral microbial diagnostic tests performed [ , ] . postulated prohibitive factors against the routine performance of viral diagnostics tests in patients with severe cap may include a lack of clear clinical guidelines, perceived low cost-effectiveness and the paucity of effective anti-viral therapies for respiratory viruses other than influenza. the primary aim of our study was to determine the risk association of microbial aetiologies with hospital mortality in severe cap, utilising a diagnostic strategy that incorporated molecular testing. our primary hypothesis was that respiratory viruses were important causative pathogens in severe cap and was associated with increased mortality when present with bacterial pathogens in mixed viral-bacterial co-infections. this was a retrospective cohort study performed in an -bed medical intensive care unit of a -bed tertiary teaching hospital, in singapore. ethics approval was obtained from the singhealth centralised institutional review board (cirb / /fp), with waiver of consent. adults above the age of admitted to the medical intensive care unit with severe cap from january to july were included. written and electronic medical records were reviewed. cap was defined as an acute infection of the lung parenchyma associated with acute chest radiographic infiltrates and or more of the following: fever of °c or higher or hypothermia of °c or less; a new cough; dyspnea not explained by other reasons; worsening cough or change in respiratory secretions in a patient with pre-existing chronic cough. these symptoms should have been present at the time of, or within h of, hospital admission. chest radiographs reported by the hospital radiologists were obtained to confirm the presence of radiographic pulmonary infiltrates. patients with shock requiring vasopressor support, mechanically ventilated or who have out of minor criteria for pneumonia severity as defined by the infectious diseases society of america/american thoracic society, are considered to have severe cap [ ] . the minor criteria include: respiratory rate of breaths per minute or greater; pao /fio ratio equal or less than ; multi-lobar pulmonary infiltrates; confusion or disorientation; blood urea nitrogen equal or greater than mg/dl; leukopenia with white blood cell count of less than , cells/mm ; hypothermia indicated by core body temperature less than °c; hypotension requiring aggressive fluid resuscitation. all patients with prior hospitalisation within days of study enrolment; on active chemotherapy for neoplastic diseases; receiving renal replacement therapy by haemodialysis and prisoners were excluded. immunocompromised patients were not excluded as they were likely to have cap microbial pathogens as well as opportunistic infections. the time of admission to the intensive care unit was verified with electronic records. all patients had at least set of aerobic and anaerobic blood cultures performed within h of admission. routine collection of endotracheal aspirates for gram stain and semi-quantitative bacterial cultures occurred within h of intubation. viral and atypical pathogen polymerase chain reaction (pcr) amplification tests were collected from endotracheal aspirates. sputum samples were sent for bacterial aerobic cultures, while nasopharyngeal swabs were performed for atypical pathogen and viral pcr amplification tests for subjects who did not require intubation. urinary samples were tested for the presence of urinary streptococcus pneumoniae antigen and legionella pneumophilia serogroup antigen in patients without anuria. where indicated, acid-fast staining and mycobacterial cultures of respiratory samples were performed. nucleic acid was extracted from swabs or respiratory samples using ez virus mini kit (cat no. , qiagen, germany) performed on a semi-automated system (ez , qiagen, germany), and stored at - °c for more detailed molecular testing. besides routine standard clinical testing, additional multiplex real-time pcr testing was performed for each extracted sample. anyplex written and electronic medical records were reviewed for clinical data and laboratory indices. the most severe value was recorded for analysis if any blood test was repeated more than once within h after intensive care admission. serum c-reactive protein was measured by particle enhanced immunoturbidimetry (cobas® crpl ) and serum procalcitonin was measured using sandwich immunoassay (cobas® elecsys brahms pct). variables such as the acute physiology and chronic health evaluation ii (apache ii) severity score, presence of shock, mechanical ventilation, intensive care unit length of stay, and mortality were captured prospectively as part of administrative clinical care audits. empirical antibiotics were deemed to be adequate if it adhered to the hospital antimicrobial guidelines. combination therapy with beta-lactams and macrolides were recommended. where melioidosis was suspected, carbapenems were preferred to other classes of beta-lactams. categorical variables were expressed as number (percentages) and normally distributed quantitative variables were expressed as mean (± standard deviation). categorical variables were compared with chi-square test or fisher's exact test, and quantitative variables were compared with t-test or mann-u whitney test. the primary outcome measure was all-cause hospital mortality. demographic and disease variables were included in the univariate and multivariable logistic regression model. pneumonia symptoms were excluded from regression analysis as there were no known associations with mortality in severe cap. the variable for number of co-existent pathogens was excluded from regression analysis due to significant overlap with mixed viral-bacterial co-infections, which was included in the multivariate model. the following severity indicators: shock, altered mentation and ventilator support were not included as these variables are incorporated in the apache ii severity score. mycobacterium tuberculosis was classified as a bacterium for the logistic regression. demographic and disease variables were then compared between patients with and without virus infections, for identification of potential risk factors associated with acquisition of respiratory viral infections. a p-value of < . was considered significant. missing data was excluded from univariate and multivariate analysis. stata special edition version . (statacorp llc, texas, usa) was used for statistical analysis. one hundred seventeen patients were admitted to the medical intensive care unit for severe cap within the study period. baseline characteristics of patients who suffered in-hospital mortality (n = , . %) were compared with patients who survived, in table . of note, the patients who did not survived were older compared to survivors ( . years vs . years, p = . ). other baseline demographic and co-morbidity variables were not significantly different. blood cultures, respiratory specimen cultures and respiratory specimen pcr testing for viruses and atypical bacteria were performed in all patients. results of microbiological tests are presented in table . causative microbial aetiologies were identified in . % of patients with specific organisms presented in table . majority of pathogens identified were respiratory viruses and bacteria. viruses were found in . % of patients (n = ) with the most common virus being influenza a. . % (n = ) of patients with influenza a received empirical oseltamivir on the basis on clinical suspicion. bacterial infections were found in . % of patients (n = ). . % of patients (n = ) had mixed virus and bacterial co-infections. patients ( . %) had only viral pathogens and patients ( . %) had only bacterial pathogens, found as the microbial aetiology for severe cap. using hospital mortality as the primary outcome, univariate logistic regression was performed (table ). patients who did not survive had a higher apache ii score and higher serum procalcitonin levels. the microbiological aetiology that was significantly associated with increased hospital mortality was the detection of mixed viral-bacterial co-infections. on multivariate analysis, apache ii severity score, serum procalcitonin and mixed viral-bacterial co-infections remained significantly associated with increased adjusted odd ratios for hospital mortality. while apache ii severity score is known to be predictive for mortality in severe cap, the study found that the presence of mixed viral-bacterial co-infections was associated with increased hospital mortality by an adjusted odds ratio of . ( % confidence interval . , . , p = . ). the patients with respiratory viruses detected (both isolated viral pathogens and mixed viral-bacterial co-infections) were compared with patients who did not have any respiratory virus infections, in table . the mean serum c-reactive protein was found to be greater ( . ± . mg/l vs. . ± . mg/l) in patients without respiratory viruses compared to patients with detection of respiratory viruses (table ). respiratory virus infection as a cause of severe acute respiratory distress syndrome is well-established in literature data was missing for patient [ ] [ ] [ ] . however, its significance as a contributory pathogen in the outcomes of severe cap is uncertain. in this study, we showed that respiratory viruses were as commonly found as bacteria ( . % vs . %), as an aetiological pathogen. mixed viral-bacterial co-infections occurred in . % of patients and was associated with an adjusted odds ratio of . for hospital mortality. the impact of respiratory viruses on the prognosis of severe cap remains unclear with recent studies demonstrating contradictory results. fisher et al., in a prospective -year microbiologic survey of nosocomial pneumonia and cap complicated by respiratory failure, showed that . % of patients had viral infections, which was associated with a hospital mortality of . % [ ] . however, siow et al., found that viral infections were independently associated with lower hospital mortality compared to other microbial aetiologies, with an adjusted odds ratio of . (ci . - . ; p = . ) [ ] . in light of these findings, accurate characterisation of the impact of microbial aetiology on the outcomes of severe cap is required to influence future development of rapid molecular diagnostics assays and novel antimicrobial therapies that would target both viruses and bacteria [ , ] . piralla et al. reviewed the microbiological data of severe cap in northern italy during winter-spring seasons over years and found that . % of patients had one or more respiratory viruses identified [ ] . the most common viruses isolated were influenza a and rhinovirus, similar with our findings. while our study was performed in a tropical country, local microbiological surveillance has shown that influenza epidemics occur twice annually [ ] . this would mean that influenza seasons occurred over the course of this study. the molecular mechanisms in the viral pathogenesis of severe pneumonia are most well studied in influenza a and streptococcus pneumoniae co-infections. viral infections alter host immune responses that increase susceptibility to bacterial infection through viral-induced interferons [ ] [ ] [ ] . on clinical suspicion alone, only . % (n = ) of patients with influenza a in this study received empirical oseltamivir. the authors postulate that incorporation of early influenza a diagnostic tests may decrease the time to effective anti-viral therapies. the second most common virus detected in our study was rhinovirus (n = ). the association of rhinovirus with severe pneumonia has previously been shown in a surveillance program for middle east respiratory syndrome coronavirus in saudi arabia [ ] . its genotypes a to c are associated with severe pneumonia, with in-hospital mortality rates from . to . % [ ] . patients who are immunocompromised or who have chronic lung disease are most at risk [ ] . there are several reasons why viral infections may have been less prominent as a cause of severe cap in prior decades. firstly, grève et al. performed a prospective observational study on physician practices in the use of respiratory virus diagnostics demonstrating that despite clinical guideline recommendations on testing of respiratory viruses during influenza season, less than half of patients admitted to the intensive care unit with pneumonia were tested for viral pathogens [ ] . this may have led to under-recognition of the true significance of viral pathogens and mixed viral-bacterial infections, on outcomes in severe pneumonia. other factors which may have contributed to underdiagnosis of viral pneumonias include the unavailability or cost of molecular diagnostic assays, and the lack of effective anti-viral therapies [ ] . however, the authors argue that in this age of globalisation, highly virulent respiratory viruses have the potential to spread rapidly. constant surveillance is required to detect outbreaks and for the implementation of isolation precautions in a timely manner [ , ] . understanding the significance of respiratory viruses in the pathogenesis of severe cap would guide administrators with resource allocation when implementing vaccination programs for at-risk populations. the high rates of compliance with performing respiratory aerobic cultures, blood aerobic and anaerobic cultures in this study, were in accordance with sepsis guidelines [ ] . the most common bacterial pathogen found was streptococcus pneumoniae ( . %), which is consistent with the known epidemiology of cap globally [ ] [ ] [ ] . gadsby et al., in a prior study, was able to demonstrate a bacterial yield of . % when respiratory specimens from patients with cap were tested with bacterial multiplex pcr [ ] . incorporating the use of bacterial multiplex pcr in future studies, may increase the rate of bacteria detection, and shed light on potential molecular synergisms between specific viruses and bacteria in the pathogenesis of severe cap. pulmonary tuberculosis is endemic in the region where this study was performed. in our study, patients had tuberculosis, one of whom had concomitant adenovirus infection while another had streptococcus pneumoniae. the initial presentation of pulmonary tuberculosis with clinical features consistent with severe cap has been described by tseng et al. [ ] , where % of patients with pulmonary tuberculosis presented with respiratory failure or septic shock. the authors postulate that pulmonary tuberculosis may play a role in increasing host susceptibility to severe infection with cap organisms. the authors recognise that there were several limitations to this study. firstly, while first-dose antibiotics would have been administered as soon as sepsis is identified, we are unable to accurately define the time between administration of antibiotics and collection of specimens for microbiological assessment. this may affect the yield [ , , ] . secondly, the inherent retrospective nature of this study increases the risk of bias in data collection. however, as part of pre-established intensive care unit clinical audits and with the availability of national health records, clinical data such as participant demographics, co-morbid illnesses and severity indicators such as apache ii were established at the time of intensive care admission and stored prospectively. thirdly, given the high population density of the country ( rd in the world) where this study was performed, the microbial epidemiology of severe cap may only be extrapolated to urban settings. the study is a single-centre survey conducted in of the acute general hospitals serving a population of . million in a land area of . km . hence, the epidemiology may lack generalisability when extrapolated to other tropical countries. the fourth limitation is that we included patients who were immunocompromised with typical cap organisms and survived. they may potentially have had opportunistic infections that were not detected, however, as they did not contribute to the mortality outcomes, we felt that the microbiological data contributed by these patients were useful in the understanding of the prevalence of various data are presented as number (%), mean ± standard deviation apache acute physiology and chronic health evaluation a data was missing for patients. b data was missing for patient classes of cap organisms. lastly, another potential limitation was that vaccination records could not be retrieved, and we were unable to ascertain its influences on microbial aetiologies of severe pneumonia in our study population. the main strength of the study is the characterisation of the epidemiology of microbial pathogens in severe cap. we were able to show that in a tropical environment, the viral and bacterial pathogens associated with severe cap were similar with regions with a seasonal climate. despite a lower-than-expected mortality rate for severe cap in our study ( . %) compared with international data [ ] [ ] [ ] [ ] , we were able to demonstrate that mixed viral-bacterial co-infections were independently associated with hospital mortality. respiratory viruses are important causative pathogens in severe cap and are associated with increased risk of mortality when present with bacterial pathogens in mixed viral-bacterial co-infections. abbreviations apache ii: acute physiology and chronic health evaluation ii; cap: community-acquired pneumonia; pcr: polymerase chain reaction etiology and outcome of severe community acquired pneumonia in immunocompetent adults etiology of community-acquired pneumonia and diagnostic yields of microbiological methods: a -year prospective study in norway epidemiology of community-acquired respiratory tract infections in adults the intensive care global study on severe acute respiratory infection (ic-glossari): a multicenter, multinational, -day inception cohort study viral-bacterial coinfection affects the presentation and alters the prognosis of severe community-acquired pneumonia molecular diagnosis of respiratory viruses the filmarray® respiratory panel: an automated, broadly multiplexed molecular test for the rapid and accurate detection of respiratory pathogens rapid viral diagnosis for acute febrile respiratory illness in children in the emergency department clinical utility of on-demand multiplex respiratory pathogen testing among adult outpatients burden of respiratory viruses in patients with acute respiratory failure community-acquired polymicrobial pneumonia in the intensive care unit: aetiology and prognosis clinical characteristics and outcomes in hospitalized patients with respiratory viral co-infection during the h n influenza pandemic management of severe communityacquired pneumonia: a survey on the attitudes of physicians in iberia and south america clinical practice of respiratory virus diagnostics in critically ill patients with a suspected pneumonia: a prospective observational study infectious diseases society of america/american thoracic society consensus guidelines on the management of community-acquired pneumonia in adults acute management and long-term survival among subjects with severe middle east respiratory syndrome coronavirus pneumonia and ards virus-induced acute respiratory distress syndrome: epidemiology, management and outcome influenza a (h n ) vs non-h n ards: analysis of clinical course a prospective one-year microbiologic survey of combined pneumonia and respiratory failure the use of polymerase chain reaction amplification for the detection of viruses and bacteria in severe communityacquired pneumonia the past decade in bench research into pulmonary infectious diseases: what do clinicians need to know? laboratory diagnosis of pneumonia in the molecular age frequency of respiratory viruses among patients admitted to intensive care units in seven consecutive winterspring seasons ( - ) in northern italy characterization of influenza activity based on virological surveillance of influenza-like illness in tropical singapore the immunology of influenza virusassociated bacterial pneumonia integrated clinical, pathologic, virologic, and transcriptomic analysis of h n influenza virus-induced viral pneumonia in the rhesus macaque kinetics of coinfection with influenza a virus and streptococcus pneumoniae etiology of severe communityacquired pneumonia during the hajj-part of the mers-cov surveillance program clinical and molecular characterization of rhinoviruses a, b, and c in adult patients with pneumonia human rhinoviruses and severe respiratory infections: is it possible to identify at-risk patients early? viral pneumonia and acute respiratory distress syndrome burden of acute respiratory disease of epidemic and pandemic potential in the who eastern mediterranean region: a literature review global threat of animal influenza viruses of zoonotic concern: then and now surviving sepsis campaign: international guidelines for management of severe sepsis and septic shock global and local epidemiology of communityacquired pneumonia: the experience of the capnetz network global changes in the epidemiology of community-acquired pneumonia community-acquired pneumonia requiring hospitalization among u.s adults comprehensive molecular testing for respiratory pathogens in community-acquired pneumonia empirical use of fluoroquinolones improves the survival of critically ill patients with tuberculosis mimicking severe pneumonia impact of antibiotic therapy in severe community-acquired pneumonia: data from the infauci study predictive factors of mortality in severe community-acquired pneumonia: a model with data on the first h of icu admission viral infection in patients with severe pneumonia requiring intensive care unit admission etiology of severe pneumonia in the very elderly assessment of prognosis in patients with community-acquired pneumonia who require mechanical ventilation the authors would like to thank the following: ms. carmen kam, the resident biostatistician for verification of the study statistical analysis; nurses ms. wang xi qin, ms. goh yuan xuan, ms. lim qian ru for assistance in data collection; senior medical technologist ms. heng ying xuan, ms. lee hui zi for assistance in performance of pcr tests; dr. tay tunn ren for her tutelage in manuscript writing. the study was performed with a grant awarded from changi general hospital research grant in (grant reference number chf . ), for an amount of $ singapore dollars (equivalent to usd , conversion usd = sgd . ). the datasets analysed during this current study are available from the corresponding author on reasonable request. authors' contributions jlq contributed to the design, analysis, interpretation of the study; drafting and revision of the manuscript. bj contributed to the design, interpretation and microbiological analysis of the study. pct contributed to the design and data acquisition of the study. cs contributed to the design of the work, drafting and revision of the manuscript. tyt contributed to the conception, analysis and interpretation of the study, drafting and revision of the manuscript. all authors have given final approval of the version to be published and agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.ethics approval and consent to participate ethics approval was obtained from the singhealth centralised institutional review board, singapore. reference number: cirb / /fp. waiver of consent granted for this study. not applicable. the authors declare that they have no competing interests. springer nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. key: cord- -xhx pzhj authors: nan title: nd world congress on pediatric intensive care rotterdam, the netherlands, – june abstracts of oral presentations, posters and nursing programme date: journal: intensive care med doi: . /bf sha: doc_id: cord_uid: xhx pzhj nan we present the results of a prospective population-based audit of paediatric intensive care activity in two comparable communities with markedly different delivery systems. in the trent region of the uk ( . million people), children receive intensive care largely without the supervision of a paediatric intensivist in a variety of hospitals, few of which have designated paediatric intensive care units (picus). critically ill children otherwise receive intensive care in children's wards, special care baby units (scbus) or adult intensive care units. in the australian state of victoria ( . million people), children receive intensive care almost exclusively in one centre -a picu staffed by full time paediatric intensivists. the two regions are otherwise demographically comparable. in both groups, data were collected on all children admitted to an intensive care unit between / / and / / and children who received intensive care (defined by levels of intervention and nurse dependency) in other sites during the same period. values of each variable at first contact with the icu, and the highest and lowest values over the first hours were recorded. the principal outcome was survival to discharge from the intensive care unit. severity of illness was assessed using pim (paediatric index of mortality) and prism. risk-adjusted mortality was compared using flora's z test and logistic regression. the rate of utilisation of intensive care (> admissions in each region) were similar. there was some variation in case mix between the two groups, but crude mortality rates were similar ( . % in trent and . % in victoria). however severity corrected data and other measures of picu performance were dramatically better in' the centralised delivery system. the substantial excess mortality in the trent region provides strong evidence for the benefits of centralisation of paediatric intensive care services. there are considerable difficulties in evaluating the efficiency and effectiveness oflcare in children presenting with respiratory failure during acute medical illness. optimal outcomes for such episodes include survival and the shortest length of stay (los) in intensive care with negligible risk of readmission. we have tried to determine whether or not the time course of acute severe medical illness with respiratory failure is predictable. study i (n= ): a retrospective study of intubated and mechanically ventilated children (> days, < years) with acute severe medical illness. measures: diagnosis, intensive care los in calender days, and survival. results: the underlying diagnosis fell within one of three broad categories: respiratory disease (n= , mortality . %), central nervous system (cns) disease (n= , mortality . %), and systemic inflammation or multisystem (sims) disease (n= , mortality . %. the los in survivors was: respiratory -median (interquartile range) ( - ) days, cns ( - ) days, £p, £ ( -g) days. :i'~'-+cen diag~,~is-rc!ated-grnnp~ (drgs) were identified ( respiratory, cns, sims disease) and each have been characterised by mortality and los. study ii (n= ): a prospective study of patients supported by the hypothesis that los for the above drgs was predictable (compared with study i data). in certain instances attributable causes for variances in los were identified: e.g. disease severity, timing ofdrug therapy, and associated disease. with daily paediatric risk of morality scoring within each drg, four profiles of instability were identified. discussion: the time course of acute severe medical illness with respiratory failure is predictable and variance may be attributable to specific care or diagnostic factors. we are now developing a means of linking drg-specific clinical care pathways with an integrated computerised decision support and education facility at the bedside. the objective of this open, prospective study was to assess the relation between basic patient characteristics as well as effectiveness of treatment on the one hand and resource utilization in pediatric intensive care on the other. as universal, non-monetary indicators of resource utilization we used the therapeutic intervention score system (tiss) and length-ofstay (los), from which indicators for total resource utilization per admission (tisstot) and average daily resource utilization (tiss-mean = tisstot/los) were obtained. overall admissions, totalling days, were included. mortality was . %; non-survivors accounted for . % of overall resource utilization. in non-survivors, both total resource utilization per admission and average daily resource utilization were higher, whereas los was not different from survivors'. severity of illness, surgical status, the presence of substantial chronic comorbidity, emergency admission and transfer from another hospital constituted the major predictive determinants of tisstot (r:= . ) and tissmean (ra= . ) in multiple regression analysis (p< . ). hence these indicators are appropriate non-monetary measures of resource utilization, a considerable proportion of which are determined by a concise set of basic clinical characteristics. subsequently we analysed the relation between effectiveness of care and resource utilization by assessing severity of illness corrected mortality in low, medium and high resource users, respectively. these categories were delineated by percer/tiles of resource utilization (< p , p -ps , > ps ). despite on average long los and high resource utilization in the high risk group, a relatively low standardized mortality was found, probably warranting prolonged intensive treatment in this patient category. summary: objective:the primary purposes of intensive care are to provide treatments to patients with life-threatening physiological dysfunction or to monitor and observe patients perceived to be at significant risk of dying. this collaborative study was performed to describe our patients and their outcome. in order to improve our results we tried to identit~ high risk groups, patients and methods: picus entered the study, the data included all the admissions with > hs. during a days period between the l°june and the th september . the records included: age, sex, weight, mechanical ventilation (mv), post-operative condition (p.op), malnutrition, diagnosis, length of stay, prism score and outcome. student test, mann-whitney or wileoxon were performed for univariate analysis. fisher exact test or chi square for dicotomic variables. risk group analysis was performed by logistic regression, odds ratio and % confidence interval. results: patients entered the study. mean age was . months (ds hh¢# ) and median months. we found significant statistical differences in calculated ,is observed mortality rate comparing malnourished with euthrofic patients; mechanical ventilated (mv) with non mv patients. no differences in ter ~,h of stay or di~ noses were found. effect of the un sanctions on the morbidity rate araong the iraqi small children ( below years old of age ) in bagdad. abdulsamad a.abood / institute of medical technology, bagdad. meningitis is essentially a childhood disease (i). the risk of infection are increased by powerty and overcrowding ( ). the impah'ed immunity may be an important pathogenic factor underlying the susceptibility to infections in undernourished subjects ( ). in general, malnutrition is a man made disease and it begins quite in the womb and ends in the grave (i). small children, below years of age were admitted to the pediatric hospital in washash with meningitis over cold months in i , in contrast to only child admitted with meningitis over the same period in . all of the children who admitted in were frankly undernourished, % of them were infected with enterobacteriae, because they were exposed to faulty hygiene and lack of asepsis. these facts showed precisely that our small children had suffered at most from the un_ sanctions against iraq, because of food, milk and drug shortage, since years which had resulted a severe undernutrition among them, which impaired their immune status. m wells, of riera-fanego, j lipman. baragwanath intensive care unit, university of the witwatersrand, south africa. background the use of prism or other scoring systems in the icu is of great importance for evaluating the efficacy and efficiency of a particular icu, the prism score was developed and validated in the usa and europe but has recently been shown to be inaccurate in a south american population, a south african population as well as several european studies. part of the poor performance of the prism score is as a result of differences in the case mix between the reference population and other paediatric icus. since scoring systems should generally be used only in populations similar to the reference population from which the prediction model was developed, a modification of the prism score is necessary to improve its discriminatory ability in a wide range of patient groups, aim to improve the predictive power of the prism score in a south african paediatdc icu population. patients & methods we analysed prism, demographic and clinical data collected prospectively from consecutive paediatrie icu admissions. the prediction of actual mortality by prism was evaluated by standard statistical methodology (goodness-of-fit test and receiver operating characteristic (roc) analysis), the components of the prism logistic regression equation (prism score, operative status and age) and the physiological variables making up the prism score in addition new variables analysed (nutritional index, the need for inotropes and institution of mechanical ventilation) were subjected to discriminant analysis to determine their association with outcome. results the goodness-of-fit test showed a significant failure of prism to accurately predict mortality over a wide range of expected mortality (chi [ ] = , p = ). prism underpredicted mortality at lower prism scores, but overpredicted mortality in patients with high prisms. similarly roc annysis indicated apoor predic~jve power (az = . ± . ), with an area under the curve significantly less than that for the prism reference population (p = ), prism showed equally poor discriminatory function at all age groups and diagnosfic categories. '~mth the addition of an index of nutrifional status (proportional weight-far-age), and indicators of early respiratory and cardiovascular failure to the logistic regression formula, and a recalibration of the acute physiological score component, the roc can be improved to . ± . , with a good fit described by the goodness-of-fit test (cn ] = , p = . ). discussion the prism score is not accurate in our patient population has been recalibrated in view of the poor discriminatory function that we have shown. part of the inaccuracy derives from the different demographic characteristics of our icu population and a different pattern of diseases. in addition to assessments of acute physiological aberrations, an assessment of nutritional status and early respiratory and cardiovascular failure significantly improve the discriminatory ability of the prism score, these parameters have been devised with a view to improving the accuracy of prism in our population, while not decreasing its accuracy in icus similar to the reference population. in interviewing parents regarding how physicians have communicated bad news, the response i have received is that it has not infrequently been done without appropriate care, understanding and compassion. personal experience and the lessons learned from parents, chaplains and others who deal extensively with these situations have provided me with an approach that has been supportive, compassionate, and caring. an especially difficult communication situation for the intensivist occurs when the parents have to be informed of the death of their child. for the parent, death is the hardest loss of all -the ultimate unalterable loss. circumstances surrounding the death are an important consideration (e.g., a fatal crash caused by a drunken driver, a prolonged illness, a suicide, aids). each produces a different grief reaction. the physician needs to inform parents of their child's death sympathetically coming right out with the news and leaving details until later. allow pauses and time for the paren~ to express sorrow and grief, the best communication may be thoughtful silence and a tender touch. there is disbelief that this happened. it is necessary to repeat oneself. acknowledgment of the parent's "feeling terrible" and the physician's acknowledgment of how terrible he/she feels that the life of the child could not be saved is an important first step in the parent's dealing with this tragic loss. with prolonged resuscitation, it is helpful to have a member of the icu team talk to the parents while the resuscitative efforts are ongoing so that the parents are not left unsupported at this time. a progress report should be delivered in a caring, lucid, and sensitive.manner, indicating that every effort is being made to save the life of their desperately injured child. after a child has died, it is helpful to the family if the physician maintains some contact with them. this should take the form of follow-up telephone calls at approximately , , and months. this can help to screen for depression in the parents. in giving bad news to the family and making every effort to support them through this tragic time, it is necessary to remind oneself that the intensivist has personal needs for dealing with grief and will also require support to pass through this stage. direct evidence that child mortality is lower in specialist pediatric icus comes from studies. a study in oregon (ccm ; : - ) found that mortality adjusted for severity of illness was % of expected in pediatric units and % of expected in general units (p< . ). a study in holland (ccm ; : - ) found that mortality in high risk patients was % of expected in tertiary pediatric units, and % of expected in nontertiary units (p< . ). a third unpublished study, has found that children in victoria (who almost all receive intensive care in a pediatric icu) have a much lower standardised mortality rate than children in the trent region of the uk (where many children receive intensive care in adult icus). there is indirect evidence that icus looking after many children are likely, on average, to perform better than icus looking after few children: numerous studies in many specialities have found that units looking after many cases of a particular disease have better results than units with few cases. see luft hs, "hospital volume, physician volume, and patient outcomes", happ, ; and farley d, medical care ; : - . compared to general icus, medical and nursing staff in pediatric icus are likely to be better at looking ~fter children, and plcu rmos have greater skills in pediatric intubation, ventilation, iv drip insertion and drug doses. picus are more likely to have appropriate equipment to manage children -especially for uncommon but life-threatening situations. icus in pediatric hospitals are more likely to have physicians and surgeons with pediatric expertise available for consultation at all times. the american academy of pediatrics, the society of critical care medicine, the british paediatric association and the australian nh&mrc have all said that children should receive intensive care in'specialist pediatric units. the weight of authoritative opinion, and direct and indirect evidence is strongly in favour of looking after children in dedicated pediatric icus. neurological deficit showed higher cbf values ( . / . ml/ g/ rain.) than the patients with good outcome (mean cbf . sd + . ; cbf . sd _+ . ml/ g/rain}. discussion: in asphyxia decrease of ph is due to reduced tissue oxygenation and indicates the severity of metabolic derangements. co reactivity in newborns with perinatal asphyxia correlates with the lowest ph and therefore may reflect severity of asphyxia. continuous monitoring of cerebral activity is carried out in our unit on all admissions at risk of cerebral dysfunction, a number of monitors are commercially available and we report our experience with the cfam which provides in addition to amplitude integrated eeg analysis, continuous raw eeg display and frequency distribution. bilateral recordings are commenced as soon as possible and continued while clinically indicated. forty one children ranging in ages from weeks to years were monitored for periods from hours to i days, diagnoses included traumatic brain injury ( ), sepsis/meningitis/encephalitis ( t), status epilepticus ( ) and miscellanous others ( ). results are tabulated below. patients status epilepticus * beta activity * background voltage * < i o/zv or more of above * (*z p < , ) asymmetry developed in children, all of whom died. positive predictors of good outcome included a mean background activity of > zzv, the presence of faster frequencies (usually ) in response to sedative drugs and the absence of seizures. all monitoring is performed by the picu staff and increasing expertise in interpretation has resulted in earlier therapeutic and diagnostic interventions. regional it was previously found that histamine, a vasoactive mediator, accumulated in brain compartments (kov~ics et al neurosci lett : ) , and antihistamines prevented brain edema formation (dux et al. neuroscience : ) in asphyxiated newborn pigs. in the present study we investigated the effect of intracarotid histamine injection on the blood-brain barrier (bbb) permeability, left internal carotid artery of newborn pigs ( - h; , - , g; ketamine anesthesia, mg x kg ) was catheterized through the external branch and different doses of histamine ( , - , xi - , - , x , m, respectively, in groups of animals; n= in each) diluted in . ml isotonic saline was injected into the vessel through rain. bbb permeability was determined for a small (sodium fluorescein, sf, da) and a large (evans blue/albumin, eba, kda) tracer ( %, mlxkg , rain circulation time for both dyes) concomitantly in frontal, parietal and occipital cortex, hippocampus, and periventrieular white matter both on left and right sides h after the challenge. then, intravascular dyes were removed by perfusion and bbb permeability for both tracers was quantified by fluorescence spectrophotometry (wavelengths for excitation and emission were nm and nm for sf; and nm and nm for eba, respectively). histamine injection, in doses higher than . m, significantly (p< . ; kruskal-wallis one way anova on ranks followed by dunn's test) increased bbb permeability for both tracers in each brain region. changes in left hemisphere were more intense (p< . ) than those in right one after the doses of xi - and - m in each region, i m histamine administration induced similar edema in both sides. increased intracarotid histamine levels resulted in a dose-dependent vasogenic brain edema formation. histamine might have a pathogenetic role in neonatal hypoxicischemic cerebral injuries. supported by otka f- and h-u.s,-jfno. , $ in coma caused by traumatic brain jnjury, an indication of the likely outcome is provided by the best motor response to pain in the first .$ hours after the insult. in a study in our picu, the proportion of children who died or had a severe disability was % in who had no response to pain, % in with an extensor response, % in with a flexor response, and % in who localized in response to pain. the long term outcome of traumatic brain injury appears to be worse in children < years old. other risk factors in traumatic brain injury are absent basal cisterns, midline shift or subdural haemorrhage on ct scan (or loss of grey-white differentiation in nontraumatic injury); or an intracranial pressure > mmhg despite hyperventilation, mannitol and barbiturate infusion. apart from brain death, there are two findings implying such a poor prognosis that consideration should be given to stopping treatment: first, after traumatic injury, the absence of any motor response to painful stimulus in the cranial nerve distribution (providing drug effects and a post-ictal state have been excluded); and second, in acute brain injury from trauma, infection, hypoxia, or ischaemia, the b{lateral absence of short-latency somatosensory evoked potentials (providing brain stem haemorrhage, subdural and extradural effusions, and decompressive craniectomy have been excluded). in children over months of age, recovery from prolonged coma or a vegetative state is exceedingly rare when more than months have elapsed after traumatic brain injury, and when more than months have elapsed after nontraumatic injury. overproduction of nitric oxide (no) via an inducible isoform of" no synthasc (inos) produces profound vasodilatation in adult septic shock. high nitrate levels have been reported in hypotensive children with sepsis syndrome ]. cardiovascular collapse is a prominent feature of severe meningocoecai disease (mcd). however, systemic vascular resistance (svr) was slightly higher in a group of non-survivors ~ and the rote of no in ivicd remains unclear. children with a presumptive diagnosis of mcd were enrolled. parental consent was obtained. blood was drawn on admission and hrly thereafter. plasma was separated immediately and stored at - °c. the final concentrations reported represent the product of nitrite and nitrate (nox). nox was measured spectrophotometrically using the greiss reaction. children were studied (median age (range); m ( - )). the diagnosis of mcd was confirmed in children, of whom had a glasgow meningococcal score (gms) of" ~ . in this group with severe mcd there were deaths. peak nox was significantly higher (,. ( - ) vs ( - )nmol/ml, median) and systolic btood pressure was significantly lower in children with severe mcd than mild mcd (p< . . wilcoxon rank test). there was a significant correlation between peak nox and gms (spearman's rank correlation r= . (p= . )) and prism (r= . (p: . )). nox production from adm.ission onwards was also higher in the severe mcd group (p: . , kmskal ~wallis). we have demonstrated that plasma nox levels are elevated in children with mcd, correlate directly with the severit ' of disease and are inversly related to systolic blood presssure. similar to hypotensive septic syndrome, mcd appears to be associated with an up-regulation of the l-arginine-no pathway.. non-survivors with mcd have higher svrs and may be relatively hypovolaemic. in our group of severe mcd there was a significantly lower systolic pressure and increased no formation. excess inos expression at different stages in mcd may contribute to the pathology of the disease. the identification of agents which can boost and/or inhibit no reiease may therefore represent different treatment strategies for mcd. u. merz, th. peschgens, g. kusenbach, m. b hle, h. h rnchen in this controlled, prospective study ventilated premature infants with a birth weight < g were randomized to receive treatment with dexamethasone (dex) either on day of life or on day of life. dex was given over days tapering from . mg/kg/day to . mg/kg/day. the infants treated with dex on day of life could be weaned earlier from the ventilator -in median after days (range - ) versus days (range - ) in the [ate treatment group (p = . ). the need for supplemental oxygen was shorter in the early treatment group -in median days (range - ) versus days (range - ) (p = . , ns). the incidence of chronic lung disease was lower in the early treatment group - of infants ( . %) versus of patients ( . %) (ns). to evaluate the long-term efficacy of early dex treatment we performed a respiratory function test in the age of - months using an infant whole body-plethysmograph. the intrathoracic gas volume (itgv), the airway resistance (r.w) and the airway conductance (gaw) were measured and no significant differences could be detected between the groups. the frequency of adverse effects due to dex therapy was found to be without significant differences between the early and the late treatment group. we conclude that early dex treatment had short-term improvements in pulmonary outcome in our study population, long-term efficacy however, remained unproven. several factors contribute to the development of chronic lung disease (cld) in premature infants including structural immaturity of the lung, mechanical ventilation, and oxidative stress. reactive oxygen species are formed during normal cellular metabolism but they are generated in higher concentrations during inflammation or inhalation of high oxygen concentrations. to study the relationship between increased oxidative stress, antioxidants and the development of cld we examined ventilated premature infants with birth weights below t g. infants developed severe chronic lung disease of prematurity (cld), defined by radiological signs of cld and an increased oxygen requirement at a postconceptional age of weeks, and infants had moderate cld with an increased oxygen requirement on day but not at an age of weeks. ventilator settings (fio , peak inspiratory and mean airway pressure) and the incidence of early-onset-sepsis were significantly higher in the severe cld group than in infants with moderate cld or without cld (n= ) during the first week of life. plasma concentrations of the two antioxidative substances bilirubin and uric acid (ua) were comparable in all groups during the first days of life. however, on day seven bilirubin and ua were significantly decreased in the plasma of infants with severe and moderate cld compared to the non cld group (p<o. ). there was no difference between both cld groups. in serially obtained tracheal aspirate fluids (taf) malondialdehyde (mda) was measured by hplc as an indicator of lipid peroxidation reflecting oxygen injury of the lung. ua was found in higher concentrations in taf than bilirubin. ua has been proven to be a potent antioxidant, capable to react especially with hydroxyl radicals and hydrochlerous acid, which play an important role in the induction of lipid peroxidation. infants with severe cld had significantly higher mdnua ratios in taf from day to compared to the non-cld group (p<o. ). mdnua ratios in taf of infants with moderate cld were significantly higher from day on but still lower than in the severe cld group. in addition, the ratio oxidized / reduced glutathione was significantly increased in taf of infants with cld compared to infants without cld between day and . the data support the hypothesis that an imbalance of oxidative stress and antioxidant defense contributs to the onset of cld at the end of the first week of life. introduction. rocuronium bromide (roe), a neuromuscular blocking agent with an onset time dose to that of succinyleholine l'z, has been studied in american society of anesthesiologists' physical status (asa) i or ii pedlatrie patients under non-critical elective conditions. there is a paucity of studies looking at the pharmacodynamics of roc under emergent conditions, involving pediatric patients who are asa iii or iv, without the use of adjuvant inhaled anesthetics ~. methods. with approval of our committee on clinical investigations (irb), roe . mg/kg was administered to asa iii/iv pediatric intensive care patients (age days to months; m:f= : ) on intravenous sedation. the time to reach % a~d % of b~elinc tetanie stimulus response was recorded using a r.elaxogra'ph/qmt- nerve stimulator/recorder (datex, helsiaki). depth of clinical relaxation at the % level was recorded. results. the time from rapid bolus to % was . + . see with a range of - see; and to % was . + . see, with a range of - see. at the % level, excellent clinical relaxation was seen. conduslon. in critically ill children, use of roe at . mg/kg produces rapid onset of neuromuscular blockade and excellent clinical relaxation, such that roe should be considered for rapid sequence intubation. int anesth clin. ; ( ): - . anesthesiology. ; ( ) computed tomography of the chest (ct) in mechanically ventilated adults can detect intrathoracic pathology not appreciated on chest radiographs (cxr) and influence intensive care management. no such data exists for ventilated children. aims: ) to assess if ct provides additional information over cxr regarding extent and nature of intrathoracic disease in ventilated children. ) to determine whether such information alters clinical management, inclusion criteria : ventilated children with cxr changes inconsistent with their respiratory status underwent ct if either a) pao :fio ratio < and/or mean paw> cm h or b) there was an unexplained increase in ventilatory requirement. methods : high resolution ct was performed in patients and spiral ct in patierits, to ensure minimal transport related morbidity, patients were transferred to the ct scanner by a specialised mobile intensive care team. results: in / patients ct demonstrated greater extent of disease than appreciated on cxr but did not significantly alter clinical management. in / patients ct provided additional information regarding the nature of disease present, in / children this involved a further diagnosis and in / children the exclusion of a suspected pathology. new information led to a positive therapeutic intervention in children, prevented inappropriate manoeuvres in , and had no significant effect on acute management in children. conclusions: initial data suggests that in a selected group of mechanically ventilated children chest ct can add to the sensitivity and specificity of intrathoracic diagnosis provided by the chest radiograph and directly influence acute management. case selection criteria and choice of the most appropriate protocol requires further study. pressure control ventilation (pcv) utilizes a decelerating flow pattern which may improve gas distribution and lead to alveolar recruitment. in contrast, volume control ventilation (vcv) employs a constant flow. in children, the effects of pcv as compared to vcv are unclear. the purpose of this study was to determine how these two modes compare in terms of dynamic compliance (cdyn). peak iaspiratory pressure (pip), and mean airway pressure (paw) at equivalent minute ventilation. methods: sixteen infants and pediatric patients ranging in age from day to years were studied. diagnoses included ards ( ), postoperative cardiac surgery ( ), head trauma ( ), and resfrictive lung disease ( ). patients were randomized to pcv ( ) or vcv ( ). initial measurements of gas exchange (abg's) and respiratory mechanics (ventrak, novametrix medical systems) were obtained after a minute stabilizadon period. respiratory mechanics included pip, peep, paw, delivered tidal volume, and cdyn (avolume/apressure). the patients were then crossed over to the alternate mode of ventilation holding delivered tidal volume, peep, inspiratory time, minute ventilation, and fio constant. data were collected after minutes, in each mode the absence of intrinsic peep was confirmed. to assure that the measurements were not affected by changes in clinical status, the patients were returned to the initial mode of ventilation and measurements repeated (final) . patients were ventilated with a siemens c or sv . reselts: data were analyzed using -way analysis of variance with repeated measures. ~ < . vs. vcv) vcv pcv ~ initial ] final ! cdljn . _+ . . _+ . * . _+ . . _+ . i , pip + . l-_t. * _+ , +- , paw . _+ . . i-_ . * . + . . -!-_ . pao _+ +- _+ _+ discussion: at the same minute ventilation, the decelerating flow pattern of pcv resulted in a % increase in cdyn and an % increase in paw while decreasing pip by %. the lack of a significant change in oxygenation may be a result of the limited time in each ventilator mode as well as the inclusion of patients with both normal and abnormal lungs. there was no significant difference in initial and final measurements indicating patient stability. the beneficial effects of iecre~l~iug cdyn and paw while decreasing pip indicate that pcv may be a preferable mode of ventilation in patients with lung injury. further randomized studies examining the effect of pcv on respiratory outcome measures in pediatrics are indicated. prolonged positive pressure ventilation following repair of cdh is associated with a high prevalence of iatrogenic lung injury, in our unit dudng - late deaths after repair of cdh were due to chronic lung disease. since babies requiring assisted ventilation for more than days following surgery were transferred to a cnep chamber to limit lung injury. cnep of - cm of h was combined with positive pressure ventilation via an endotracheal tube dudng the transition phase. immediate reduction of peak inspiratory and positive end pressures were possible and following extubation respiratory support was maintained by cnep v~th appropriate inspired oxygen. overall outcome: [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] n= deaths before surgery (%) ( ecmo during - / who were ventilated for more than days received cnep and there were no deaths and no chronic lung disease in that group. cnep assisted ventilation may be an important management option for babies who require prolonged respiratory support to avoid the adverse effects of chronic positive pressure ventilation, introduction so far modes of liquid ventilation (lv) have been used in experimental animals and, exceptionally, in humans: . total liquid ventilation (tlv)-functional residual capacity (frc) is filled by perfluorocarbons (pfc), and slow tidal volume (tv) breathing is performed by pfc. . partial liquid ve, ti,la~ion (page) -only frc is filled by pfc. gas tv is delivered by conventional mechanical ventilation (cmv), high frequency jet ventilation (hfjv) or high frequency oscillation (hfo). the aim of our study is to present our limited experience with page in newborns and infants. page was used in two groups of infants: , in infants with brain death before disconnection from cmv, because recipients for organ transplantation were not available. these infants have relatively normal lungs (fio~ less than . ). infants stayed on page for hour, during that period no ventdator manipulations were made. after page, infant were switched to cmv for next hours. . very critically diseased infants with ards (rds) - on ecmo more than days, before cannulation for ecmo, on hfo because of intractable respiratory failure, preoxygenated rm (miteni, italy) was used in the doses up to ml/kg intratrachealy. blood gases and parameters of pulmonary mechanics were followed (dynamic compliance -c dyn, airway resistance -raw, bicore monitor). page was combined with no inhalation ( - p.p.m, in infants). in both groups ad hoc an approvement from e local ethical commission and informed parental consent were obtained. in the first qroud with relatively normal lung parameters of oxygenation drops after pfc instilation intratracheally and stayed depressed for - hours. slight pco retention occured in both cases during page. c dyn increased almost double during page period, raw drops transitorily after pfc instilation but in minutes they were identical like in prepage period, parameters of oxygenation (peo /fio ) after - hours after page improved and were better than in prepage period. after that time infants were disconnected and died. in the second group no improvement of oxygenation was seen in one ecmo baby, in spite ()f transient improvement of c dyn. in the second ecmo baby, oxygenation improved and flow of pump could be decreased by more than %. none of these babies, however, survived, improvement was only transient in spite of repeated dosis of pfc. in these babies serious problems were to maintain the adequate frc by liquid, because of severe air leak, in babies on hfo/hfjv with severe ards/rds the improvement of oxygenation were seen in all the cases immediately after pfc instiletion for the period of - hours. after that period, pfc dose had to be repeated. two babies of this group survived. conclusion. page is going steadily from tabs to clinical practice. it is simple, could be performed anywhere, cheaper than tlv. however, because liquivent -perflubren (aliance pharmaceutical) is not available in europe, rm of (mitenti, italy) is the only solution, which could be currently used here. before the widespread use of page in clinics, liquid network among most nicus and picus must be built up, the criteria for page must be defined and ethinal-legal problems resolved as well. after resolution of these particular problems page can be life saving procedure for very special part of critically ill newborns end infants. catherine caronia, peter silver, laura nimkoff, cad quinn, jack gorvoy, and mayer san. division of pediartic critical care, medici,, schneider children's hospital, new hyde park, ny , imroduetiun: cystic fibrosis (cf) patients awaiting lung transplantation present a therapeutic dilenuna when severe respir, aory decompemalion occurs, endotracheal intubation and mechanical ventilation is known to have no long term benefits and is associated with high morbidity and mortality. noninvasive respiratory support appears to be a beneficial alternative. methods: we instituted bipap (respironics, inc,, murrayville, pa) in end-stage cf patients who were admitted to the pediatric icu with severe respiratory decompeusation. all patients were awaiting tung transplantation. after a control period, bipap was applied via a tight fitting nasal or facial mask, using the spo~aneous breathing mode, expiratory pressures were set at - cm hhzo. inspiratory pressures were started at cm ~i o and increased in cm i-i increments until the patient's respiratory comfort was achieved and substantiated by non-invasive monitoring. patients were instructed to use bipap during night sleep and whenever subjectively required, data are reported as mean _+ s.d. results: all patiems utilized nocturnal bipap for - hours/day during a follow-up period of - months. compared to their pre-bipap status, the patiems' oxygen requirement and respiratory rate both oz~ cundusion: bipap tl~rapy improves the respiratory status of decompeusatir!g end-stage cf paacnts. it is well tolerated for long term use at home, and provides an extended period of respiratory comfort and stability for cf patients awaiting lung transplantation. l. bindl*, g. kiihl**, p. lasch***, appel**, j.m er**** and the "arbeitsgemeinschaft ards im kindesalter" background acute respiratory distress syndrome (ards) is a therapeutic challenge in pediatric intensive care in view of the high mortality, in about german paediatlic hospitals founded a working group aiming on collaborative clinical research in this field. aims and methods the aim of both a prospective and retrospective survey conducted in german pediatric intensive care units in was to accumulate data on the epidemiology, risk factors, natural history and treatment strategies in a large group of pediatric ards patients who were treated in the tt~ee year period from to .all patients had acute bilateral alveolar infiltration of noncardiogenic origin and a po ~io ratio < mmhg. the influence of sex, underlying disease and single organ failure was analyzed using the fischer's exact test, the influence of additional organ failure on mortality was tested with the cochran-mantel-haenszet statistics. results patients were reported giving an incidence of cases per admissions to pediatric icus. median age was month. in % of the cases, ards was associated with a pulmonary, in % with a systemic underlying disease. in % immunocompetence was impaired. mortality was % and not dependent on age, sex and triggering event. the number of associated organ failures, however, strongly influenced mortalib,. mortafity in immuno-compromised patients was t %. the analysis of treatment modalifies employed in the patients revealed a lack of uniform therapeutic strategies. on the other hand, the patients were exposed to interventions not yet supported by controlled trials. conclusions the observation of the lack of uniform treatment strategies led to the elaboration of recommendations on ventilator therapy and patient monitoring within the working group. the data gathered in this survey provide the basis for the design of prospective multicenter studies urgently needed to evaluate innovative treatment modafities in pediatric ards. recurrent apnea and respiratory failnre due to severe lower respiratory tract disorders such as bronchiolitis or pneumonia are the most common reasons for mechanical ventilation during respiratory syncytial virus (rsv) infection. acute respiratory distress syndrome (ards) has been described as a complication of severe rsv infectionj in contrast to the low mortality rates associated with rsv infection (< %), mortality rates in the range of - % have been reported in pediatric patients with ards. however, studies on ards are usually lumped in respect to causation and the disease course of rsv induced ards has not been previously studied. we examined the lung function abnormalities of infants with rsv induced respiratory failure requiring assisted ventilation, measurements included respiratory mechanics, maximal expiratory flow-volume curves and lung volumes, ards was defined clinically using the criteria which were recently proposed by the american-european consensus conference on ards~: acute disease onset, pao /fio~ ratio _< mrn hg, bilateral infiltrates on chest radiograph and absence of clinical evidence of left atrial hypertension. we calculated the murray lung injury scores modified for use in pediatric patients from total respiratory system compliance, radiographic findings, ventilator settings and blood gas results. we identified infants with severe restrictive lung disease that fialfilled the clinical criteria fbr classification as ards. all had lung injury scores above . which is the recommended cut-off for a diagnosis of ards, twenty-seven infants had obstructive disease consistent with a clinical diagnosis of bronchiolitis. the ards patients were significantly younger, had a longer time of assisted ventilation (p < . ) and a greater proportion of infants with preexisting illnesses (p= . , odds ratio = . ) when compared to the patients with obstructive disease. with the exception of one immunodeficient patient, none of these infants died. given the low mortality despite a clinical picture of severe lung injury, there is evidence that rsv induced respiratory failure may represent a relatively benign cause of ards in pediatric patients, bachmann an audit of patients with severe acute bypoxic respiratory failure (ahrf) receiving highfrequency oscillatory ventilation (hfov) in our unit ( n= , mortality %) revealed that sub-groups with severe underlying disease (n= , mortality %)and those with mu~pie organ failure ( > systems failing, n= mortality %) accounted for all the deaths beyond the neonatal period. v~ therefore hypothesized that in a modem paedistric intensive care unit (picu): a) children greater than one month of age with ahrf do not die in the absence of severe, pre-existing disease or multi-organ dysfunction syndrome, b) respiratory parameters alone will predict outcome poorly in ahrf. method prospect~/e sty/of all adm~ns to our tertiary picu. data it, citing the respiratory parameters (oxygena~n index [ol] , aiveolar-artedal oxygen tension gradient , pao /fio ratio) were collected hourly from the bedside charts throughout admission. patients were included in the study if ahrf was present at admission either none or in combination with other organ dysfun~on. ahrf was defined as the acute (< hour) onset of respiratory dysfunctk:~l with a pao /fio ratio.< for six consecutive hours dunng the first hours of admission (with no evidence of left anal hypertension), x-ray review defined a sub-group of patients with acute respiratory distress syndrome (ards) by the presence of bilateral interstitial infiltrates. results to date children (ages - months, weight . - kg) have been admitted in ahrf. of these also had ards. the overall mortality was . % ( / ), and greater in the ards group than the non-ards group ( t , . % vs, , . %, p< o. ) . it was not possible to predict survivors from non-survivors on the basis of the seventy of the respiratory failure alone, the a-ado on the day of admission (best in hours) was not significantly different between survivors and non-survivors: (mean, + sd)( mmhg +_ , vs mmhg _+_ ). kdl non-survivors were immunodeficient (n= ), previously extmrnsly premature infants (< ),(n= ) or suffedng fcom chronic metabolic or gastrointestinal disease (n= ). no previously normal child died. conclusion the severity of respiratory failure does not allow predioljon of outcome in our patients. we believe that this reflects that modem picu is so effective at providing respiratory support that pre-existing pathology alone de~ prognosis. this suggests that an abnormally regulated host response or abnormal persistence of a pathogen may be required to induce lung injury of sufficient severity that the resulting respiratory failure cannot be supported in a modem picu. introduction: postural changes (supine to prone) is a therapeutic intervention that could be useful in children with adult respiratory distress syndrome. objective: to determine the effects of postural changes in the oxygenation of young children with ards. method,s: a prospective stud ," was performed in eleven subjects aged to months (mean= ) with the diagnosis of ardsreceiving vendlatory support. (mean peep and fio of and . respectively). postural changes was performed every - hours, during a period of time ranging from to days. arterial blood gases were determined before and - n~n after the postural change, no modification in the mechattical ventilation other that changes in the fio were performed. the oxygenation was determined by the index pao /fi (p/f). to study the differences between the oxygenation mean, before and after the postural changes the wilcoxon test for paired samples was used, results: changes were performed ( from supine to prone and from prone to supine). a % increased p/f ratio was obtained after the change from supine to prune. although, not all the patients receiving postural changes improved their p/f. six of them (group i) showed an improve in the p/f when changed from supine to prone, returning to their base line when positioned from prone to supine. no improvement on the p/f was observed in the remaining subjects (group ii)after postural changes (table ) . during the maneuver no complications were observed. two patients had a pneumothorax, not related with the postural change. conclusions: postural changes (supine to prone) is an easy way to improve oxygenation in some children with ards. change to prone change to supine introduction: the common noninvasive diagnostic efforts to identify possible obstruction of the intrathorucic airway, are of limited value. invasive procedures such as bronchoscopy and bronchography may also be noncontributory and entail risks. we evaluated the usefulness of d-ct in the diagnosis and management of pediatric patients with suspected intrathoracic airway obstruction (itao). methods: we used a diagnostic algorithm (see diagram) in patients with suspected itao resulting in respiratory distress. three-dimensioual imaging of the tracheobronchial tree was reconstructed, following high speed spiral ct scan, by specific computer software (advantage window computer work station, general electric, milwaukee, wisconsin). non-ionic contrast medium was injected, in some patients, to delineate the intrathoracie large vessels.. results: eight patients were studied. in patients the d-ct revealed intrathoracic airway abnormalities. these patients underwent further invesive studies which confirmed the following diagnoses: patients had bronchomalacia, had bronchial stennsis due to a dilated pulmonary artery mad patients had subglottie stenosis extending to the thoracic cavity. three patients had no significant disruption in the configuration of the tracheobronchial tree and thus did not require invasive diagnostic procedures. conclusion: computer reconstruction of three dimensional images of the tracheobronehial tree is a safe and reliable diagnostic tool for itao. ards and ecmo; preliminary data from a randomized clinical trial. j fackler, c steinhart, d nichols, d bohn, m heulitt, t green, l martin, k newth, m klein, j ware. many suggest ecmo be considered experimental for ards and undertaken only with careful data collection and reporting. a mtflticenter pediatric rct is in progress to determine whether ) ecmo and/or ) permissive hypercapnia, offer significant advantage for the treatment of ards. methods: all patients aged wk to yr (without congenital heart disease) are eligible for study. data collection begins when a patient receives at least % oxygen and a peep of cm h for hours (stage t). if the predicted mortality reaches % within days (stage ), eligible patients are asked for written consent for randomization. patients are excluded from randomization with significant chronic lung disease, immune compromise, cardiac disease; or profound acute central nervous system damage. the prime outcome variable is survival. at the studies onset, pts were estimated to be required so that pts were randomized per arm. results: patients are enrolled from centers. data are complete on . patients never reached stage (i.e. % mortality). patients improved and died. of the latter, had randomization exclusion criteria even if stage was reached. patients reached stage . had exclusions from randomization and all died. eight patients ( survivors were eligible for randomization; consent was obtained in no case. two patients received ecmo. overall survival is % ( / ). in patients without randomization exclusions, survival is % ( / ). morbidity m survivors (discharge -admission popc or pcpc score >_ ) was seen in none of the stage surviviors and % ( / ) of those who reached only stage !. conclusion: the rct requires completion. the records of hospital in-patients at king faisal specialist hospital and research center who received external cardiac massage as part of their cardiopulmonary resuscitation were reviewed. success of resuscitation was analyzed as ( ) short term (restoration of spontaneous circulation), and ( ) long term (discharge from hospital). of such patients, ( . %) survived the initial resuscitation, and ( . %) were discharged. success of outcome was not related to age, location of patient, time of day, or rhythm at arrest, including asystole. longer resuscitation time was associated with less chance of restoration of spontaneous circulation (p< . ), but not associated with hospital discharge rate. results for patients with congenital heart disease were similar to those with other medical or surgical conditions. in this series, . % of ward in-patients survived to discharge, compared to two "*;'~r ~r;~' ,.,.'her,, the r-e~ult~ were c/ "'~d ~, ~,°(. overall, % of patients who survived the initial resuscitation were discharged from hospital. where resuscitation continued for more than minutes, . % of patients had tong term survival. outcome from asystole was no worse than for other cardiac rhythms, we believe that previous reports of poor outcome from asystole in pediatric cardiac arrest should noi influence decisions to stop resuscitation for pediatric in-patients prematurely. successful restoration of spontaneous circulation with long term survival can be achieved after prolonged resuscitation. abdelmoniem~ lindsey jahusou~,mariano fiallos, university of florida, prudential drive, suite jacksonville, florida usa central acidosis is well recognized as a marker of inadequate tissue perfusiou, and ventilation. however, obtaining central venous blcod is difficult and fraught with complications in the child undergoing cardiopuimonary resuscitation. intraosseous blood may be used instead of central venous blood to judge ph and pcoz during short durations of cardiopulmonary resuscitation and during hemorrhagic shock. the purpose of this study is to compare the ph and pcoz status of intraosseous and central venous during prolonged cardiopulmonary resuscitation after fluid and drug infusion. we hypotbesized that there would be no difference in ph and pco values of simultanecusly obtained intraosseous and central venous blood samples. eighteen ( ) introduction: cardiopulmonary arrest (cpa) in children is usually preceded by a deterioration of cardiac or respiratory function due to sepsis, dehydration and hypovolemia. early recognition of clinical and laboratory signs followed by immediate intervention are essential for prevention of cpa. the purpose of the present study was to identify factors which contributed to high rates of mortality from cpa in patients admitted to a paediatric intensive care unit (p cu). methods: a prospective study was done of all non-surgical patients with cpa who were admitted to the picu, hospital baca ortiz, quito ecuador from january to october . clinical and laboratory variables before and after admission to the picu, time from hospital admission to picu admission and the pediatric risk of mortality score (prism) were recorded on a questionnaire designed specifically for this study. results: of the non-surgical patients admitted to the picu, ( %) were admitted after developing cpa on the general pediatric wards. mean age was + . months, with of patients under months of age. initial diagnoses upon picu admission included meningitis (n= ), respiratory failure (n= ), congenital heart disease (n= ), severe neurological impairment (n= ), end stage neoplastic disease (n= ), hypovolaemic shock (n=l), peritonitis (n=l) and sepsis (n=l). mean time from hospital admission to p cu admission was _+ . hours. the mean prism score upon hospital admission was + . (score > = > % mortality). % ( / ) of the patients died. one of the three survivors had severe neurologie injury. prior to picu admission, patients experienced tac~,cardia (n= ), hypotension (n= ), neurological deterioration (n= ), respiratory, distress (n= ), oliguria (n= ), bradycardia (n= ), metabolic acidosis (n= ), hyponatremia (n= ), hypokalemia (n= ), hypocalcemia (n= ) and severe hypoglycemia (n= ). there were serious delays from the time of development of clinical and laboratory abnormalities to the time of admission to picu. conclusion: in the critically ill pediatric patient, rapid recognition of clinical and laboratory signs of deterioration, followed by immediate intervention, are required to prevent end stage shock and cpa. we found serious delays in intervention following development of important premonitory clinical and laboratory abnormalities in patients less than months of age on the general pediatric wards, which iikely contributed to the dismal % mortality rate. hospitals throughout ecuador should institute immediate improvements in ctinical supervision, and provide training in paediatric advanced life support (pals) to decrease excessively high rates of and mortality from cpa. intraosscous access is recommended by the american heart association and american academy of pediatries as a means of rapid access to the vascular system for childhood emergencies. bone marrow and fat embolism is a concern and has been reported post intraosseous infusion in stable animals but has never been studied in animals subjected to cardiopuimonary resuscitation. we undertook this study to investigate the incidence and magnitude of lat and bone marrow embolism with the use of intraosseous infusion during prolonged cardiopuhaonary resuscitation and after fluid and drug infusion. we hypothesized that there will be no difference in the magnitude of fat embolism between cardiopulmonary resuscitation only and other cxperirnental conditions. thirty-one ( ) piglets were anesthetized, mechanically ventilated, and instrumented (carotid artery, pulmonary artery and intraosseous earmulas ). the animals then underwent bypoxic cardiac arrest followed by chest compressions with the mechanical thumper (michigan insmunents) and mechanical ventilation for a minimum of minutes. the animals were divided in groups: a (n= ) which had no intraosseous, ~'oup b (n= ) had intraosscous with no infi~ion, and groups c (n= ), d (n= ), e (n= ) had intraosseous with infusion of adrenaline, normal saline and sodium bicarbonate, at cessation ofcardiopulmonary resuscitation, representative lung samples were collected fi'om upper and lower lobes of each lung, embedded in ocp and firozen immediately. ltmg specimens were stained using oil red-o dye and observed for fat globules and bone marrow elements. the amount of emboli present was rated as a percentage in relationship to iung tissue, by a pathologist blinded to the experimental groups. buffy coat specimens were collected before and at cessation of cardiopuimonary resuscitation, stained with oil red-o dye and observed for fat globules. percentage of fat present were compared using analysis of variance. fat globules were seen in the prebronchial blood vessels and in intravascular areas throughout all lung fields. there was no difference in appearance or distribution of fat globules between groups. quantity varied in the different groups[(a) %, (b) %, (c) % (d) %, (e) %], but were not statistically significant (p = . ). fat globules in the buffy coat were few and inconsistent with lung findings. fat and bone marrow emboli were present in all experimental conditions, the use of the intraosseous cannula does not increase the magnitude of embolization during cardiopuimonary resuscitation. the decision to use the intraosscous route should not be influenced by the risk of embolization. tzareva iv/,, md*, nedialkova r, md**, *dept. of pathophysiol, *~dept. of child surg. and icu, emergency medical institute pirogov, sofia, among children with blunt abdominal trauma, treated in emi pirogov during the last five years, children had serious disturbances of the basic vital functions, connected with the trauma, and most often with massive haemorrhage, for this reason being an object of reanimation and intensive care. in the group of children who survived - , predominated the trauma of only one abdominal organ (mainly the spleen, rarely the kidneys, the intestine) and only children had injuries of more than one abdominal organ. in the same group, in children the abdominal trauma was combined with chest or head trauma or bone fractures. in the group of children who died - , a profound combined trauma was present. the haemodynamic parameters in all children showed a characteristically significant tachycardia along with normal or even high blood pressure, while hypotonia was present in only % of the children on the first trauma day. despite the fact that only . % of the children had direct chest injury as well, the gas exchange was considerably disturbed - ' of the children were hypoxemic during the first, and % during the third trauma day -in % significant -below . kpa ( mmhg). together with the markable decrease in haemoglobin levels, this determines the pronounced disturbance in oxygen transport. during the first trauma day all the children were acldo~c, and a metabolic alkalosis was present during the following days. twelve of the children with severe combined trauma died within several hours, with the symptoms of irreversible haemorrhagic shock, or in the next - days, developing multiple organ failure. in conclusion, the intensive therapy of children with severe abdominal and combined trauma, should take in consideration the special haemodynamical trauma answer in children, and requires dynamic monitoring of the most influenced homeostatic parameters -blood gases, acid-base metabolism, haemostasis. introduction: endocrine emergencies, other than diabetic ketoacidosis, are uncommon causes of pediatric intensive care unit (picu) admissions. we report our experience of children diagnosed of adrenal insuficiency (ai) admitted in the picu, during the last four years. subjects: five eases of ai requiring intensive care unit admissions are presented. four females anna male, with ages ranging from days to years, none of them had a previous systemic or endocrine diseases that could suggest al the initial clinical manifestations were: dehydration ( ), vomits ( ), abdominal pain ( ), seizures ( ), lethargy ( ) and hyperpigmentation in the muco-genitat area in a newborn male and ambigna genitalia in a newborn female. the reason for their admission in the p cu were: shock in two subjects; three because of hyperkalemia and hyponatremia (k/na: . / ; / ; , / meq/l); and two with severe hyponatremia (na: ; meq/l). laboratory findings: severe hyponatremia ( ), increased concentration of urinary sodium and chloride ( ); metabolic acidosis ( ); hyperkalemia ( ); increased levels of urea ( ) and hypoglycemia ( ). in all of them, the electrolytes abnormalities did not normalize with replacement and only normalized after the administration of hydrocortisone. tile ai was due to: autoimmtme disease in two subjects, congenital adrenal hypoplasia, congenital adrenal hyperplasia secondary to alia hydroxylase deficiency and in one no etiology was found, at the present time, comments: aiis an uncommon disease in the pediatric age. anearly diagnosis is crucial, as if the treatment is delayed could lead to patients death. in subjects with arterial hypotension and electrolytes abnormalities refractory to the usual treatment, they should be treated with corticosteroids, if no etiology is found. although, previously samples must be obtained to make the diagnosis, : denotes the number of cases. gerbaka b; hakme c; akatcherian c. toxics are frequently involved in domestic accidents during childhood; among non medical products ingestion, carbohydrate poisoning is a serious injury often made possible by inadequate stocking. over years, children aged years and less were examined in the emergency department of hotel-dieu de france hospital for carbohydrate ingestion. , % are boys; age goes from months to years (moan = , years). kerosene is found in , % of cases; all were admitted (mean = , days). , % were symptomatic on first examination but % of all children presented signs of gastric ( %) or respiratory ( , %) irritation sometime during their history; , % had neurological signs and , % presented some fever. leucocytosis is found in % of cases; , % of the children received antibiotics. chest x ray was abnormal in , % of cases: mainly parahilar infiltrates were found, all children survived; , % with a normal course ( , days of hospital stay) whereas those who presented complications (severe pneumonia, coma) stayed in the hospital for days (mean) with short course of assisted ventilation for two of them; long term follow up was not possible. we fonnd nick's criteria for hospital admission to be of value: -symptomatic children with normal x ray } to hours monitoring -asymptomatie children with x ray abnormality } -symptomatic children with x ray abnormality: hospital admission -asymptomatic children with normal x ray : no admission. these criteria would have helped to avoid admission in children and would have allowed a short t hours stay for more. we found chest x ray to be mandatory in carbohydrate ingestion; other tests were not helpful, aside arterial blood gases measurement in case of respiratory involvement; we now also advocate more restriction in antibiotic use. prevention remains efficient and should be stressed on. severe liver failure [slf] is a rare but severe condition in infants. we report our experience. patients: slf was defined as liver insufficiency with hepatic encephalopathy and a decrease in the level of factor v to below %. between and , infants (mean : mo) were admitted for slf (neonates excluded). main causes were metabolic disorders ( . %) (tyrosinemian= , hemochromatosis n= , reye's syndrome n= , other n= ), virus-induced flf ( . %) and hematologic diseases ( . %). in cases, the causes remained undetermined. results: olt was contraindicated in cases because of multiple organ failure (n= ), or underlying disease. all of them died within days after admission. patients had no indications for olt, all but one are alive. ( of them was transplanted later for tyrosinemia and died lately (virus induced-slf). among the t infants who underwent emergency olt, are alive and died because of primary non function of the graft. conclusion: slf in infants admitted before their first birthday is a severe condition with an overall mortality rate reaching %. inherited metabolic disorders are the first cause of slf at this age. contraindications for olt are frequent because of underlying disease or multiple organ failure. a number of children undergo primary graft failure after liver transplantation. it is unknown if there is any increased morbidity or mortality following retransplantation. this study seeks to explore these issues. methods: a pediatric intensive care/iiver transplant database is in formation. records of all liver transplant patients are reviewed and abstracted. this data is then computerized to allow analysis. this data provides the source for this study. statistical analysis was performed via student's t-test where appropriate. results: of the patients who have thus far received at our center orthotopic liver ransplants, the records of who underwent transplants form the basis for this review. twenty-three patients underwent multiple transplants, required one additional, three required organs, and one patient survived after a fourth organ transplant, there was no significant difference in age at first transplant between those who received multiple organs and those who did not ( vs, months, p=ns). the anesthesia time for the procedure did not significantly increase tbr subsequent transplants ( . vs, , hours), nor did time in the intensive care unit (t . vs. . days), nor did time on the ventilator ( . vs. . days) subsequent transplants did not predispose to having more bleeding in the intensive care unit for usage of packed red blood cells or platalets was not significantly altered ( vs ml and vs ml respectively). patients who required retransplantatior~ did receive mere fresh frozen plasma (ffp)daring their first transplant than in the subsequent ones ( vs ec, p < . ). however ffp use was not significantly different than patients who did not require retransplant. patients who underwent retransplant had a markedly increased mortality ( %) than the overall mortality for liver transplants at our center ( %), conclusion: children who require another liver transplant have a markedly increased mortality. bleeding and prolonged icu stay is not significantly different between the first and subsequent transplants, fulminant hepatic failure and ortothopic liver transplantation.dr.sasb n,j;centeno,m;entin,e;acarenza,m;ciocca, m:gofii,j;bianco,g;weller, g;imventarza,o. unidad de cuidados intensivos.hospital de pediatria "dr.j.p. garrahan" .buenos aires.argentina. introduction:fulminant hepatic failure (fhf) is a clinical syndrome, defined by the development of hepatic encefalopathy within weeks from onset of illness in a previously healthy person.by far,the most comun cause of pediatric fhf in all series, is acute viral hepatitis.we report our experiences with the pediatric fhf and ortothopic liver transplantation (olt) as attemative of treatment. patients: childrens with fhf diagnosis were admitted at the picu from / / to / / .symptomatic treatment was given to all children and all were put on list for olt,) following the king's college criterion (protrombina time,age,atiologies,bilirrubin,and encefalopathy state). results:etiologic causes corresponded to the childrens were: , hav ( %); , noa nob ( %); ,autoinmune ( %).the age was mean: years (range: month- years).seventeen patients were transplanted, chidmn were discarded because:no donors: ;withdrow of the list: ,because sepsis in and bleeding of cns ;and no admission at list: because genetic syndrome ,massive intestinal necrosis, ,mitral valvulopathy and sepsis, . patients ( %) had at least one complication dudng the post operative period.the most frequent was the acute renal insufficiency(ari) and patients requiered continuos hemofiltration.the gtobal mortality rate was %.the mortality of patients without olt was % and the mortality of patients with olt was %, patients dayed because sepsis, ( candidiasis) and the others because mof.the actuarial survival at year is % and the follow up of months. conclusions:the fhf is a very severe and frequent disease at picu. supportive treatment only is associated with a very poor prognosis and high mortality rate.the most frequent etiology in our country is the hav. the olt is applicable in this cases and is a valid alternative of treatment (mortality in our series %).the ari is the most frequent complication during the post opeative period.in argentina,due the high prevalence of hav,prevention must be considered the main and only way to avoid this catastrophic illness.- to assess the efficacy of gastric intramucosal ph (phi) for evaluation of tissular perfusion and prediction of hemodynamic complications m critically ill children. patients and methods: thirty critically ill children ( boys and girls) whose age ranged from month and years old were studied. a tonometry catheter was placed in the stomach of all patients at their °admission in pediatric icu. intramucosal ph measures were made at the admission and each - hours during the study: a total of determinations were made. the catheter was removed after extubation and/or checking of hemodyrmmic stability of the patient. the intramucosal ph was derived from application of the henderson-hasselbaeh formula using the pco value from the tonometer and the arterial bicarbonate. values of phi between . and . were considered normal. the relationship between phi and severity of patient measured through prism, presence of major (cardiorespiratory arrest, shock) and minor (hypotension, hypovolemia or arrhytlmtias) hemodynamic complications, mortality and stay in the picu, was analysed. results: the admission value of phi was . -t- . (range . - . ). five patients ( %) had an admission phi < . . no relationship was found between an admission phi < . and a higher incidence of hemodynamic complications. sixteen patients ( %) showed some values of phi < during their evolution. patients with phi < . had a higher number of hemodynanuc complications than the rest (p< . ). every cardiorespiratory arrest (cra) and shock cases were related to a phi < . . patients with major complications (cra and shock) had a phi lower (p= . ), as well as a higher number of measurements of low phi (p= . ) than patients with minor hemodynamie complications. the value of phi lower than presented a % of sensibility and % of specificity with regard to hemodymanic complications. there was no relationship between phi < . and prims score and stay in picu. patients with phi < . presented a prims higher than the rest of patients (p< . ). conclusions: the phi value may be an early sign of presence of hem dyaaimc complications in the critically ill child. we tested the hypothesis that gastric intramural ph (phi) can be used as an early sign of failure m weaning pediatric patients because the blood flow from nonvital areas is diverted to meet the increased demands of respiratory muscles. methods: children (mean age ( . _+ . ) years + sd) who were thought by their physicians to be weanable from mechanical ventilation (mv.). these patients were ventilated on serve c ventilators, receiving ranitidine, and had intestinal tonometer (tonometrics, inc.) minutes before obtaining a sample.. all children were placed on pressure support (ps) at levels judged to overcome the resistance of the endotracheal tube and ventilatory circuit ( em h.,o). a sample of arterial blood and a sample oftonometer were obtained during vm and weaning (ps). phi, hemodynamic and respiratory data were recorded during vm and weaning we did not interfere with the primary caretaker's decisions regarding extubation. patients were considered to be successfully weaned if they were able to sustain spontaneous ventilation for more than hours after extubation. paired t-test were used to compare the values obtained during mechanical ventilation with those obtained during weaning trials. unpaired ttest were used to compare values from the group that was successfully weaned (a=i ) with those from the group that were not (b= ). results: we did not find statistical differences in any of those variables mesured during mv for patients who were successfully weaned(group a) and those who were not (group b). gastric phi was in group a: . + . (vm) and + . (weaning); in group b: . _+ . (vm) and . t _+ . (weaning). discussion: although we did not find differences in gastric phi during vm, the group a had a lower value than group b because of the number of cardiac patients ( %) and transfusion therapy, in fins group. in group b % of patients showed a problem in upper airway (subglottic edema, and enlarged tonsils). we found it after extubation. conclusion: ) gastric phi is a good predictor of risk in critically ill patients but maybe because of the small size of the sample, in our study is not of practical value as a predictor of failure in weaning pediatric patients from vm. ) this test is not a predictor of problems in upper airway~ important etiology of failure weaning in children. objectives: i-to determine the prognostic value of the gastric intramueesal phi in mortality and multiple organ dysfunction (sdmo) in critically ill children. -to compare this value, with the pediatrics risk index mortality score (prims). methods: aprospective study was performed with critically illcbildren, aged from mouth to years. the athnittiug diagnosis was: post-surgery ( neurosurgery, spinal fusion and thoracic or abdominal surgery), sepsis, polytraumatism, adult respiratory distress syndrome and with miscellaneous. all the subjects were monitorized on picu admission and treated for their underlying condition. gastric intramucnsal pt{ was measured following the tonometric method, ou admission and every - hours depending on the patients state. the severity of the clinical condition was evaluated using the the prims, on admission (prims-i) and during the first hours, when the clinical condition deteriorate, the worse score was utilized for the statistical analysis (prims- ). to perform the statistical analysis the subjects were divided in two groups, one with the phi< . and the other with phi> . .aunivariate analysis (student's tand wilcoxon two tailed test, chi-square) and multivariate analysis were used. results: out of the subjects dyed. of children developing multiorgan failure (mof) expired. % of the patients admitted to the picu with sepsis, ards and miscellaneous had a phi < . . in contrast, with % of post-surgical and none of the postqraan~atism. the mortaliry rate, in children with a phi< . was % (ci %: . ; , ) and . % (ci %: , ; . ) in children with phi> . (p= . ). mofwas observed in , % of children withphi< . v.s, . % with phi > . .no relatiouship was observed between the phi and the score of prims-i and . perforating an unconditional logistic regression analysis, two independent variables have mortality predictive value: the phi and the prism- . (table i) following induction of anaesthesia, a laser doppler probe (moorsoft instruments ltd) was inserted cm into the patient's rectum, the probe's special design ensuring that the optical prism lay against the mucosa. continuous monitoring of rectal mucosal perfusion ("flux") was continued throughout the operation. after rain cpb at °c, "steady state" readings of nasopharyngeal temperature, mean femoral arterial pressure (map) and flux were recorded over a further min before cpbinduced core cooling to - °c. steady state was defined as a rain period with no change in core temperatures or map. other rain steady state recordings were taken immediately prior to low flow, immediately prior to rewarming and after rewarming to °c, before initiation of any vasoactive drugs. the cpb flow rate was kept at m l k g - min q, the pcv at _+ %, the p~co at . + . kpa and the pro at + kpa. results: initial warm and rewarm map (both mmhg) were significantly lower ( = . ) than during the cold cpb periods ( & mmhg). the mean cold flux before ( ) and after ( ) low flow were both significantly lower (p= . ) than the mean initial warm cpb flux ( ). the mean rewarm cpb flux ( ) was significantly lower than all other flux values (p= . ). there were no siglaificant correlations between map and flux except at the first warm cpb period (r= , , p= . ). conclusions: although hypothermia significantly reduces rectal mucosal perfusion, rewarming produces an even greater reduction in gut perfusion which, considering that mucosal oxygen constmaption is highest during this time, may prove crucial in the postoperative development of mof. therapy aimed at improving gut perfusion during cpb should be directed at the rewanning period in particular. abstract this work is aimed at establishing a clinical procedure for the diagnosis of enteritis necroticans (en), even at the communal level, and to define criteria for diagnosis able to distinguish between acute forms. subjects and method : cases admitted at the institute for protection of children's health dpch), having characteristic symptoms, were examined clinically, by roentgenography of the abdominal cavity, with the analysis of the blood (total protein, electrolytes, hematocrite) and cultures of intestinal fluid and faeces. through surgical operations, the pathological lesions were observed and recorded. results: common epidemiological features: the average age is - years old ( - ) ; male/female : . ; in % of the cases, the disease occurred after a meal rich in protides. the acute toxic form accounted for % : severe shock appearing early, with very severe dehydration associated with profoundly decreased blood protein concentration and lowered natriemia as well. the lesions of the small intestine were expanded, all of them were necrotic. in the surgical form ( %), the predominant feature was an obstruction -peritonitis syndrome, the peritoneal fluid showed a characteristic inflammatory reaction. for the rest of cases % were the internal form, the shock syndrome was less severe, the abdominal distention was light and disappears gradually, the inflammatory reaction of the peritoneal fluid was not so characteristic. conclusion (ino) is a selective pulmonary vesodilator that is rapidly inactivated compared to intravenous vasodilators. these qualities make ino an attractive agent for the treatment of pulmonary hypertension (pittn). the efficacy of ino has been studied in persistent fetal circulation, acute respiratory distress syndrome (ards), and congenital heart disease (chd). potential adverse effects oflno include: nitrogen dioxide (no toxicity, methemoglobinemia, and platelet dysfimction. our objective was to evaluate the safety of ino in pediatric patients (pts). methods: pediatric pts. with phtn from ards or chd were studied under an established, approved protocol conforming to fda guidelines tbr an investigational new drug. informed consent was obtained for each child prior to treatment. no was sequentially titratad from parts per million (ppm) to , , , and ppm at ten minute intervals. parameters monitored before and during therapy included nitric oxide (no) and no~ concentrations (cone.), mean arterial blood pressure (map), and percent methemoglobin (mhg). no and noz levels were continuously monitored using an inline dr~ger electrochemical detection device. ~,litp was continuously measured with an indwelling arterial catheter. mhg was measured by co-oximetry. a mhg level e % or no cone. ~ ppm were considered adverse effects by study criteria. pretreatment map was compared to map at and ppm ino using paired t-tests. ap value < . was considered statistically significant. results: thirty-two mechanically ventilated children with phtn ( with ards, with chd) were studied. five pts. were treated following cardiopulmonary bypass. methemoglobin (met-hb) levels were routinely measured in two prospective clinical studies on no inhalation in pediatric patients with pulmonary hypertension following heart surgery with extracorporeal circulation and in pediatric and neonatal ards patients, the observed differences between the groups prompted in an in vitro study, red blood cells (rbc) of patients sampled before and after surgery with and without extracorporeal circulation (ecc), respectively, were incubated with ppm no for rain, met-hb, atp, and nadht nadph concentrations were compared, during therapeutic exposure no increased met-hb from . - -_ . to . _+ . % in cardiac surgery patients and from . ± , to . ± . % in ards patients (p < . ). rbc's having undergone ecc were more susceptible to met-hb formation (p< , ) whereas intracellular coenzymes did not differ neither between the groups (table) nor before and after no exposure. ecc predisposes to increased methemoglobinemia upon exposure to no both in vivo and in vitro. our data suggest a reduced activity of met-hb reducing enzymes rather than diminished availability of energetic substrates, variation of the inhaled nitric oxide concentration with the use of a continuous flow ventilator. anne pmc de jaegere ~, frans im jacobs , nico gc laheij , john n van den anker t . dept. of paediatrics ~, central instrumentation , sophia children's hospital, erasmus university rotterdam, rotterdam, the netherlands. objective: to investigate the homogeneity of nitric oxide (no) concentration in a delivery system with a continuous flow ventilator. design: bench study, setting: biomedical laboratory. interventions: a nitrogen/nitric oxide (njno) gas mixture was injected at three different sites in the patient circuit: just before and just behind the humidifier, and centimetres before the y-connector. ventilator flow ( , , l/rain), ventilator rate ( to , increments of ) and compliance of the testlung ( . ; . ; . ml/cm h ) were changed. carbon dioxide (co ) instead of n /no was injected at the same points in the circuit. measurements and main results: a) though the flow ratio of the njno and the ventilator gas were kept constant, the no concentration ([no]) raised with increasing ventilator rates. the increase in [no] was up to % when the n /no injection site was close to the y-connector of the ventilator circuit. minimal changes in [no] were noticed when the n~/no was mixed to the ventilator gas before the humidifier. b) analysis of the ventilator flow pattern showed variations at different places in the ventilator circuit. the magnitude cf the p, ow change depended on the meas~:rement site. the closer to the expiratory valve the highest the flow change was. the duration of the flow change was inversely proportional to the adjusted ventilator flow. c) real time measurements of the co concentration ([coz]) showed variations during tile respiratory cycle. these [co ] variations were higher when the co gas was blended closer to the yconnector. conclusions: the ventilator flow variations in relation to the fixed side flow of the n /no gasmixture result in changes of the inhaled [no] during the respiratory cycle. the no concentration during inspiration is always higher then during expiration. this could not be detected with the available monitoring system. to ensure a constant [no] by blending a njno gas balance in a continuous flow ventilator, the site of injection should be as close as possible to the inspiratory outlet. nitric oxide, a potent and selective pulmonary vasodilator, has recently been successfully used to treat pulmonary hypertension of variable etiology in infants and children. side-effects and complications in infants are so far not well known. we describe here two cases in which prolonged ( and- days respectively) high-dose ( - ppm) nitric oxide was used to treat refractor~¢ pulmonary hypertension. one patient was a newborn infant with pulmonary hypertension secondary to a large leftsided diaphragmatic hernia. nitric oxide was begun under conventional ventilation (babylog ) at hours of life with a slight initial improvement in oxygenation. he was then placed on oscillation with the same nitric oxide concentration due to worsening respiratory failure. he died on th day of life. monitored nitric dioxide concentration never exceeded ppm. the other patient was a months old infant with severe pulmonary hypertension due to a complete atrioventricular septal defect. he required high-dose nitric oxide to come off cardiopulmonary bypass after surgical repair of his heart defect. he slowly improved over the week following surgery but developped suddenly respiratory failure due to massive pulmonary hemorrhage and died. surprisingly, a particular autopsy finding in both infants was a massive acute necrotizing tracheobronchitis. we conclude that nitric oxide is an excellent and sometimes lifesaving treatment of pulmonary hypertension in infants. tracheobronchitis has not yet been reported as a possible complication of nitric oxide administration. we suggest that caution needs to be taken with prolonged high-dose administration and this possible complication to be looked for at autopsy. introduction: permissive hypereapnia (ph) is a beneficial strategy for patients with acute respiratory distress syndrome (ards) to minimize barotrauma by decreasing the peak inspiratory pressure (pip). hypercapnia and hypoxia cause pulmonary vasoconstriction, pulmonary artery (pa) hypertension, and, thus, an increased afterload to the right ventricle. this increased afterload may result in increased right ventricular (rv) work load and subsequent rv dysfunction. one therapeutic approach is the use of inhaled nitric oxide (inn), a selective pa vasodilator. the objectives of this study were to test the hypothesis that in a swine model of ards with ph, inn would improve rv work load and not change intrinsic rv contractility. methods: in swine ( - kg), ards was induced by surfactant depletion. hypercapnia was achieved by decreasing the pip while increasing the peep to maintain a constant mean airway pressure, inn was administered in concentrations of , , and ppm in a random order. pulmonary blood flow (qpa) was determined by an ultrasonic flow probe. rv total power (tp) and stroke work (sw) were calculated by fourier transformation of the pa pressure (ppa) and qpa data. preload recruitable stroke work (prsw), a preload and afterload independent measure of ventriculur contractility, was determined by a shen-subtraction method and vena caval occlusion. respiratory failure with pulmonary hypertension in piglets gerfried zobel*, bernd urlesberger*, drago dacar**, siegfried rtdl*, fritz reiterer* and ingeborg friehs** depamnents of pediatrics* and cardiac surgery**, university of graz,austria objective: to evaluate gas exchange, pulmonary mechanics and bemodynamic data during partial liquid ventilation (plv) combined with inhaled nitric oxide (no) in acute respiratory failure with pulmonary hypertension. design: prospecfive~ randomized, controlled study. setting: university research laboratory. subjects: twelve piglets weighing to kg. interventions: acute respiratory failure with pulmonary hypertension was induced by repented lung lavages and a continuous infusion of the stable endoperoxane analogue of thromboxane. thereafter the animals were randomly assigned either for plv or conventional mechanical ventilation. initially perfhiorocarbon liquid ( ml/kg) was instilled into the endotracheal tube over min followed by - ml/kg~. all animals were treated with different concentrations of no ( - - ppm) inhaled in random order. measurements and results: continuous monitoring included ecg, cvp, mpap, map, san and svo measurements. during plv pao /fio increased significantly from _+ . mmhg to ± mmhg (p< . ) within rain, while pao ]fio remained constant at -+ . mmhg. qs/qt decreased significantly from -+ % to -+ % (p< . ) during plv and did not change during conventional mechanical ventilation. static pulmonary compliance (cstat) increased significantly ff~m . r± . to . _+ . ml/cmh /kg (p< . ) during plv and decreased slightly from . _+ . to . e . ml/cmh /kg during conventional mechanical ventilation. the infusion of the endoperoxane analogue resulted in a sudden decrease of pao /fio from _+ to _+ . mmhg in the plv group and from ± to +_ . mmhg in the control group. inhaled no significandy improved oxygenation in both groups (pao /fio : _+ mmhg during plv and +_. mmhg during conventional mechanical ventilation). during inhalation of no mpap decreased significantly from -+ m ± mmhg (p< . ) in both groups. there was no significant change in oxygenation and mpap during inhalation of and ppm no. conclusions : plv significantly improves oxygenation and pulmonary compliance in acute respiratory failure. the additional application of inhaled no further improves oxygenation and pulmonary hemodynamics when acute respiratory failure is associated with severe pulmonary hypertension. inhaled no is very effective in improving oxygenation and pulmonary blood flow even at low doses. the work was supported in part by grants of the austrian nationalbank nr . as in neonates, severe respiratory failure in infants and children can be aggravated by pulmonary hypertension, resulting in further deterioration of oxygenation due to increasing intrapulmonary shunting. we analysed the influence of inhalational nitric oxide (ino) in treatment, course and outcome of severe ards in a pediatric population. since infants and children (age: - months) with ards and oi > (mean value: . ± ) underwent a trial with ino (concentration: , , , and ppm) to prevent further respiratory failure. patients had a significant improvement of their oxygenation (rise of pa > mm hg) for at least hours (responders); mean best ~fficient no dose: . ppm. the non-responders had only a short-term improvement or ino had no effect. in responders and nonresponders there was no significant difference with regard to age, underlying disease, ards severity, time on mechanical ventilation, blood gases and ventilator settings before notrial, nor was there a different grade of pulmonary hypertension (estimated by echocardiography). the only difference was an higher ol in the group of the non-responders: . ± .i vs. . ~ . , p < . . in the group of the respenders there was a secondary deterioration of lung function after i - days on ino in children (transient responders): in these patients, as well as in the group of the non-responders, alternative modalities of treatment (hfov and/or ecmo) became necessary. children ( %) died: transient respenders and non-responders. in infants and children with ards due to different underlying diseases ino can acutely lead to a significant improvement of oxygenation in about % of the cases. the right selection of patients for no therapy and the influence of ino on the survival rate of ards in childhood has to be evaluated in further studies. and pediatric cardiology, university of graz, a- graz purpose: after fontan procedure cardiac output is critically dependent on the pulmonary vascular resistance. even minor elevations of the pulmonary vascular resistance may significantly decrease cardiac output. inhaled no is an effective, selective pulmonary vasodilator in experimental and clinical situations of pulmonary hypertension. the aim of this study is to evaluate the effects of inhaled no on oxygenation and pulmonm , circulation in children after a bidirectional glenn-anastomosis (n-~) or a fontan-like operation (n= ). material and methods: from june t to january children with a mean age of . +~ . (sem) yrs and a mean body weight of . -+ . (sem) kg were treated with inhaled no after glenn-or fontan-like operations. all but one had complex cardiac malformations with single ventricle. all children were mechanically ventilated with an fin > . . inhaled (no) was applied using a rrdcrdproeessor based system which additionally allowed measurement of no/nox using the chemihimniscence method. methemogtobin concentrations were determined times a day. the major indication for postoperative inhalation of no was a high (> mmhg) transpulmonary pressure gradient (tpg--cvp-lap). severe myocardial dysfunction of the single ventricle was excluded by echocardiography. results: the mean duration of mechanical ventilation was . _+ . (sem) days the. mean dose of inhaled no was . -+ . (sem) ppm, the mean duration of no-inhalation was _+ (sem) hours. the mean methemoglobin concentration was . -+ . (sem)%. hemodynamic data and arterial oxygen saturation before inhaling no and minutes later are given in table acute hypoxaemic respiratory failure (ahrf) in children occurs in a heterogenous group of diseases with pulmonary pathophysiological processes ranging from reversible physiological intrapulmonary shunting to fixed structural lung damage. we hypothesized that inhaled nitric oxide (ino), a selective pulmonary vasodilator, might identify those patients with potentially reversible disease, i,e, large response may indicate a greater likelihood ef reversibility and thus survival. a retrospective review of the early response to ino in infants and children (aged month to years, median months) with severe ahrf( with ards). the mean p(a-a)o , pao / fio , oxygenation index (oi) and acute lung injury (all) score prior to the commencement of ino were +_ . , +_ . , _+ , and . +_ . respectively, the magnitude of response to ino was quantified as the % change in oi occurring within minutes of ppm ino therapy. this response was compared to patient outcome data. results. there was a significant correlation between response to ino and patient outcome, kendall tau b r= , , p< . (table) conclusion. in ahrf response to ino appears te define a subgroup of patients with improved outcome compared to nonresponders. we speculate that response to ino may be useful in selecting patients with potentially reversible lung disease for special support therapies such as ecmo. randomised controlled trials are needed to define the role of ino in paediatric ahrf. between may and december , patients (pts) were treated for mas. treatment groups were: group i only : pts; group i conventional mechanioal ventilation (cmv): pts; group ii hfo: pt; group iv hfo+no: pts. therapy was stepwise intensified until oxygenation improved ( i -) ii -) iii --) iv). "high volume strategy" was used with hfo (mawp - cm h ). the initial no-concentration was - ppm, with rapid reduction down to - ppm once oxygenation improved. results: one pt (group it) died of hypoxic-ischemic encephaiopathy (termination of therapy); all other newborn babies survived. in group iv pt and showed barotrauma prior to hfo. pt , and were treated with additional mgci (max. mg serum concentration . - . mmol/i). following the identification of inhaled nitric oxide "no) as a selective pulmonary vasodilator (frostell et al ) [ .+ , + . data are compared to baseline values within each group. *=p< . , **=p< . , ***=p< . l among patients who fulfilled ecmo criteria, improved with no and did not required extracorporeal life support. tltree out of ecmo patients eventually survived. conclusions: m our study low-dose of irthaled no showed a variable effect on oxygenation in newborns with acute respiratory failure. an acute response to no appeared to be correlated with a better short-term outcome and the avoidance of extracorporeal support in ecmo candidates. differently, lack of acute and/or sustained response was associated with death or need for ecmo. although the nature and severity of the underlying disease or the degree of prematurity may play an important role in these patients, we believe lack of acute response to no may be an early predictor of bad outcome, prompting toward alternative treatments such as ecmo or liquid ventilation. *picea s., °bartuli a.,°dionisi-vici c., *dello strologo l., §villani a., §bianchi r., ^salvatori g.,*rizzoni g, °sabetta g. *div. of nephrology, °div. of metabolism, §intensive care unit, ^div. of neonatology. "bambino gesfl" children research hospital. rome, italy. successful prevention of handicaps or death in newborns with ~ depends on rapidity and efficiency of treatment. poor response to nutritional and/or pharmacological treatment requires extracorporeal removal of nh . efficiency and cardiovascular tolerance are often difficult to obtain with peritoneal or hemodialysis in neonates. we report the results of cavhd in newborns with hc. methods: vascular access: femoral vessels. blood flow: - ml/min, dialysate flow: - ml/h. filter: amicon minifilter plusrm(polysulfone membrane; . sq.m.). no ultrafiltrate(uf) production, patients: case with carbamoytphosphate synthetase deficiency (body weight -bw-: . kg) showed hc at day , a relapse of hc occurred at day due to an infectious event. case and (bw: . and . kg), both affected by propionic aeidemia, showed hc at day and day , respectively. plasma nh (~tg/dl) decrease is shown in the complications: transitory ischemia of arterial cannulation limb and transitory thrombocytopenia occurred in case ; surgical repairing of artery after cavt-id was necessary in case ; no cardiovascular instability was observed during cavhd . outcome,'all patients recovered from hc in less than day: case : alive, mild b)iootonia at mos; case : dead after days from cavhd withdrawal for pulmonary hemorrhage; case : alive, normal development at mos. conclusions: ) in newborns with hc, ca~q-id provides good cardiovascular tolerance,high efficiency and quick removal of nh , even without uf production (i.e. only by diffusion). this allows easier management (no need of fluid and electrolyte balance). ) arterial complications seem frequent in neonates treated by cavhd. venovenous circulation could overcome this problem. vb nguyen, m jokie, c leeaeheux paediatric intensive case service, hospital university centre, avenue c te de nacre, caen cedex, france background, the implication of polymorphonuclear neutrophils (pmns) in the physiopathology of children's haemolytic.uraemie syndrome (hus) becomes more and more evident. the purpose of the present study is to role out their impact among other pronostie elements during the course of the disease. patients and methods. diarrheal prodrome and its duration, patient's age, maximal blood nitrogen level, anuria and dialysis time, extra.renal involvements, white enll and pmn counts and thrombopenia duration have been retrospectively analysed in infants with good outcome and in another children with unfavorable outcome. results. neither diarrhoea or its duration, nor children's age, nor blood nitrogen level, nor anuria or dialysis time had any predictive value for the disease evolution in the acute phase of our patients. adversely, extra-nenal involvements was accompanied by severe and complicated courses of the disease (p< , ). the elevation of white cells and pmns (heyon x /i) and pmns (more than x / ) as well as its persistence beyon a week were most frequently observed in complicated forms (p< , , p< , and p< , , respectively). a transient thrombopenia (less than day@ in patients with elevated counts of white cells may be a filrther obvious sign of an unfavorable course of the disease ( < , ). conclusion. the elevated count of white cells and pmns, either alone or associated to one rapid regeneration of platelets, seems enabled to predict an unfavorable evolution of the hus in children. msud results from an inherited impairement of catabolic pathway of branch chain amino-acids. high leucine blood levels may induce acute brain dysfunction. this dramatic complication led us to propose leucine removal procedures as continuous hemofiltration. patients and methods three newborns in acute msud onset were treated by hf, hdf and hd. extracorporeal circulation was performed through a . fr catheter, a circuit with a blood pump (priming volume = ml). patients and procedures characteristics are summarized below in the sucralfate (an aluminium salt of sucrose octa sulfate) is used to prevent and treat upper gastrointestinal bleeding in critically ill patients. with minimal absorption, the potential for side effects is thought to be limited, though aluminium toxicity has been reported in patients with chronic renal failure. these patients may already have had high body stores of aluminium. we report critically ill children with high serum concentrations of aluminium following sucralfate therapy. all had renal impairment. the normal aluminium level is < . gmol/l and in patients with chronic renal failure < . ].tmol/l. none of these patients had known preexisting chronic renal disease. cpb was conducted under deep hypothermia (t,° °c) and cardiocirculatory arrest (cca) or under hypothermia (t,° °c) and low-flow perfusion. continuous holter-electrocardiograms (h-ecg) were recorded from the ilranediate postoperative (po) period on for hours. h-ecg were also recorded prior to the operation and before discharge. following dr were observed: snpraventricutar (sv) and ventricular (v) extrasystoles (es) (> / h), sv and v tachycardia (svt and vt), accelerated junctional rhythm (ajr) and junctional ectopic tachycardja (jet), and nd and rd degree atrioventricular block (avb and avb ). the incidence of po dr was % in the pre-op h-ecg, % on the st, % on the rid, % on the rd po day and % befbre discharge. compared to the pre-op findings, an increased incidence of sves, ves, svt and avb on the st po day was observed, whereas vt and a jr or jet were exclusively observed po. all types of dr were observed up to the rd po day. ty e of dr before discharge was similar to pre-op findings and there was no definitive avb . considering patient groups according to the most frequent isolated op-procedure, the incidence of dr on the first po day was % after asd ii-closure (n= ), % after stthaortal vsd-closure (n=lg), % after correction of a complete avsd (n= ), % after correction of a tetralogy of fallot (n= ) and % after fontan-operation (n= ). incidence and type of dr were not significantly different between groups. longer cpb-dttration and use of cca were risk factors for po ves and vt (p< , and p< , , respectively) whereas use of cca and degree of hypothermia were risk factors for the development of a jr and jet (p< , and p< , , respectively). -our results indicate that po dr after cpb in children m'e frequent but mainly transient. in our series, specific cpb-related parameters are of greater influence than surgical procedure itseif for the development of dr and are discriminant risk factors for particular types of dr. the course of anp, cgmp/anp (as indicator for atrial natriurefic peptide biological activity), and no and no (as indicator for endogenous nitric oxide (no) synthesis) was investigated in i infants (median age months) undergoing cardiopulmonary bypass (cpb). patients were divided into groups according to whether they had (group , n= ) or not (group , n= ) preoperative heart failure (hf) and pulmonary hypertension (pht). group patients had preoperatively significantly higher levels of anp (p< . ), cgmp (p< . ) and no and no (,p< . ) but had significantly lower cgmp/anp (i < . ) than group patients. during cpb, anp was significantly higher in group patients ~< . ). as compared with prebypass values, cgmp/anp was reduced in both groups during cpb (p< . ). cgmp/anp inversely correlated with duration of cpb and aortic clamping time (p< . , respectively). no and no were significantly higher in group than in group patients (p< . ) without any intraindividual change during cpb. from the early postoperative period on anp, cgmp/anp and no and no were similar in both groups. after cpb, anp correlated in both groups with blood pressure (p< , ) and diuresis (p< . ). no and no inversely correlated with pulmonary arterial pressure immediately after cpb ( < . patients after a fontan-type of procedure have elevated central venous pressures (cvp) leading to congestion in the gastrointestinal system and often ascites. purpose of this study was to evaluate whether this causes a different postoperative gastric mucosal ph (phi). methods: we evaluated a series of patients, who underwent cardiac surgery with cardiopulmonary bypass (age: days to years (mean , yrs), weight: . to kg (mean . kg). a commercially available tonometer (tonometics®) for sigmoidal use in adults was inserted into the stomach after induction of anesthesia. the phi measurements were done according to manufacturer recommendations we compared three groups of patients: ) aeyanotic (n= ), among them p with vsd and p with avsd; ) cyanotic (n= ): tof: p, tga: p; ) cyanotic after a fontan-type procedure (n= ). phi were measured at picu arrival and after h. fudhermore we compared lactat levels at these time points. differences between the groups were evaluated with one way anova on ranks with pairwaise multiple comparisons (dunn's method). the relationship between cvp and phi was investigated by regression analysis. results: the median phi for groups i, and were . , . and . at ardval and . , . and . after h respectively. at picu arrival group was significantly (p< . ) different from groups and . there was no significant difference between the latter two groups, after h group was different from group , there were no other significant differences. the median lactate levels for groups t, and were . , , and . at ardval and . , . and . after h respectively. at ptcu arrival group was significantly (p< . ) different from group , after h there were no significant differences. there was a weak negative correlation between cvp and phi: r= - . ; p< . . conclusion: patients after a fontan-type of procedure have lower phi than patients after other cardiac surgical procedures, however, this is only in part due to the elevated cvp and venous congestion. eleven children were investigated months (median) after postoperative mof. iviof was defined as the failure of at least two vital organ systems (kidney, liver, lung, central nervous system) in addition to cardiac insufficiency and high fever. underlying surgical procedure was repair of tetralogy of fallot (n= ), fontan-(n= ) or seuning procedure (n=l). all patients fulfilled criteria for mof in the first postoperative (po) days. six patients needed peritoneal or hemodialysis for days (median) during the po period. one patient showed cerebral infarction due to thromboembolism in the territory of the right internal carotid artery immediately after the operation. the follow-up protocol consisted of extensive investigations of heart-, renalliver-, and lung functions as well as complete neurological and psychological examinations. all patients had adequate cardiac examination. lung function was normal in all but patients who had an obstructive syndrome. only patient showed an isolated decreased creatinine clearance. abnormalities of the liver ftmction tests were only noticed in patients after fontan procedure. severe neurological sequels such as paraplegia (n = ) and diplegia (n-i) were observed in of the patients. the remaining children presented with a delayed graphomotorical and speech development associated with normal intelligence. -in our series the most frequent and severe sequels after postoperative mof were neurological. -abnormal liver fimction tests are more likely to be a consequence of the fontan hemodynamics than a sequel of mof. the optimal dosing schedule of surfactant therapy for the treatment of neonatal respiratory distress syndrome (rds) remains unclear. goal: surfaetant function and the concentration of phospholipids (pl) in tracheal aspirates are compared in a prospective randomized trial involving neonates with rds who received either two or more ( or ) doses of survanta. methods; ventilated neonates < w with rds were treated with survanta oo mg/kg if fio >_ % or mean airway pressure _> , cm hzo, after h a nd dose was given (same criteria), if the support still exceeded the criteria h after the nd dose, the patient was randomized to no extra dose (two}, or to an extra dose of survanta (morel (and a th dose h later; same criteria), pl was measured in tracheal aspirates and corrected for dilution with the urea method. "active" large aggregates and "non-active" small aggregates of surfactant were separated by centrifugation and quantified. surface tension of the large aggregate fraction was measured by pulsating bubble surfactometer, results: neonates were randomized, x two and x more ( x and x doses), gestational age was , ± , w and birth weight ± g. most patients had severe rds with initial ventilation: rate . _+ , , peak inspiratory pressure (pip) , -+ . cm hzo, fio . ± . %. at randomization: rate . ± . , pip . -+ . cm hzo, fio . ± . %, and h after randomization: rate . ± . , pip . _+ . cm hzo, fio . ± . %, without signif, differences between the groups. there was relapse (again fio _> % within h) in group two and t bpd in group more. in total, tracheal aspirates were analyzed. pl was not signif, different before randomization (two . ± . vs more . ± . /jmol/ml), but neither after randomization (two . -+ . vs more . ± ,o /~mol/ml). there was no difference in the % small aggregates (two . ± . vs more . ± . %), the surface tensions (ran/m) were not signif, different (each time two vs more): before randomization . ± , vs . -+ . , in the h after randomization . ± . vs . -+ , , or - h after randomization . -+ . vs . ± . , or - h after randomization . _+ . vs . -+ . . conclusion: neonates who received more than two doses of survanta did not have higher pl, nor a better surfactant function than neonates who received only two doses of survanta. continuation of the trial is necessary to evaluate clinical outcome. may not indicate need for treatment p.c. clemens s.j. neumann university of hamburg, department of pediatrics, klinikum schwerin, wismarsche str.. , d- schwerin. aim of the study: the finding of elevated tsh and decreased t in the newborn usually is classified as "transient hypothyroidism", thus the elevation of tsh is classified as consequence of the lowered t . but on the other hand several data sets show that tsh elevation as well as low t , one independently of the other one, are associated with different kinds of perinatal stress. each of these laboratory deviations, if not associated with the other value being abnormal too, is generally accepted not to be an indication for treatment. from this we conclude, that more pefinatal stress, as in intensive care neonates, may produce tsh elevation as well as low t , but only coincidentially, not the tsh elevation being the consequence of low t , thus not to be classified as "hypothyroidism", thus not indicating treatment. if this hypothesis is right, we should find an association of increasing pefinatal stress with an increasing number of neonates from tsh and t normal via tsh or t abnormal to high tsh and low t . method: in the newborn screening program in germa w we determine primarily tsh, and only in the neonates with elevated tsh, in addition we determine t . thus in our study we asked whether we find an association of increasing perinatal stress with an increasing number of neonates from tsh normal via tsh abnormal while t normal to high tsh and low t . definitions for this study were: tsh elevation = > mu/ (as usual in the german screening programs), t lowered = < p_g/dl perinatal stress score was or or or in dependency of the neonate having stress in none to all of the following three categories: (a) forceps or vacuum extraction or sectio co) birth weight below g (c) at the th day existence of a relevant neonatal disorder (rds, ictems gravis, infection/sepsis, vitium cordis with hemodynamic relevance, severe malformation). results: our data of neonates show a high significant association (chi = , p < . ) of, on one hand, perinatal stress score with normal tsh, versus, on the other hand, perinatal stress score or with high tsh and low t . discussion: facing the background given above, in the intensive care newborn, the constellation of high tsh and low t may be only a coincidential addition of two independent abnormalities. in tbese cases -the high tsh not being the consequence of low t -the classification as "hypothyroidism" is not justified, thus a therapy not indicated. on the other hand of course there exist rare cases with high tsh as consequence of low t thus with hypothyroidism tlms with indication for therapy. unfortunately we have no criteria, that enable a certain discrimination of these two categories thus in respect to the question of therapy or not. conclusion: further research has to be done to learn how to discriminate the coincidential high tsh and low t from the causal constellation of high tsh and low t . until we have certain discrimination criteria we have to treat both groups of neonates. few studies have focused on fa composition of surfactant pc in preterm infants before and after surfactant therapy. methods: tracheal aspirates were collected in venttlated mfants from birth until extubatlon ( / _ /twk ga, .+ g bw). after lipid extraction, t.l.c,, and methylation, fas of pc were quantified by gaschromatography. intralipid a ( . % linoleic acid, : • ) was started h after birth. results: six infants developed respiratory distress syndrome (rds) and received survanta r i mg/kg (sr), all doses within h after birth (ix s r n=l, x s r~ n= , x s r n= ). one child did not develop rds. in alt patients, the patmitate % in pc was ~ % (before sr<=natural composition), increased to ~ % after s r, and remained > % for i h after lx s a, . .+i . h after x, and . .+ . h after doses. in patients, intubated long enough, the palmitate % decreased with a half-life of . _+ . h to a new plateau which was still higher than baseline after week. linoleic acid % was . _+ . (with rds), decreased after s r~ and returned to baseline due to the decrease in patmitate %. thereafter the linoleic acid % increased linearly with . % per h, in patient even up to . %. other fas did not increase after return to baseline. in neonatal medicine the current parameters, arterial oxygen saturation and arterial oxygen pressure, are poor indicators for oxygen delivery and oxygen demand. the purpose of this study was to obtain venous blood samples from the inferior vena cava in stable neonates with respiratory failure and to determine a parameter that reflects more adequately the balance between oxygen delivery and oxygen demand. "l~e study included neonates requiring mechanical ventilation tbr severe respiratory insufficiency. an umbilical venous and arterial catheter were inserted in the inferior vena cava and in the aorta respectively. paired blood samples were obtained at the time that the patients were hemodynamically stable. fifty paired arterial and mixed venous blood samples were analyzed. jnear regression analysis showed the following correlations: in a neonatal intensive care unit adjacent to a delivery room caring for mothers per year, (with a referral of mostly for preterm delivery), virtually every neonate <ls g was seen by the same ophtalmologist, j.o. the incidence of retinopathy of prematurity (rop) was in - of ii cases ( preterms <ls g with % surviving to discharge, and % rop in survivors; % if less than weeks). in - , cases of rop were diagnosed by the same ophtalmologist. were born at less than weeks, of which ( %) survived to discharge, and among those survivors, rop was present in ( %), with rates varying from % at weeks to % at . objective: to compare a method of teicoplanine (teico) infusion using an electrical syringe-pump with a manual injection. methods: infusion of teico was simulated through a standart neonatal i.v. system. the infusion system consisted of an life care infusion pump (abbo'it lab.) with its i.v. set for maintenance intravenous fluid (flow < mi/h) connected to a -way stopcock. an meter extension tubing (vygon lab.) was placed between the stopcock and a neonatal catheter, an another meter tubing (injection tubing) was connected to the stopcock. the volume of the injection circuit was . ml. simulations were realised for weights ( or kg), with a doses of mg/kg and a injected volume of , ml to ml. our goal was to perform a complete infusion in minutes. we compare one method of infusion with an electrical syringe-pump with manual methods: a: teico was infused with a syringe-pump pilot c (becton & dickinson lab.) according to a protocol using ) a priming before connecting the syringe to the tubing (for immediate starting of infusion), ) a programmed volume injected in minutes (the drug volume was greater than the dose prescribed to avoid loss of teico into the syringe), ) a ml wash out was performed over minutes with the syringe pump. b: teico was manually injected in a few seconds in the tubing followed by a ml wash out over minutes. c: idem as b but a nu site valve (medex) was connected to the proximal end of injection tubing to avoid leakage between removal of the teico syringe and connection of the wash out syringe. during each run, serial samples were collected every ten minutes over a one hour period. the samples were assessed using hplc method. results: the amount of drug delivred at minutes was calculated and expressed as a perventage of the total amount of teico prescribed. results are a mean of to rons. a b c kg . + , % , + . % , -+ . % kg -+ , % -+ , % . -+ % conclusion: with a drug volume injected < circuit volume and a valve, a direct i.v. administration of the drug in the circuit followed by a wash out seems an accurate alternative to drug infusion conbroled by a syringe pump and is easier to perform. doxapram hydrochloride is a central and respiratory stimulant which is used in neonatal intensive care units to treat caffeine-resistant apnoes of the newborn. routinely, doxapram is administered intravenously. oral administration would be much easier but data on bioavailability after oral administration are limited (two studies) and show a large variation. we therefore studied the oral bioavailability of doxapram in preterm infants. doxapram treatment was started with an intravenous loading dose of , mg/kg during minutes, followed by a continuous infusion of mg/kg/h. after h the intravenous administration was changed to continuous oral administration with the same dosage, blood samples were collected h after both the intravenous and oral administration and analyzed by hplc-assay. apnoea rates per hours were listed before and after starting doxapram treatment. the median apnoea rate per hours declined from to after both the intravenous and oral administration. the oral bioavailability of doxapram is between and %, indicating that the oral route can be used effectively in these infants . there were no differences in the decline of the apnoea rate after intravenous versus oral administration of doxapram, infasurf r has been comnared with exosurf r in two studies: one as drodhvlaxis and the other a rescue trial. similar. althouah non-sianificant benefits were found for the natural surfactant. in trials there was a sianificant reduction in dulmonarv air leaks for . : % ci . - . for babies treated with natural comnared to svnthetic surfactants. for trials ( babies~ comdarina natural and synthetic surfactants for rds { comdarina survanta r and exosurf r. and one infasurf r and exesurfr% neonatal mortalitv was sianificantlv reduced (or . : % ci . - . with natural compared to synthetic surfactant treatment. in two further trials different natural surfactant drenarations were compared. reduced oxvaen needs for hours after treatment were found for r r infasurf and curosurf respectively when each was compared to survanta r. andarent lonaer-term benefits from these surfactants were not statisticallv sianificant. further trials will be needed to ascertain differences between various surfactant preparations. introduction bacterial meningitis is a common cause of admission in the paediatric age group, and one which still carries a high morbidity and mortality. these complications are strongly associated with a delay in antibacterial therapy. a significant proportion of these patients require mechanical ventilation; it is in this particular group of patients that survival is greatly diminished. the identification of risk factors for mechanical ventilation associated with bacterial meninges may help improving survival by shortening the delay to icu referral. the paediatdc risk of mortality score (prism) is the most widely used scoring system in the paediatric icu literature; the accuracy of prism on predicting outcome in meningitis has been shown to be poor. aim we aimed to describe our population of patients with bacterial meningitis requiring ventilation, and then to identify predictors of survival. methods this study was conducted at the baragwanath hospital in south africa which is an universityaffiliated institution with end average annual paediathc admission of patients. baragwanath icu admits non-neonatal paediatdc patients per year. a retrospective chart review from january to december of consecutive paedistdc icu admissions with bacterial meningitis was performed. results approximately cases of bacterial meningitis are admitted to the general paediatric wards every year; this constitutes ± % of all admissions. the mortality of these patients is + %. during this time ( m, f) patients ( %) were accepted for icu admission, with a mortality of . %. the median age was months (range days to years). the median delay to hospital admission was hours, and the median delay to icu admission was a further hours. the median duration of tcu stay was hours ( for non-survivors, for survivors). the main presenting features were convulsions ( %), altered mental state ( %), fever ( %), respiratory symptoms ( %), headache ( %), and diarrhoea and vomiting ( %). the most common indications for icu admission were seizures ( %), coma ( %), shock ( %), respiratory failure ( %) and acidosis ( %). in % of patients there was a cardiorespirstory arrest prior to admission. the most common organisms in the csf was pneumococcus ( %), haemophilus influenza ( %) and e coli ( %). the presence of leucopenia (wcc < ), a low platelat count (< ), acidaemia (ph< . ), and the need for inotropic support were strongly associated with nonsurvival. tachycardia and hypotension were not significantly essorjsted with poor outcome, as has been previously reported. discussion early diagnosis of meningitis and prompt institution of antibiotic therapy requires a high level of clinical suspicion. paediatric patients with bacterial meningitis that require mechanical ventilation have a poor prognosis. there is little evidence to suggest that nomsurviva[ can be predicted prior to icu admission, but a rapid deterioration requiring ventilation and inotrepic support with evidence of severe sepsis (low platelet count, leucopenia) is nmost invariably associated with a fatal outcome. the long term functional outcome of these patients make this disease even more devastating; the rote of icu in the management of these patients has yet to be fully estanished. objective and study design: a retrospective study was pertbrmed for all patients diagnosed with hemorrhagic shock and encephalopathy syndrome (hse) during to . soroka medical center is the only medical facility in the southern negev region of israel serving a population of - , residents, consisting primarily of jews and bedouins. results: patients ( bedouins and jews) were diagnosed with hse. main features on arrival included profound shock and coma with con~alsions. active bleeding and/or disturbing coagulation tests with falling heuroglobin levels and thrombocytopenia was noted in every case. all infants developed diarrhea shortly after arrival. elevated urea, creatinine and liver enzymes was noted in all cases. annual incidence tbr inlhnts < year of age was : , for bedouins and . : , for jews. patients ranged in age from - wks and arrived at the hospital late night/early morning ( : am-ll: am), during winter/early spring (november-april). all were healthy prior to admission, with short prodromal symptoms of upper respiratory tract or gastrointestinal infection noted in cases. most infants had markedly elevated body temperature on arrival. a history of overwrapping and/or excessive hearing was obtained in of infants. bacteriological and virological cultures were negative in all infants. one infant died and neurological sequelae were observed in all survivors. the high prevalence of hyperpyrexia during sleep in the presence of negative microbiological results with no evidence of excessive hearing, and the high incidence of rise among a closed, culturally isolated society known to have a high incidence of congenital malformations, may support previous assumptions that hse results from hyqperpyrexia, originating in most cases from a "physiologic" heat induced trigger which starts and peaks during the night in previously healthy infants with susceptible, predisposing genetic underlie. approximately % of hospitalised infants with respiratory syaacytial virus (rsv) infection require mechanical ventilation. our general practice is not to use specific antiviral therapy. we have undertaken studies of rsv-infected mechanically ventilated patients without underlying congenital heart disease or immunodeficiency. study i (n= ): a retrospective review of infants (mean post-conceptual age weeks; median duration of intubation was days (interquartile range - ). blinded review of chest x-rays during the first three days of mechanical ventilation revealed a spectrmn of lower respiratory tract findings from marked diffiase consolidation in all zones without hyper-inflation (n= ) to gross hyperinfiation without consolidation (n= ), with the remaining patients have an intermediate picture (n= ). death occurred only in patients who were at the poles of this continuum ( / consolidation and / hyperinflation). patients at these poles could be differentiated by gender predominance, birth gestation, alveolar-artarial (a-a) oxygen gradient (p< . l), oxygenation index (p< . l), peak inspiratory pressure (p< . ), and intubation days (p< . ). study ii (n= ): prospective audit of infants with the aim of verifying the above differentiation. separating patients based on their early x-rays and a-a gradient we confirmed the above predictable difference in duration of supportive therapy (consolidation v intermediate: ( - ) v ( - ) p< . ). severe lower respiratory tract rsv-infecrion in young infants results in different distinctive partems of disease with characteristic radiological and clinical features, and each has a predictable timecot~se. conflicting reports on special therapy should be interpreted with respect to these observations before making conclusions about the efficacy of any specific treatment. the clinical data of children wich suffered from a severe mycosis between jan. -dac. and wich were treated with amphotericin b in spanish hospitals were collected. mean age was + . years; were mate ( %) and female ( %). the most common underlying diseases were leukemimymphoma ( %) and congenital heart defects ( %). the major pathogens isolated were candida albieans ( %), other candida ( %) and aspergillus ( %). and liposomal amphotericin (la) in ( %). in the ca group, the starting dose was , ~ , mg/kg, reaching a maximal dose of . + , mg/kg after . + days. maintenance time was + days and the cumulative dose: ~ rag. in the la group, the starting dose was . ± mg/kg, reaching a maximal dose of . _+ . mg/kg (p< . ) after + days. maintenance time was ~ days (p< . ) and the cumulative dose _+ mg (p< . ). the reasons for la use were: elective in % of cases, previous renal failure in %, and ca inefficacy in %. hepatic or renal function impairment was two times more frequent in la group than in ca group, the antifungal efficacy was % for ca and % for la. therapeutic failure or toxicity induced withdraw of ca in % of cases, vs, , % for la (p< , ). mortality was not different in both groups ( vs. %), adverse effects were related to ca in events vs, only one in la group (p< , ). the commonest side effects in ca patients were fever ( %), chilis ( %) and nausea ( %). in ca group, cases ( %) developped analitical anormalities related with amphotericin, vs. cases ( %) in la patients (p< . ). in the first group, the commonest serum alterations were hypokaliemia ( %), raised creatinine ( %) and bilirrubin ( %). in the second one, hypokaliemia presented in % of cases (p< . ), raised bilirrubin in % and creatinine in % (p< , ), the antifungal efficacy of la is at least the same of ca. from the clinical and analitical points of view, la is much less toxic than ca. la can be used at a greater dose than ca and is safe in children with renal failure. laurence desplanques -serge gottot -christian dageville d~artement de sant~ publique -facult~ xavier bichat - rue henri huchard paris -france -the french << reaped ~> network was created to implement a nosecomial infections (ni) quality care program in nicu and picu, the first objective was to describe the annual ni incidence rate in each icu population : all patients stayed more than hours in icu. methods : n] criteria were defined by the reaped group according to cdc criteria. all data were collected by a medical and nursing team. all infection data were validated by an external investigator. results : patients were admitted over a months period. % were newborns. ni were identified among patients. the overall ni incidence rate (ir) was . % and . °/ person day (from . to . °/ according to age, lowest rate for newborns). septicemia ( % of ni) and pneumonia ( % of ni) were the two main ni. according to age, the septicemia ir varied from . to . °/oo catheter day (lowest rate for newborns) and the pneumonia ir from . to . °/ ventilator day (lowest rate for newborns). there were very few other infections (uti : %, ir : . °/ catheter day). gram positive cocci were isolated in % of septicemia ( % of them were coagulase negative staphylococcal). gram negative bacilli were isolated in % of pneumonia ( % of them were pseudomonas). % of ni were caused by candida, mostly septicemia. the septicemia and pneumonia ir varied according to unit even after adjustment for age. discussion the aminoglycoside antibiotics are frequently used in newborns for the treatment of severe infection and sepsis due to gram-negative microorganisms. the currently recommended dosage schedule for tobra ( . mg/kg q h) does not take into account differences in gestational or postnatal age during the first weeks of life. we questioned the validity of these recommendations and studied the population kinetics of tobramycin to establish predictive equations that enables the clinician to select the appropriate initial dosing schedule. methods tobra trough (t= ) and peak values (t= ) were taken on day - after birth in newborns. tobra was administered as a -minute intravenous infusion already in an adapted dosage schedule: . mg/kg q h in infants with gas < weeks; . mg/kg q h in infants with gas between - weeks and . mg/kg q h in infants with gas > wks, tobra concentrations were analyzed by tdx-assay, a one-compartment model was assumed and non-linear mixed effect modelling (using nonmem) was applied to the data, a trough level < mg/l and a peak level between and mg/l was required, with the present dosage scheme % of the trough levels were too high and almost % of the peak levels too low. calculations showed that the following dosage schedule should result in optimal levels of tobra. preterm infants gas < wks: mg q h preterm infants gas - wks: . mg q h preterm infants gas > wks: the currently recommended dosage schedules for toeira result in high trough and low peak levels. prolongation of the dosing interval and increasing the amount of drug per dose according to the above scheme will improve tobra level control. since january british clinicians have been conducting a randomized controlled trial of neonatal ecmo. mature infants (>- weeks gestation and birthweight kg) with severe cardiopulmonary failure have been randomized to receive continued care in their referring institution or referral to a designated ecmo centre for further management. we now present the preliminary results which have prompted closure of recruitment to this trial. the final outcome will be assessed as intact survival against death or severe disability at one year of age for all the recruited patients. patients were categorised by diagnosis such as isolated persistent fetal circulation, secondary persistent pulmonary hypertension of the newborn or congenital diaphragmatic hernia and by severity of illness at the point of first contact with the clinical coordinators of the trial -judged primarily by the oxygenation index ( before randomization). patients were randomized ( in each arm). hospital outcome data are reported for all patients and year outcomes on t ( survivors). at this stage of the babies allocated to ecmo are known to have died compared to of those allocated to conventional management (rr . ; % ci . - . ; p= . ). fewer deaths have been obsea-ved amongst ecmo allocated babies in all the diagnostic categories used. a % incidence of disability and impah~nent has been observed amongst survivors. this rate is similar in both groups and the survival advantage is not offset by an increased rate of disability or impairment following allocation to ecmo. we consider that these data combined with those available from other studies provide conclusive evidence that the survival to discharge from hospital is substantially higher in patients allocated to ecmo than in comparable infants not so allocated. therefore recruitment to this trial has been closed whist awaiting complete one year outcome data. sigston pe, goldman ap. #keating j. crook r. ~e dj~. great ormond street hospital for children nhs trust, and ~biochemistry department, kings college hospital, london, united kingdom. isoflurane is a safe and effective means of long term sedation in both children and adults in the intensive care setting. the use of isoflurane, by adding it to the sweep gas allows the use of this volatile anaesthetic agent in patients on ecmo, enabling rapid control and weaning of sedation. a potential problem with the long term use of isoflurane is fluoride ion accumulation with the possibility of renal toxicity, the purpose of this study was to assess plasma fluoride levels in patients receiving prolonged isoflurane on ecmo. method: fifteen infants and children (aged day - years, median weeks) receiving ecmo support for either cardiac or respiratory failure were recruited to this study. the patients were sedated with isoflurane as well as intravenous agents (morphine and midazolam). isoflurane was administered ( % - %) via a calibrated vaporiser to the sweep gas, adjusting the level to maintain adequate sedation. blood samples were obtained on a daily basis for plasma inorganic fluoride assay. the relationship between plasma fluoride and amount of isoflurane administered, as %-hours (vaporiser setting in % x hours) was calculated by linear regression. results: the duration of ecmo ranged from to (mean ) hours, during which the amount of isoflurane administered varied from to (mean ) %-hours. blood samples were anaiysed, demonstrating individual peak plasma fluoride levels of . to . #mol/ , mean , p.molli (toxic threshold = gruel/f). the plasma fluoride positively co;related with the %-hours of isoflurane (r = . , p = < . ). conclusion: this study shows that although there is a dose related accumulation of inorganic fluoride ions in patients sedated with isoflurane on ecmq, the peak fluoride levels are well below the suggested toxic threshold. merzel y, lev a, bar yosef g, halbertal m, lorber a ecmo center, picu, emek medical center, israel. the mortality rate of pediatric patients with acute myocarditis is - % according to the severity of myocardial damage. a month old gzrl presented with high fever, respiratory and cardiac failure. diagnosis of acute myocarditis was made and the patient was ventilated with high pressures and fio of . . she required high doses of inotropes. echocardiography revealed a dilated la and lv with severe mr. lvedd was mm and lvsf %. calculated oxygenation index was . she was resuscitated after a cardiac arrest. she was commenced on ecmo (using biomedicus centrifugal pump and avecor oxygenator) at a flow of ml/kg/mm with immediate improvement of hemodynamlcs, oxygenation and pc . resptratory assistance and vasoactive drugs were reduced. the patient was transported by air, on ecmo, to the ecmo cevter. she developed arf and cvvh-d was performed. cardiac fimction started to improve after days. ecmo was discontinued on day . echo revealed lvedd mm and lvsf %. ippv was discontinued on day . on discharge, a month later, her lvedd was mm and lvsf %. she behaves normally for age without neurologic or other medical sequellae. literature search revealed no case of acute myocarditis, as severe, that was treated successfully. survavors of disease this severe usually suffer dilated cardiomyopathy and permanent disability. the use of ecmo allows myocardial rest which prevents long term myocardial damage. introduction ecmo is increasingly used in the care of critically ill newborns. despite the frequent use of betalactam antibiotics in the treatment of these infants there are no data available on the dispbsition of cefotaxime (ctx) and amoxicilfin (am) d ring ecmo. the purposes of this study were to determine the pharmacokinetics of these two drugs in infants on ecmo and consequently formulate appropriate dosing regimens. we therefore studied the pharmacokinetics of ctx ( mg/kg ql h) and am ( mg/kg q h) in term infants on day after birth, blood samples were taken before (t-o) and . , , , , (am) and t h (ctx) after the intravenous bolus injection and analyzed by hplc-assays. . ctx mg/kg q h results in adequate serum levels of ctx in fullterm infants on ecmo, am mg/kg q h results in very high serum trough levels. recalculation based on the known volume of distribution and elimination serum half-life of these infants resulted in the following dosage recommendation: mg/kg q h. persistent pulmonary hypertension of the new-born (pphn) is characterised by rapid fluctuations in pulmonary artery pressure (pap) and a clinical impression of stifflungs. lung mechanics were measured in term infants, mean age . +_ . days who were paralysed and ventilated within the first three days of life. fourteen infants had pphn with systemic or suprasystemic pap measured by echocardiography. in these patients, the respiratory system resistance was . % higher (p < . ) and compliance . % lower (p = . ) during systemic or suprasystemic pap compared to when the pulmonary hypertension had resolved. in contrast, there were no changes in resistance in the infants with respiratory distress syndrome (rds) and no pulmonary hypertension or in the seven infants with normal lungs, where two readings were taken hours apart. the changes in lung mechanics interfered with mechanical ventilation, resulting in a . mmhg rise in paco (p= . ) during pulmonary hypertension. inhalation of nitric oxide ppm resulted in a % decrease in respiratory system resistance and an improvement in oxygenation. the bronchial and vascular smooth muscle was increased by % in postmortem lung samples from eight infants with pphn compared to six age matched post-mortem controls with normal lungs (p< . ). these findings suggest a co-constriction and co-hypertrophy of bronchial and vascular smooth muscle during pphn. anatomically the pulmonary vasculature and bronchi lie in close proximity to each other. thus mediators such as endothelin- released locally may act on both vascular and bronchial smooth muscle to produce the observed vasoconstriction, bronchoconstriction and smooth muscle hypertrophy. prince of wales children's hospital university of new south wales, randwick, n.s.w. australia. introduction an increasing mortality in asthmatic children has been reported. the increased severity of asthmatic illness leads to an increased demand for icu admission, and a corresponding increased need for mechanical ventilation. geographic end environmental factors are thought to be partly responsible for differences in disease sevedty throughout the wodd. for this reason, epidemiological studies from diverse areas are important, risk factors for icu admission, and for the institution of mechanical ventilation should be identified, to optimise icu admission criteria and to avoid unnecessary delays in admitting at-risk patients. aim to document the clinical characteristics of ventilated and non-ventilated asthmatic patients admitted to icu. methods this is a retrospective study of all paediatric asthma icu admissions from january to december . results there were patients admitted to the icu for acute severe asthma in the study period. the male:female ratio was : , the mean age . • . months, the mean prism . - . %, and the mean duration of admission . hours. there was no seasonal variation in admissions. only % ( / ) patients required mechanical ventilation. in % of all patients this was the first presentation with asthma. there were some significant differences between ventilated and non-ventilated patients (see table) . there was a significantly higher incidence of concomitant and nosocomial pneumonias in the ventilated patients ( . % vs . %) as well as segmental lung collapse ( . % vs . %). there were no deaths. discussion the need of mechanical ventilation significantly increases the morbidity of and duration of icu stay of asthmatic patients. younger asthmatic paediatdc patients have a significantly higher risk of ventilation. the need for ventilation is predicted principally from a worsening pco and respiratory acidaemia, which is often independently interpreted by the clinician as respira ory exhaustion. this study has shown that icu admission is important in the management of young paediatdc patients with acute severe asthma and respiratgry fa!!ure. intravenous salbutamoi in the emergency, department management of severe asthma in children. g.j.browne,a. perma,x. phung,m.soo westmead hospital, sydney, australia. it is postulate that if an initial intravenous loading dose of salbutamol is given in severe asthma, a more rapid clinical response will occur, reducing requirements for continued high doses of nebulised salbutamoi with fewer side effects. this double blinded study was conducted in the emergency department of westmead hospital a university hospital in sydney, australia. all children with severe asthma had initial nebuliser therapy ( rag of salbutamol with ml of saline). if asthma remained severe minutes later, they were given a dose of intravenous hydrocortisone ( mg/kg) and either normal saline or salbutamol microgm/kg intravenously. frequent nebulised salbutamoi therapy continued during the initial first hour if clinically indicated. continuous respiratory and haemodynamic monitoring occurred in the first hours. serum potassium and glucose determinations were made at study commencement and hour after intravenous therapy. salbutamol determination was made at study commencement. children remained clinically monitored for the next hours, with their ongoing treatment determined by clinical response. children with severe asthma months to years of age were studied, with given intravenous salbutamol and given intravenous saline. the intravenous satbutamol group (ivsg) showed rapid reduction in asthma severity scale in the first hours, with reduced need for high frequency nebuliser therapy ( _< hourly), occurring . hours.earlier. no clinically significant side-effects were found in either group, although, tremor more frequent in the [vsg. biochemistry and salbutamol concentrations were similar in both groups. the use of intravenous salbutamol (i microgm/kg) in the management of severe childhood asthma is a safe and effective therapy with no significant side-effects and the potential to abort severe asthma attacks in the emergency department. intravenous terbutaline in picu piva j., amantra s, rosso a., zambonato s, giugno k, maia t. introduction: the admission to a picu of children with respiratory failure secondary to an acute obstructive lower airway disease is a common event, especially during winter seasons. these diseases have several causes, but most of them (especially asthma and chronic airway disease) have a good response to the administration of b -adrenergic drugs. objective: to find the dosis of intravenous terbutaline that is safe, efficient and with minimal adverse effects when used in children admitted to a picu with acute obstructive lower airway disease and respiratory failure. material and methods: we study the records of all children that were admitted to our picu during the winter of . only the patients that had respiratory failure and acute lower airway disease and who needed the use of iv terbutaline were selected. the records were divided in two groups: less than months and more than a year old these two groups were compared in the following aspects: the minimal and maximal dosis, and the length of time of use of iv terbutaline, frequency of tachycardia, hypokalemia, and mechanical ventilation. to establish any difference in the two groups we use the t exact test of fisher and x , with p< . , results: during the period of study were admitted patients to the picu, and ( , %) of them used of iv terbutaline. the mean age was . + . month, used iv terbutaline during . + . days ( . to days), the initial rate was . + . p~g/kg/min, and the means of therapeutic dosis was . +l. ~g/kg/min (ranged from . to . ). twelve ( . %) patients had tachycardia art obstacle to the increases in the rate of use of iv terbutaline during any time. mechanical ventilation was necessary in patients ( . %) and ( . %) patients died. the children under year of age used initial dosis of iv terbutaline lower than the children up of year old ( . p.g/ kghnin x . ~tg &g/rain, p< . ), but without difference in the length of use, the maximal dosis, the rate of mechanical ventilation and tachycardia. the frequency of hypokalemia was most common in the group of children under year of age. acute respiratory failure during status asthmaticus may require mechanical ventilation. current therapy includes paralysis, pressure control ventilation (pcv) and permissive hypercapnia to limit pulmonary barotranma and its hemodynamic consequences. asthmatic children exert a significant amount of respiratory effort during exhalation. with paralysis, this expiratory effort is lost. unloading the inspiratory work of breathing while maintaining the patient's expiratory eftbrt using pressure support ventilation (psv), may be beneficial. methods: children receiving pcv (peak inspiratory pressure (pip) = kpa. rate breaths/min) and pco > kpa were switched to psv. children were initially ventilated with psv . kpa and peep = . kpa (servo c). all children received beta agonist therapy, ipratropium and anesthesia with ketamine or inhalational anesthesia, and were breathing spontaneously. respiratory parameters and blood gases are shown be~bre psv, within minutes (start) and when the ph had normalized (during). data are presented as median and range, * p < . compared to before psv. results: children with hypercarbia during pcv responded to psv, normalizing pcos and ph within hours. the mean respiratory rate decreased from a median of ( - ) to ( - ) while the pip was decreased to . ( . - . ) kpa within hours. the i:e ratio also significantly decreased. conclusion: psv permitted patients to active/y exhale while unloading the inspiratory work of breathing. perhaps this strategy shifts the patient's respiratory effort from inspiration to exhalation, thus permitting the child to meet the excess work of breathing caused by bronchoconstriction. maged z. youssef, peter silver, laura nimkoff, and mayer sagv. division of pediatric critical care medicine, schneider children's hospital, new hyde park, ny . introduction: mechanical vemiladon of patients with severe bronchospasm can be difficult, due to poor chest compliance and increased airway resistance. ketarmne is a cormnonly used anesthetic agent that has been shown to have bronchodilator properties. the purpose of this study was to determine ifa continuous infusion of ketamine had an effect on the oxygenation and chest compliance of children with severe lironchospasm who were mechanically ventilated. methods: a retrospective chart review was conducted of pediatric patients in severe bronchospasm who were mechanically ventilated in our picu and treated with a continuous ketamine infusion. all patients were receiving aggressive bronchodilator therapy and adequate sedation prior to keramine. patients were excluded if any new bronchodilator or sedative agents were started within hours of initiation of ketamine treatment. all patients were simultaneously treated with benzodiazepines. for each patient, the pao /fio ~ ratio and dynamic compliance [tidal volume/(peak imp. pressure -peep)] was determined immediately prior to ketamine, and at , , and hours post-ketsmine initiation. data are presented as mean ± s.d., and were a~yzed using one way anova and the multiple comparison method of bonferroni. patients (age . ± . yrs.) received * p< . ketamine for severe bronchospastu during mechanical ventilation in our picu. both . .xto-* * the pao /fio ratio and dynamic . . -.... . compliance increased significantly following initiation of the ketamine infusion (see figure) . the mean ketamine dose was ± mcg/kg/min, and the -, mean infusion duration was ± too-[/ hours. one patient required glycopyrrotate ~' to control excessive airway secretions, and " one patient required an additional dose of o--j i ~-~ ~/me diazepam to control hallucinations after i cessation of ketamine. all patients were t~n~,mr~ *~am~ successfully weaned off mechanical ~l~s ~,~s~on ventilation and discharged from the picu. conclusion: continuous ketamine infusion to mechanically ventilated pediatric patients with refractory broncliospasm results in a significant improvement in oxygenation and dynamic compliance of the chest. reports of adults with status nsthraaticus document significant morbidity and mortality, whereas studies in children have had more varied results. different centers report mechanical ventilation (mv) in to % of admissions, occurrence of pneumothoraces or paeutuomediastinums in to %, and mortality in up to % of patients ~'t . we retrospectively reviewed status asthmaticus admissions to the pediatric intensive care unit (picu) between january and december . seventy-five of these patients were admitted fr~an the emergency department of chla (er admit). the mean length of stay in the picu was . days and the mean length of stay in the hospital was . days. based on patients who had arterial blood analyses, patients had hyperoapnia (pco > ). all patients received oxygen, inhaled albuterol (alb), and cortieosteroid therapy. ninety-five percent of patients also received methylxanthine (mx) therapy. of the admissions, patients ( %) required mv. only of these patients were admitted through our emergency department, whereas the remaining patients were intuhated at outside facilities. twenty-three cases required intr:wenous beta-agonist therapy, either isoproterenol osop) or terbutaline (terb). h~ff of the ea.~es re~%wed were complicated with hypokalemia (k+< . ). c,', ,~lications ofpoeumothoraces or pneumomediastinums were seen in % of ,'r:u~ported patients, but in only % of er admit patients. only % of these were in mechanic.all, )atients. there were no deaths in the review. respiratory mechanics measurements 'are useful in mechanically ventilated children to optimize ventilator settings. nevertheless, the transducers used to measure flow (f) and pressure (p) remain expensive. objective. to evaluate the performances of piezoelectric p transducers ( us dollar) in measuring f and p. methods. we used a previously described monitoring system measuring respiratory parameters [ ] . in this study f was obtained by a differential piezoelectric p transducer (_+ . cmi-i , honeywell) whose sensitivity has been reduced to +_ cmh by an electronic amplification equipment and p by a piezoelectric p transducer (_+ (). cmhzo, honeywell) connected to a grid pneumotachymeter &nt) ffleisch or ). volume (v) ( to ml) obtained by numeric integration off ( . to l/rnin ) and p ( to cmh ) were respectively delivered through a calibrated seringe and an electronical manometer (pic premier) and calculated by the computer. bland and altman analysis was used for assessment of results bias. coefficient of repeatability (cr) was estimated by the standard deviation of repeated measurements of the parameters as calculated in a oneway analysis of variance. results. mean difference (mdi between injected v ( to ml) and measured v using pnt was . ml, sd = . ml. difference and mean v were not correlated. sd of repeated v measurements were not correlated to v. cr was . ml. mdif between injected v ( to ml) and measured v using pnt was lrd, sd = ml sd of repeated v measurements were not correlated to mean v. cr was ml. mdif between injected p and measured p was . cmi-i , sd . cm h sd of repeated p measurements were not correlated to mean p. cr was . cmh . conclusion. inexpensive piezoelectrical transducers can be used to measure f and p and evaluate respiratory mechanics in ventilated children. previous studies have already shown the problem of the reproducibility of pft in preterm ventilated babies. were studied preterm ventilated babies {mean weight gr) in the first week of life in clinically stable condition, measuring flow, airway pressure and esophageal pressure simultaneously. each baby was studied twice with an interval of one hour and each study was done increasing the rate till to inhibit spontaneous breaths. none sedative has been used. only mechanical breaths were analyzed. compliance and resistence were calculated with a computer system using the linear regression method. we expressed quantitatively the intrapatient variability as the percentage of variation of tidal volume, compliance and resistence between the two studies in each baby. then intraclass correlation coefficient test (icc) was applied to confirm qualitatively our results (total agreement = , good reproducibjtity > . ). we h~£ed, an a eept~ble ~efiabirl¢, ~-~r;= '~ . during mechanical ventilation, an air leak (al) and plateau phase duration (pl) may influence dynamic and static compliance (cdy and cst, respectively). this study evaluated the effect of al and pl on two methods of measuring c.dy and est. methods. intubated, ventilated patients in a pediatric intensive care unit were evaluated after obtaining informed consent. patients were intuhated with a cuffed endotracheal tube and ventilated with a serve ( ventilator. cdy and cst were determined using the serve ands~rmedics . objective: evaluate the repercussion in respiratory mechanics and arterial blood gases and the impact of the ventilator adjustments on the auto-peep magnitude. material and methods: the measurement of the auto-peep was performed using an eletronic-pneumatic controlled device with a oclasion valve installed between endotracheal canutla and the ventilator circuit. the d~'ice was connected to a solenoid to detecte the end of inspiratuo phase and thus, the activation of the oclusion valve. the signs of pressure and flow were monitorized using a diferential transducer and it was processed using a pc computer and tmeumoview® software. the stud were divided in phases: phase a. where the ventilator adjustments was performed using the routine of the unit and phase b, where the targets of mechanical ventilation were to minimize the auto-peep. static compliance (crs) was ineasured by the single-breath occlusion technique, using a mean of ten occlusions for analysis. passive respiratory resistance measurements and the tidal breathing flow-volume loops were also obtained., while the ventilatory settings were siguificantly reduced soon atier ecmo was started. before ecmo crs measured in all patienls was . _+t). ml/cmh /kg (mean_+sem). for each patient the ecmo course was divided into four periods, proportional to the duration of the treatment, and the best ~alue of crs in each period was chosen for analysis. as shown on the figure. crs significantly improved (*p< , ) from the second half of the ecmo course in the group of patient that finally were successfidly weaned from ecmo. no change ill compliance was measured in the group of patients who failed to respond to the extracorporeal hmg support our data suggest that compliance measurements during ecmo can be useful togelher with overall clinical evaluation to predict both outcome and duration of cxtracorporeai support in the neonatal and pediatric population. objectives: brain temperature determines the amount of neuronal damage caused by hypoxic insults. thus measuring brain temperature at standardised conditions is in request. we investigated whether brain temperature of neonates varies with head insulation environmental temperature, body activity and time course. patients and methods: we investigated non-invasive brain temperature analogues in healthy prematures tess than two weeks of age in an incubator (gestational age . + . wks; x + sd, weight + g). we measured nasopharyngeal temperature (tnasoph) by a thermistor placed in the nasopharynx via a feeding tube, zero-heatflux temperature (zht) at the temple by a thermistor and healflux transducer, insulated by two pads, as well as rectal and incubator temperatures. patient activity was documented by video taping. measurements were performed during periods of increased insulation ) by turning the head with its measuring site on to the mattress ( ( ) ( ) - ( ) ( ) ( ) ( ) . ( ) ( ) { ) ( ) ( ) - ( ) ( ) ( )i ( ) ( ) ( ) ( ) . ( ) ( ) t ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) web (lmg/kg) at rain ( ) - ( ) ( ) ( ) ( ) - ( ) the vehicle had no effect. paf caused dose dependent rise in ao and pa pressure and reduction in flow to lpa (up to % like the vascular endothelium, the endocardial endothelium (ee) has a significant impact on adjacent myocytes, and may critically alter myocardial function.~ we have previously shown that ee cells are capable of sensing and responding to hypoxia by the release of prostacyclin (pgl). potassium channels in other cell types have been reported to be oxygen sensitive. to determine whether potassium channels modulate the ee hypoxic response, we investigated the effects of three potassium channel inhibitors on hypoxia-induced pg] release from ee cells. methods: ovine endothelial cells were harvested and passaged onto ,~ microcarriers. cells were constantly perfused with normoxic and hypoxic kreb's solution, and with three potassium channel blockers: glibenclamide (gb, #g/ml), tetraethyl-antmonium (tea, ram) and aminopyridine ( ap, i mm), perfusate was assayed for prostacyclin (ria). data were compared by analysis of variance. * p<. compared to normoxic control; # p< . compared to hypoxic control. adrenaline is extensively used for resuscitation in neonates with rds. however, effects of adrenaline on systemic, pulmonary and cerebral hemodynamics have not been defined in newborns with rds. thirteen anesthetized, and ventilated newborn piglets were subjected to repeated saline lung-lavage series while mean systemic arterial pressure (abp), mean pulmonary arteriat pressure (pap), mean left atrial pressure (lap) and mean central venous pressure (cvp), cardiac output and blood flow in the internal carotid artery (ica) were measured. systemic vascular resistance (s~), pulmonary vascular resistance (pvr) and cardiac index (ci) were calculated. sixty minutes after luug-lavage, the adrenaline group (a) (n= ) received adrenaline as a continuous infusion of . lag/kg/mi, while the control group (c) (n= ) received saline. none of the varlables were changed by saline. however, significant increases in abp (p< . ), pap (p< . ), ci (p< . ) and svr (p< . ) were observed after administration of adrenaline, whiie pvr and ica were not modified. mean±sd for abp/pap (p/a), fvr/svr (p/s) and ci (ml/mirdkg) were: ratios of pap/abp and pvpjsvr significantly increased following infusion of adrenaline. these data suggest: ) the cerebral perfusion is preserved during the infusion of adrenaline; ) effect of the adrenaline infusion on the systemic circulation is more pronounced than its effect on the pulmonary circulation in newborn piglets with surfactant deficiency. s demirak~a, ch knothe, kj hagel, j bauer department of pediatrics, justus-liebig-university giessen, frg inhaled no is a short acting selective pulmonary vasodilator. we studied the effects of ppm no and % oxygen during heart catheterization in children (age - years, median years) with heart defects and elevated pulmonary vascular resistance index (pvri) in order to asses the value of no as a tool of decision making for corrective cardiac surgery. patients were eligible for testing when they were more than one year old and had a pathologically elevated pvri in a previous heart catheterization. intubation, 'anesthesia and muscle paralysis were performed in all patients during testing of pulmonary reagibility. calculations of pulmonary vascular resistance and flow were based on the fick method. response to no was assumed when pvri declined more than %, of the patients were responders to no. effects of no and oxygen on pvri, mean pulmonary arterial pressure (mpap) and pulmonary vascular flow (qp) in all responders are described in the table below. cardiac surgery was offered to all responders, and of them were successfully operated. surgery is planned in another patients and parental consent for surgery was not given in one patient. in ebstein disease, during the first days of life, the ability of right ventricle to propel blood to the pulmonary artery is impaired due to high pulmonary vascular resistances. the flow is mainly directed to left atrium through tricuspid insufficiency, right atrium and foramen ovale. to decrease pulmonary resistances and increase pulmonary blood flow, high frequency oscillations, mechanical ventilation, nitric oxide and prostaglandin are required. after few days, a forward circulation is normally established. we cared two newborns with ebstein disease where this approach was hindered by a large pulmonary valve insufficiency. both of them were diagnosed in utero, showing a large tricuspid insufficiency with a non opened pulmonary valve and a ductal left to right shunt. one fetus was hydropic. at birth, blood stream from the ductus arteriosus was directed to the right ventricle through the pulmonary valve insufficiency then to right atrium, left atrium and ventricle, aorta and ductus arteriosus. a low pulmonary blood flow was demonstrated by low mean velocities ( cm/sec). a high reverse flow was seen in descending aorta with a negative flow in the renal artery. both of these newborns were oliguric because of ductus arteriosus steal. pulmonary blood flow doppler evaluation allowed different strategies of ventilation, switching between hfo and conventional ventilation, modulation of pge doses, inhaled pulmonary vasodilators (nitric oxide) and surfactant. the hydropic baby died, the other survived after weeks of intensive care complicated by supraventricular arythmia (wpw). in conclusion, during neonatal period, in ebstein disease, a large pulmonary insufficiency leads to a vicious circle where lungs are excluded, inducing severe asphyxia and high pulmonary resistances. the blood is backward propeled from the aorta through the ductus arteriosus to the right ventricle and atria, then left cavities to aorta. arec must be considered when pulmonary blood flow does not increase despite optimal therapy. guti~rrez-larraya f*, mandoza a*, velasco jm*, zavaneua ( **, gatindo a ~, s&nchez-andrede r, s&nchez jl***, mellon a***, mar f***. pediatric cardiology*, pediatric cardiac surgery**, pediatric intensive care unit***. hospital de octubre. madrid. background: transesophageal pacing (tp) is effective and sate both for diagnosis and treatment of pediatric arrhythmias. material and methods. eleven consecutive patients are included. a tri or quaddpolar or f temporal transvenous catheter with an interpolar distance of to mm was advanced through the nares and positioned to the point with the largest amplitude of atrial deflection, surface ecg and a bi or monopolar electregram were recorded simultaneously, selecting filters when needed ( to mhz). pacing was performed with a programmable stimulator (medtronic ) beginning with ms and increasing ma to and then increasing up to . ms. narula method was selected to diagnose sinusal node disfunction (snd) and overdrive pacing to treat tachyarrhythmias. results. tp was useful in all the patients and no complications were observed: in patients a snd was diagnosed (one needing a definitive pacemaker), in two patients with atrial ratter (ripe ) sinus rhythm was recovered, in one patient with a postoperative junctional ectopic tachycadia we were able to get atrial synchrony with marked bemodinamic improvement, and patients with paroxysmal supraventricular tachycardia sinus rhythm was easily and quickly restored ( of them recquirad repited episodes of tp until pharmacelogycal levels of antiarrhythmic drugs were raised). mean age and weight were months and . kg (one patient had . kg). there was a close relation between height and depht insertion (r= . ). mean stimulation parameters were , ms and . ma. discussion. in experiencied hands tp is an effective and safe way to treat and diagnose cardiac arrhythmias even in newborns. it should be tried before endovenous pacing is stablished and it is faster than pharmacologycal treatment. bailing g., eicken a., sebening w., vogt m., schumacher g., bl~hlmeyer k.; kinderkardiologie, deutsches herzzentrum m nchen, germany to assess the outcome of balloon valvuloplasty in infants with cardiac failure caused by critical aortic stenosis a retrospective study was performed. between and neonates, aged - days (median d), weight .t - , kg (median , kg) with critical valvar aortic stenosis were dilated by balloon (aovp) as the first line treatment. patients received prostaglandin el, needed inotropic drugs and mechanical ventilation. associated cardiac lesions : persistent ductus arteriosus (pda) in patients (restrictive pda in cases), a mitral regurgitation (mivr) in cases ( severe and moderate or mild mivr), angiographic findings of endocardial fibroelastosis (efe) in patients, mitral stenosis (mivs) in , coarctation of the aorta (coa) in , and finally a small musculary ventricular septum defect (vsd) in i patient. vascular approach for ballooning : a. axitfaris in cases ( %) a. femoralis in t ( %) and v. femoralis in cases ( %). the median ratio between inflated balloon and aortic valve diameter was , . dilatation was achieved in all cases. the peak systolic gradient across the aortic valve (pre aovp) ranged from to mmhg (median mmhg) and was reduced to to mmhg (median ; gradient reduction is significant (p < , )). aortic regurgitation (aovr) was absent or mild in , moderate in and severe in patient after aovp. children survived (actual suwival rate: %; early mortalffy: n = ; late mortality: n = ). mid term follow up ( - , years; mean , years) showed an increase of the systolic peak doppler gradient across the aortic valve (median mmhg) but no increase of aovr. re-interventions (re-aovp: n = , commissurotomy: n = , mitral valve replacement n = , resection of subaortic stenosis: n = , resection of coarctation: n = ,vsd-closura: n = ) were performed in patients. rv contractility and pulmonary vascular mechanics(pvm) in immature animal models are poorly underslood. we developed an acute rv injury model to measure rv contractility and pvm in response to commonly used cateehalamines. ten anesthetized piglets ( - kg) were instrumented with micromanometers in the lv, rv, pa, and la. a pulmonary artery flow probe was placed to measure cardiac output(qpa). ultrasonic dimension crystals were sutured to the myocardium and dynamic chamber volumes estimated using shell subtraction methodology. rv injury was induced with - cryoprobe injuries at - to - °c for - minmes each. da at mg/kg/min, db at mg/kg/min, and ep at . mg/kg/min were infused in random order. rv contractility was evaluated by calculating a load independent measure of contractility, the preload recmitable stroke work(prsw), during vena caval occlusions. to describe pvm, input resistances), characteristic impedance(z ), total pewer(tp), and efficieacy f=qimo"p) were measured. measurements were made pre-and post-injury, during infusions, and between infusions. clyoablation decreased prsw ( . _+ . to . + . , p< . ). at the end of the experiment, prsw remained depressed to this level indicating stability of the model. one factor contributing to organ dysfunction for infants undergoing repair of congenital heart defects (chd) is their "inflammatory response" to cardiopulmonary bypass (cpb). this response is characterized by an increase in cytokine release, complement activation and endothelial injury. modified ultrafiltration (muf) is a method for removing tissue water and inflammatory mediators by rapid ultrafiltration followin~ cpb, muf may acutely improve post-operative end organ function. in this study, we evaluated the effects of muf on the pulmonary and cerebral function of infants undergoing cpb for repair of chd. we prosnecrivety randomized infants (.~ mos) to either muf (n= ) or no muf (n= )(control) following correction for chd. the study intervals were ) before cpb, ) immediately after cpb, and ) minutes after cpb. pulmonary function was evaluated by measuring dynamic compliance (cdyn) and airway resistance (raw). for pts (mue= pts; control= pts) exposed to a period of deep hypothermie circulatory arrest (dhca), cerebral metabolism (cmro ) was calculated at each interval using the xe clearance technique for cerebral blood flow measurements and arterial and jugular bulb saturation measurements to calculate cmro . a reduction in cmro has been consistently demonstrated after dhca. the effects of muf on cdyn and on cmro are shown below: p< . vs pre-cpb; # p< . vs post-cpb • p--o. vs. post-cpb this study demonstrates that immediately following exposure to cpb, muf will improve pulmonary compliance. raw was not different between groups. there was no significant difference in hours of post-op ventilation for either group. in those pts exposed to dhca a trend towards better cerebral metabolic recovery compared to control was demonstrated. this is the first technique applied to infants undergoing dhca where cmro after cpb was greater than precpb measm~s. although this may be beneficial to postoperative hemodynamics, ventilatory management and long-term neurologic recovery, more patients and longer follow up will be necessary to verify such an effect. the effects of conventional mechanical ventilation (cmv) on left ventricular (lv). diastolic filling in neonates are not well established. one approach to improve lv filling is the use of cmv to provide a phasic increase in airway pressure {thoracic augmentation). this phasic increase in airway pressure may result in an increase in lv filling similar to that which occurs with cpr. thoracic augmentation has not been evaluated in neonates with ventricular dysfunction who frequently demonstrate increased heart rates. attempts to maintain low peak airway pressures during cmv may result in a prolonged inspiratory time that occurs over multiple cardiac cycles. this may alter lv filling in the later cardiac cycles. to determine the effects of inspiratory time on lv diastolic filling, infants were examined with doppler echocardiography less than hrs after surgery for the arterial switch procedtme. pulsed doppler recordings of the millal valve (mv) were obtained with the inspiratory time adjusted to occur over cardiac cycles ( sec.). a pressure transducer was placed in line with the ventilator, and the respiratory cycle was recorded superimposed on the doppler tracing to provide accurate determination of inspiration and expiration. doppler recordings were obtained from the apical -chamber view and the following measurements were made: peak e and peak a velocities, eia ratio, and deceleration time. compared to the expiratory phase of cmv, the initial beat during the iuspiratory phase of cmv resulted in an increase in mv peak e (. +-. vs . -+ . m/s, p< . ) and peak a (. + . vs . -+ . m/s, p< . ) velocities with no change in mv deceleration times (p<. ). compared to the initial beat during tile inspiratory phase, the third beat during the inspiratory phase resulted in decreased peak e (. + . vs . + . m/s, p< . ) and peak a (. + . vs . + . m/s, p< . ) velocities with no difference in deceleration times. thus, cmv augments lv filling during the initial phase of inspiration. however, as the increase in airway pressure is distributed over multiple cardiac cycles, lv filling falls below baseline levels. these observations indicate that while thoracic augmentation may be beneficial, to optimize lv filling the inspiratory time of cmv must be < cardiac cycles. energy expenditure in pediatric orthotopic liver tranaplantat~on, to determine the actual calorie requirements of critically ill children and evniuate the correlations between measured, stress-p~lictod and repleted energy exponditttm and the severity of illness. des/gn: a prospective, dinlcal study. se~ng: tertiary care pediatric icu in a university hospital. patients: ten patients aged to months with disorders prompting picu admission, including sepsis, respiratory failure, solid organ transplantation, and cardiovascular surgery. inta~entions: all patients were studied within hrs of major surgery or transplantation, or following acute illness. all patienls were severely stressed clinically and all but two were intubated by cuffed tubes, in three of them, still in a stress state, the study repeated on the third day of the disease, energy expenditure mensurements (mee), as well as illness seventy scoring systems, mtfltisystern organ failure scores and various anthropemetric and clinical indices of nutritional status, the stress-predicted energy expenditure (s-pee), the basal metabufie rote (pbmr), the repleted energy (re) and the recommended dietary allowances (rda) were measured or calculated in each patient. multiple regression analysis was used to analyze the data. measurements and main results: although the mean mee was significantly lower than the mean s-pee ( . + kcal/kg/day vs. . : kcal/kg/day, p<. ), it did not differ significantly from the pbmr (mean difference - . kcal/kg/day, range - . to + . kcal/kg/day). the s-pee/mee ratio ranged from . to . , while the re/rda ratio ( . : kcal/kg/day)/( . : kcal/kg/dny) ranged from only . to . . the prism/tiss ratio was not correlated better with mee than the diagnostic category (r~=. vs.. , respectively). the re was positively correlated withthe mee (rz=. , i)=. ) while negative oarrelatian has been found between mee and age, mid-arm circumference, triceps skinfotd and the use of vaseactive agents (r~. , - , -. , p<. and -. resp~lively). concl.m~: if s-pee is used for caloric repletion in the stressed oritic~ly fll el~d, these patients will be substantially overfed by as much as %. although pbmr appears to approximate the mee by ± %, other clinical and nutritional indices should also be ennsidered. objective: to deter .mine..t.he metabpli.c and.nutritional state of mechanically ventilated intants and children m relatmn wlm severity or msease. patients and methods: mechanically ventilated infants and children, median age months (range days to years), were studied. severity of illness was assessed using prism, prism-ii~ and fiss-scores. oxygen consumption (vo ), energy expenditure (mee) and respiratory quotient (rq) were determmed by mdirect calorimetry. total urinary nitroger(tun) and creatinine excretion, levels of albumin and crp were aetermmed in patients. in these patients daily caloric intake and substrate utilization were assessed. they were categorized in subgroups: a partial feeding (recent admission to p cu); b complete feeding. results: mee of the total group (n= ) a) i=intake g/kg/day (% total intake); u=utilization g/kg/day (% total production). nitrogenba]ance was negative in all patients in group a (mean - . -- : mffkg/day) and positive in all but one patient in group b (.mean . ± .d n~g/..kg/day;p= . ). no significant correlations were round between creatinine height index, crp, albumine, jun vs v u /kg conclusions: the mean measured energy expenditure does not exceed predicted resting energy expenditure, but ~ere is a wide range. in a majority ot patients with complete feeding h.igh carbohydrate intake resulted, in high kq and lipogenesis. in patients witla partial teeding the highly negatwe nitrogen'balance suggests that in the early phase of diseasean higher protein intake should be provided. severity of illness scores ann oiocnemicm markers of physiologic stress correlatedpoorly with oxygen consumption. leite,hp; iglesias, s; faria, c; ikeda, a; albuquerque, mp; carvalho, wb pediatric icu -s~o paulo federal university -s~o paulo, brazil objectives: ) to evaluate patterns of use and monitoring of nutritional support in critically ill children; ) to evaluate an education program in nutrition support given throughout the resident physician training in the pediatric icu. patients and methods: records of patients receiving nutritional support during were reviewed. aider this first phase, knowledge and understanding of the role of nutrition support was conveyed to the residents through didactic lectures. in a second phase thedata were reevaluated in children who were given nutrition support in . results: from a total of days ofthempy, the single parenteral route was utilized in , %, the digestive route (tube feeding or oral route) in , %. of this time. a previous nutr~ional assessment was performed in children; no patient had the nutr~on goals set. the nitrogen to nonprotein calories ratio ranged among : and : . only , % of the patients had their estimated caloric needs supplied and this goal was achieved only in those patients who were on enteral tube feeding. patients did not achieved their goals for vitamins. the supply ofoligonleme~s was adequate except the zinc. nutritional monitoring parameters including weight, serum albumin and serum triglycerides were performed in almost all the patients but without uniformity. the reevaluation ofthase parameters showed adequacy of protein and micronutrients supply; however deficiency in nutritional monitoring and infrequent enteral feeding were still detected. conclusion: there were lacks in the implementation of nutritional support, which were partially corrected in the rid phase of the study, although the training of residents may have contributed to give them cognitive skills, it didn't changed policies and procedures as desired. we recommend reinforcement of the education program concerning basic nutritional aspects, and the organization ofa multidisciplinary team in charge of coordinating the providing of nutritional support. plasme free fatty acids (ffa) are the meier energy source for mast tissues. during fasting ffa are released from the breakdown af triglycefides in edipose lissue (at). lipalysis, le. the rote of release o/ ffa, has been megsured in humans by means of stable isotope techniques using labeled pa or glyeerd as traces. no information is avoilob!e io dale on the ro of la. we infused albumin hound u c-pa and u c-la in critically ill infants, receiving kcel/kg/doy of iv glucose end na oral feeding (weight . ,i., kg;, range . - . ; ego : days, range ) and measured simultaneously the ra of pa and la from (he isotopic enrichment of plasma fea by gas chromatography-mass speclrome|ry ai : , : and : hours from tile shod of the infusion. a subcutaneous gluted at biopsy was obtained far fatty acid (fa) composition. we intended to ( ) in fie infants sbjdied atipa ~'os hi her than attla (~p<o.o )' and ihe ra of pa was higher then thai of la (~p= . §). however the ratio el the re's to the respeclive fa in at was higher for la than far pa (*p= . ). in conclusion the ra af la was higher then expected from ~he fa composition of at. we speculate [hat la is preferentially released during tipolysis and its contributuion to the energy metnbolism of the sick infonl could io !orger than what: is normai!y assumed from ihe fa composilion of at and of plasma fea. preoperative starvation can be prevented in infants undergoing non gi surgery by continuing feeds till as long as it is safe to do so. some gi disorders require withholding of feeds for a long period. for these and for infants already suffering from pem~ parenteral feeds are given preoperatively. peroperatively nutrition is maintained by ensuring adequate glucose supply in the infusion fluid and its smooth metabolism. proper placement of feeding tubes are also carried out peroperatively. postoperative early resumption of oral feeds is ensured by new techniques of anaesthesia and pain relief and methods to ensure early return of peristalsis. enteral feeds of pre digested substances and feeds through transanastomotic tubes and through gastrostomies and jejunostomies can prevent starvation in many situations. parenteral feeding is carried out only when enteral feeding is not possible, for example in necrotising enterocolitis~ gastroschisis, short gut syndrome, high enteric fistula and acute pancreatltls. the artedal ketone body ratio (akbr) is established as a valuable tool in the management of adult patients undergoing surgery for severe liver disease.the redox theory emphasises the central role of the liver in preserving homeostasis and the importance of the hepatic mitochonddal redox potential (nad+/nadh) in monitoring liver function and integrity.the akbr (plasma acetoacetate/ hydroxybutyrate) is a measure of hepatic mitochonddal redox status, reduced ratios associated with poorer outcome. the venous ketone body ratio (vkbr) is influenced by peripheral ketone utilisation and therefore thought unreliable in assessing hepatic redox its use has been dismissed as no correlation with the ratio in hepatic venous blood was found. inspite of this. vkbr is used in the diagnosis of lactic acidoses and respiratory chain defects. the objective of this study was to determine the relationship between the akbr and vkbr in a paediatnc intensive care population. children admitted to the paediatdc intensive care unit, with indwelling arterial and central venous lines as part of their routine care, were recruited for the study. the median (range) age was . years ( - . years) with prism score ( - ), simultaneous akbr and vkbr were measured. suppression of ketogenesis was confirmed using a rapid -hydroxybutyrate strip test (ketofilm, genzyme diagnostics) pdor to sampling. the median vkbr . (range . - . ) was lower than median akbr . (range . - , ), there was good agreement between artedal and venous ratios, bland-altman analysis giving a % confidence interval for relative bias of - . to - . . there was a significant correlation between vkbr and akbr r , , p, . (intercept . , slope , ) conclusions; values for akbr tend to be higher than vkbr, however there is a consistent relationship in critically ill children. in paediatdc patients in whom ketogenesis is suppressed, vkbr may be considered equivalent to akbr. this simple test may prove to be a useful adjunct in predicting mortality in cdt{cally ill children. to understand the present status of pediatric intensive care in china, we conducted a survey between october and december, , involving hospitals in provinces and municipalities. the results showed that there were totally icus for pediatric patients, including pediatric icus (picu), neonatal icus (nicu) and pediatric surgical icus (sicu), with total of beds. the physicians to bed and nurses to bed ratios were : . and : . respectively. the average number of equipment per bed was . ventilator, . multi-fnnction monitor and . infusion pump. very few of the icus had portable x-ray machine, biochemical and blood gas analyzers, and hemodialysis machines. the most frequently treated diseases/conditions were pneumonia, intracraniat infections, post-operation and sepsis in picus, and neonatal pneumonia, hypoxic ischemic encephalopathy and sclerema in nicus. pneumonia and respiratory failure accounted for . % and . % of all the diseases/conditions treated in the icus and the mean case fatality rate of respiratory failure was . %. the results of the survey suggest that there is shortage of tcu beds and modern equipment, and treatment is often delayed due to excessively strict criteria for mechanical ventilation. a set of simple, and nationally acceptable criteria for evaluation of severity and cure of the diseases/conditions is urgently required. the medical reports of children were reviewed and each admission was analysed in terms of age, sex, diagnosis, management within the hosp~al, length of hospital stay and type of poisoning. the youngest age was days and the oldest was years. hundred and twelve ( . %) of the children were girls and ( . %) were boys. the most common ( . %) reason of admission was intoxication among all of the cases and the greatest group ( . %) of the poisoned children had ingested medication which was followed by another group of patients ( . %) who had eaten mushrooms. the peak incidence of drug ingestion was salicylate intoxication ( %). forty drug poisoninigs were accidental while was intentional. the majority ( . %) of the cases that committed suicide was between and t years old, and the main cause of suicide was being unsuccessful at school. we conclude that poisoning -especially salicylate intoxication -is still a major problem in turkey and we believe that emphasis on the need to stere all kinds of drugs in a secure place and re-examination of a chin resistant packaging should help to reduce childhood poisoning significantly which is a preventable condition. included were all patients who died in the hospital, except those treated in the neonatal icu (predominantly premature and sga newborns) and those who died in the emergency room. among a total of hospital admissions (excluding the neonatal icu and emergency room), patients died ( . %). of these patients ( %) died in the pediatric icu, ( %) in the ward and ( %) in the operating room. a chronic underlying disease was present in ( %). withholding or withdrawal of therapy was implemented in ( %) children, ( %) died due to failed cpr, ( %) were brain dead and ( %) died following a dnr order. justification for therapeutic restrictions in the patients of the dnr and w/w groups was imminent death in ( %), lack of future relational potential in ( %) and excessive health burden in ( %). withdrawal or withholding of therapy was carried out by extubation in %, vasoactive drugs were stopped in %, and mechanical ventilation was withdrawn in %, analgetics and sedatives were frequently used (in % and %, respectively). hence decisions concerning restrictions of treatment are common in pediatric practice, mostly due to imminent death. patients in which treatment was restricted were characterised by a longer hospital stay, a worse prognosis, a higher frequency of chronic underlying diseases and a lower acute mortality risk. despite restrictions of intensive therapy, most patients were allowed to die in the pediatric icu. background microalbuminuria (mca), a subclinical increase in urinary albumin, reflects glomerular and overall vascular permeability " . an increase in urinary excretion of albumin occurs after burns and trauma. transcanillary albumin escape rate is also increased in response to elective surgery and in critically ill adult patients . we investigated a possible relationship between urinary albumin levels and clinical instability, as measured by pediatric risk of mortality (prism) scores. method we studied consecutive patients (median age of months, range - ). patients whith nephropathies or any abnormality of urine analysis were excluded from the analysis. prism scores, mca (mg. - ) (immunonepbelometric) and urinary creatinine (cu) (mmol.l -!) (jaffb) ( hour collection sample) were determined within hours from admission to the picu. the mca/cu ratio (mg.mmol. "l) was used to correct for urine output variability. diagnoses included respiratory failure ( ), postoperative ( ), neurologic ( ), sepsis ( ), trauma ( ) and miscellaneous cases ( ). pearson's correlation was performed to correlate data. results and conclusions. mean prism score and mca/cu were . ± . sd and - sd, respectively. a significant correlation was found between mca/cu and prism scores (r= . , p< . ). our observations show that, independently of the initial insult, the paediatric unstable patient may have increased capillary permeability, which is correlated with the degree of physiological derangement, as measured by prism scores. microalbuminuria can be rapidly determined since it is routinely used in the management of diabetic patients, it is inexpensive, simple to measure and blood-spearing. therefore, it might have a role in the clinical assessment of capillary permeability and transcapillary albumin escape worldwide the demand for paediatric intensive care services exceeds the supply. in developing countries sporadic access to such services alters expectation of care and can lead to children being either mechanically or handventilated in general paediatric wards. the outcome of children requiring ventilation in a major teaching hospital in india was reviewed.children were ventilated on an adult intensive care unit (aicu) if a bed was available, otherwise in the general paediatric wards. over a year period children were ventilated on aicu with deaths. yearly mortality rates varied between - %. over a month period patients were ventilated on the paediatric wards. of the p~tients over weeks of age died (chi squared .t >p> . ) reasons for the higher mortality rate on the paediatric ward likely include the higher patient:nurse ratio, and more limited resources. a predictor of mortality based on simple physiological observations without the need for expensive blood tests and including chronic health status would be a useful tool. the establishment of a paediatric intensive care unit is proposed to redress the balance of care. to assess the performance of the pediatric intensive care unit of hospital dona estef~nia by an international standard score, the authors did a prospective study of consecutive admissions to the unit during a period of months. mean age was . _+ . months; mean lengh of stay was . + . days. the effectiveness and efficiency were determined by the admission prism. admission efficiency was defined by two criteria: a) mortality risk > % or b) the administration of at least one intensive care unit-dependent therapy. the cumulative observed mortality was . % and the expected mortality was . %, with a standardized mortality ratio (smr) = . . the overall performance of the prism score-based predictive model was found to be good (goodness-of-fit test x [ ] = . ;p= . ). of patients admitted, combining the two criteria (icudependent therapy and mortality risk) an admission efficiency of ( . %) was found, equating to ( . %) of cu days. conclusion: in our study the assessment of the admission efficiency and of the effectiveness of the unit was possible by using the prism score of admission. there was no significant difference between mean values for otiss and ntiss)in level l patients (p= . paired t-test).for level and patients mean value of ntiss was greater than otiss (p< . ). there was a significant correlation between levels using either ntiss or otiss (mean difference level and , level and , ( p < o.oool). conclusions: a new tiss has been developed and used in a picu. nurses were able to accurately score the interventions on their shift. the assignment of patients to intensive care levels correlates with tiss values allowing a quantitative measure of severity. objective : to compare the rate of cerebral palsy (cp) between monochorionic-twins, dichorionic-twins and singletons born at to weeks' gestation. design : two-year prospective cohort study. setting : geographically defined study (region of franche-comt~., france). main outcome measures : type of plasentation was obtained by anatomopathological, or macroscopic examination of placenta and comparison of twins' blood-groups. neurological assessment was performed at two years of age (uncorrected for gestational age) by family doctor (pediatrician or physician), or neonatologist of the icu at tertiary center. sample : of i survivors aged of two years ( % follow-up rate), born between / / and / . triplets and chromosomic malformation were non included. results : thirteen ( %) of the singletons had cp.vs / ( %) of dichorionic twins and / ( %) of monochorionic twins (p= . ). four of the monochorionic twins ( %), / dichorionic twins ( %) and / ( %) nngletons suffer from quadriplegia (p< . ).in a multivariate approach, monochorionic twin placentation was the strongest risk-factor of cerebral palsy (or= . , ic % = a- , p< . ). others risk-factors of cp were : lack of father's profession (or , p< . ), maternal antecedent of abortion (or . , - , p< . ), vaginal delivery (or . , - , p< . ), hyaline membrane disease (or . , . -t , ~ . ). discussion : this is the first population-based study to uplight the role of monochorial twin-placentation as a strong risk factor of cp for premature infants. cp is more severe in monochodonic twins than in other infants. mecanism of cerebrat deficiency is not clear since none of our infants with cp was survivor of an in utero cotwin's death, and none of these infants was exposed to twin to twin transfusion syndrome. were these monochorionic-twins affected by an undiagnosed neurological structural defect that could lead both to prematurity and handicap remains an open question, a vital role of the intensivist is to ensure that knowledge and practice are imparted to trainees in the icu so that patients receive optimal care. teaching effectiveness varies widely leaving gaps in knowledge and practice in the trainee. being an effective teacher should not be a "gift" of a privileged few. the icu provides a fertile ground for using a variety of methods for teaching, e.g. didactic, at the bedside, emergencies, and in the performance ofproeeaures. in this environment, much can be learned. we have embarked upon a program to facilitate this learning process. i) teaching needs to be recognized as the foundation of good clinical care, i.e., patient related, and in its ability to generate discussion and research investigation. ) teaching structurally has many components including the speaker, audience, varying situations, and the message delivered. ) establishment of a program using these components to enhance teaching abilities at all levels, a) evaluate base-line teaching skills initially, b) individualize interventions to improve teaching skills, e) demonstration of learned skills with re-evaluation. this process is analogous to the analysis of a clinical disorder in a patient which, once recognized, interventions are then instituted and then re-evaluated. ) instill the desire to use these attained skills to teach and interest others to teach. teaching excellence should be recognized through awards, honors, and academic advancement. a major emphasis of this program is to provide participants with skills necessary to teach thought processes, decision-making skills (what to do, what to avoid) and implementing appropriate management during stressful emergency situations common to the picu. introduction: many" e-mail based discussion groups exist on the internet to provide medical professionals with a rapidly responsive medium for the international exchange of ideas relating to patient care. several such lists each serve more than a thousand professionals in more than countries, each distributing a dozen or more messages each day to every subscriber. there is very little known about the time being spent by professionals interacting with these lists, and very little known about the impact of the discussions on patient care. we wished to test the hypothesis that these discussion groups provide infortuation which is being used to change the care of individual patients and the general approach to patient problems. methods: in early january a pilot electronic survey was sent to a small fraction (n= ) of the memberships of e-mail discussion groups, picu@its.mew.edu, and nicu-net@u.washington.edu (the full memberships of both. groups (n=t for nicu-net, n= for picu) will be surveyed in early february of ). participants were asked for demographic information, experience and skill level relating to e-mail, time spent with the discussion groups, perceived usefulness of different types of discussions, and the ways in which the discussions were used clinically. the pilot study was analyzed for construct validity by correlating an overall assessment question with a summary of the specific questions. scale reliability was measured by cronbach's alpha statistic. results: the pilot survey response rate was ( %). the majority of respondents were male physicians, with an average age of +_ years, who had completed subspecialty training in intensive care, and were working at a university-affiliated hospital. most had been using e-malt for more than months, and considered themselves moderately adept in that use. % felt that the list helped weekly to keep them informed about current issues and practices in their field(s), and % felt that, at least monthly, they used information from the list(s) that was not readily available in medical journals. overall, % agreed that the list improved their professional competency. when asked to compare the value of months of membership on an e-mail discussion group with more traditional educational media, % compared it with attending a national conference, and % compared it to a journal subscription. cronbach's alpha was . , construct validity testing yielded coeff=. , p <. . conclusior~: internet-based e-mail discussion groups for health care professionals can be an important part of a strategy for maintaining professional competency. despite the very low cost of this medium for most, the value is felt to be comparable to that of t~r more expensive forums for education. further study will include distribution of the full survey in early february of . fronk shann, tony slater, gale pearson and the pim study group we have developed a new score for predicting the risk of mortality in children admitted to intensive care. the score is calculated from only seven variables collected at the time of admission to icu: mechanical ventilation (yes/no), booked admission after elective surgery (yes/no), the presence of any one of specified underlying conditions, both pupils fixed to light (yes/no), the base excess, the pao divided by the fio , and the systolic blood pressure. most scores used to predict outcome in intensive care require the collection of a large number of variables (so many icus do not calculate them routinely), and they use the worst value of each variable in the first hours in intensive care. this means they appear to be more accurate than they really are (about % of child deaths in icu occur in the first hours -so they are diagnosing these deaths rather than predicting them), and they blurr the differences between traits (a child admitted to a good unit who recovers will have a low score; but the same child who is mismanaged in a bad unit will have a high score -the bad unit's high mortality rate will be incorrectly attributed to its having sicker patients). pim was developed in the picu at the royal children's hospital in melbourne, and has been tested in six other picus in australia and one in the uk. objectives: to study the characteristics of the muhiorgan dysfunction syndrome (mds) in children. methods: a retrospective study with all the children with mds diagnosed from january to june is presented. children fulfilled the wilkinson criteria (i). in all of them the number of organs affected and the prims score were determined during the first hours. several groups were performed according to the clinical diagnosis, the hospital of origin and the order of organs affected. results: the subjects studied were an % of the pediatric intensive care unit admissions. of them expired ( %). no differences in age, sex and weight were observed between the children dying and the survivals. the most common causes of mds were sepsis, both nosocomial ( %) and medingococcal (i %) and acute respiratory failure. sixty-fivepercent of the patients were from the hospital wards and the remaining were directly admitted to the pigu from the emergency room. the systems affected were: respiratory ( %), cardiovascular ( %), hematologic ( %), central nervous system ( %), renal ( %) and (hepatic) liver ( %). the organs initially failing were: heart ( %), tung ( %) and central nervous system ( %). the children dying had a larger number of organs with failure than the survivors ( . v,s. . , p< . ).the prmis score was higher in the children expiring than in the survivors ( . v.s. , p < . ). s.mmary: the mds is a common pathology in picu, with a high mortality, the mortality is higher in children with a larger number of organs affected and a higher prism score. sepsis is the most common etiulogy. methods : from june ist to july th , all patients admitted to the pediatric icu were included. the score was measured at day (d ) and day (d ) and we used variables. for each organ system, we defined categories : dysfunction or failure, which we respectively confered or points. results : patients were admitted : newborns, children. were medical and were surgical patients. ( %) patients had two or more organ failure at the admission, ( , %) patients died, which ( %) in the first hours. the mortality rate was the same for children with two or more organ faiiure at d and d : / ( , %) at d , / ( , %) at d . the mean score is different for children who survived or who died : , versus , at d ; , versus , at . when the score is > , the mortality rate is significant. conclusion : in this study, there is a good correlation between the score of severity and the mortality rate but we have few included patients. we need a prospective multicentric study to assess these results and we must compare this score to other scores of severity used in picu. back.qround: injury to the central nervous system is the cause of death in the majority of pediatric trauma victims, studies have identified a wide range of factors associated with poor outcome from brain injury. however, when single features are analyzed, they are not sufficiently accurate predictors. few studies have used a multivariate analysis of these factors and pediatric outcome, methods: clinical and radiographic features of comatose children after traumatic brain injury were analyzed, clinical parameters, the initial cranial ct scan, and demographic characteristics were analyzed for an association with death or vegetative survival at months. a tree diagram in which risk factors may differ within the study subpopulations was constructed using recursive partitioning. results: chitdren with a motor score _< had an -fold increased risk of poor outcome compared to those with motor scores > . among patients with scores of _< , those with abnormal pupillary reflexes experienced a -fold increased risk of death compared to those with normal pupillary reflexes. among patients with a motor score > , an intracranial diagnosis code (no pathology, mild shift _< mm, swelling, shift > mm, surgical mass lesions, or non-operative mass lesions) was highly predicative of poor outcome at months. children with ct findings other than normal or mild swelling had a -fold increased risk of poor outcome. of children with swelling, shift or mass lesions, the pupillary light reflex was associated with outcome. children with abnormal pupils had a -fold increased risk of poor outcome. discussion: a few clinical and radiographic features stratified comatose children into fairly distinct risk groups. information available early after traumatic brain injury in comatose children provides useful prognostic information on the likelihood of death or devastating injury. a retrospective study of children with the diagnosis of epidural hematoma was made during - period. ages ranged between days and years ( % less than year, % between and years, and % older than years), % of them were admitted at the picu. % of the cases were due to falls, % to road traffic accident and % to other causes. on admission gcs was less than in % of the cases and more than in %. diagnosis was made during first hours in % of patients and delayed more than hours in % of them. neurologic impairment was present at admission in % of patients, and delayed in %. even so, % remained without impairment. radiological findings at first ct were skull fracture ( %); epidural hematoma localization was: in the right side ( %), frontal area ( %), temporoparietal ( %) and occipital (t %). associated lesions were: several ( %) or unilateral ( %) cerebral contusions, diffuse brain oedema ( %), unilateral hemispheric oedema ( %) and % showed shifted middle line. four patients died, half of them during the first hours. fully recovered ( . %) and have sequelae of different nature : were left with severe motor disability ( %); at the follow-up t have some degree of neurodisability. next datas keep correlation with death or neurosurgical impairment: only were significative multiple cerebral contusion (p= . ) and brain oedema (p= . ), gcs less than at the admission (p-- . ), shock (p= . ) and remaining cerebral contusion in control ct correlated with death or diasability at discharge. on the other hand, neither surgical drainage volume nor first or highest levels of icp ( cases),nor pupillary abnormalities ( cases) correlated with worse prognosis. conclusion: gcs equal or less than an shock are main factors related to worse prognosis, also multiple cerebral contusions in ct and diffuse brain oedema. the results of a modified gcs were compared to outcome and intensive therapy in children (mean age , t , years) with head and associated injuries ( , % of all cases) of different causes (traffic accidents, falls). the gcs was regularly used inn the course of intensive therapy. according to our own and other experiences the gcs was divided in stages: stage ( - points), stage ( - points) und stage ( - points) palhuiugy wile sp, tdhlg c~'lcb al blood ~ w. sabgcqucntl}. rhc slat,: rerltncd to t tl, iiltlils. the p st,~pem~v~ b}i~g wij!!,:q ! ,:_a!~p!ica!j n~:. ri~;¢ ill the level of sensibflizatjou lo tile cerebn~ anhgrns up to t. -o was flofcd iu i,alicnts. there wa.~ al~ iuclt~a~e ill cerebral vdociij,. ~m d~;'ati a il~ p¢fiphc~ai re~ista/isc of the large ce~'bral ve~ds. neur h;~c ~:yn'.pt,m~at !a~, (s::mno!en~', _r_uscu!~r l~:pot ni& !ryper*'flema) was nbserwed tu lt~ese pal~enls o. cbruc~l ~ nnds. rile ple~c.ut abse~vafion~ suggesl ihal die ~tttdy at" ihe stale ~f hematocr~chcplm/itic bm~ic~ in ckil&en with on emergensy is of abviou.~ !?ece~sib; in co~.te ctin g severe pa~ lo ~-i~mnediately f u wing ne ,:~per,'~fion. background: reconstruction of the heart by three-dimensional ( d) echocardiography provided new information on anatomy of complex congenital heart defects, we assessed the utility of d ultrasound in detecting morphological changes in cerebral anatomy in newborns before and after cardiac surgery. methods: transfontanel cross-sectional ultrasound, scans were obtained in standardized coronal and median sagittal planes. subsequently, rotational scanning was used to acquire the multiple sequential crosssections of the brain. for rotational scanning, a conventional mhz transducer was rotated degrees.scanning took less than one minute and required no sedation, data was stored in the image processing computer which allowed for off-line three dimensional reconstruction of different brain regions.twelve infants aged - (median ) days were assessed before and after cardiac surgery, results: cavity of lateral ventricle, choroid plexus and the periventricular brain parenchyma could be reconstructed in all. accurate estimation of size and volume of lateral ventricle, aqueduct, and other ultrasonographic visible pathological brain lesions could be performed. reconstruction of various brain areas was accomplished in - minutes. the localisation and extension of severe periventricular hemorrhage which was detected preoperatively in one infants was better visualized than in conventional ultrasonography. epicortical and subarachnoidal space could be reconstructed in all and allowed detection of hemorrhage in one case which was not detected by conventional ultrasound. conclusion: d reconstruction of different areas of the brain may provide additional quantitative information on size and volume of the internal ventricle and choroid plexus, and better understanding of the topographical aspects and the extension of intra-and periventricular hemorrhage than conventional cross-sectional ultrasound. introduction: intracranial cerebral blood has been estimated to be % venous, the invasive measurment of venous blood saturation in the jugular bulb provides quantitative information on cerebral oxygen supply and consumption. however, routine oxymetric measurement of blood saturation in the jugular bulb by insertion of a catheter line into the internal jugtdar vein is an invasive procedure which has limited use especially in infants and young children. thus the aim of this study was to investigate the correlation between the non-invasive spectroscopic measurement of rso and the oxymetric determination of the blood saturation in the jugular bulb in infants and children undergoing routine cardiac catheterization.. methods: during routine cardiac catheterization infants and children (age day- year, median , year) the rso was measured continuously using a two chanel cerebral oxymeter (invos a). the sensor was placed in standardized location at the left temporal head side. after the routine oxymetric blood sampling in the superior vena cava the oxymetric catheter was manupilated into the left jugular bulb. after control of the catheter position simultenuous values of the rso were documented. results: over a range of ( - %) sjo , a significant linear correlation was found between the spectroscopic measurement of rso and the oxymetric determination of venous blood saturation in the jugular bulb (r= , , p< , ) and the superior vena cava (r= , , p< , ). no significant correlation was found between rso and the arterial blood saturation in the descending aorta and as well as to the standared hemodynamic parameters. conclusion: meusurement of rso by mrs may provide continuous non-invasive information on cerebral venous blood saturation and thereby possibly on cerebral oxygen supply and consumption in infants and children. these may be of clinical value particulary during and immediately after heart surgery by means of non-pulsatile cardiopulmonary bypass. information on refractory status epilepticus (rse) from developing countries is scarce. we analysed cases of rse admitted over last yrs. the objective was to study etiology end evaluate efficacy of diezepam infusion. median age of the patients was . years irange . months to t . yrs); % were boys. onset of seizures was -t hours (median hours) prior to hespitalisation. the glasgow coma scale score ranged from . (mean+sd + ). the commonest underlying causes were acute cns infections ( / , %; bacterial meningitis, , encephalitis, ) and epilepsy ( / , %). oiazepam infusion in incremental dose (range . - . mg/kg/min) was used in patients over . _+ . days. seizures were controlled n ( %), mechanical ventilation was required in ( %)only, while none had hypotension; % patients survived. thiopental infusion (holus mg/kg followed by . mglkg/min, and increments of . mg/kg/min till seizure control) was used in patients over . _+ . days; seizure were controlled in all, but five patients needed mechanical ventilation, six developed hypotension needing infusion of vasopressoi drugs, out of ( %) died, overall mortality was %, mainly due to acute cns infections (n- ) and prolonged se. the patient was a -year-old gift di~aosed of dov,~'s s~drom¢, tetralogy of fallot. (t.f.) before admission a vasovagal crisis after coughing and vomiting was seen, and she was taken to the emergency room. mother said she had eyanosis in the mucous membranes of the mouth with exercise.on physical examination, she ~as afebrile, normal fundi and neurologic examination was normal. a harsh systolic murmur was hear~ with decrased intensity during bradycardia. chest rx disclosed a decreased pulmonary vascular markings. ecg: synus rhythm, with bradycardia and nodal escape rhyflmas. she was transferred to our picu because of severe h ,pertomc seizure, lost conciousness, and deeembrate poslamng~ ~t cyancx~is. the episode lasted for ~weral seconds, and ceased v~th diazepam. on admission she was lethargy, and neurologlc exammation showed weakness of left leg without babinski, and normal funduscopic. the patient had two episodes of bradycardia and isoproterenol was begun. during those episodes the patient was cyanotic, and the murmur was heard with the same intensity. act scan disclosed a tight parieto-temporai abscess with midline shift, lnmediately after the diagnostic ct, we administered antibiotics, antiedema treatment and it was drained. the abscess culture was negative. a ct control disclosed air and midlme shift. ~ the next two days she had three episodes of h oxia and c'yauosis ceased with o@gen, morphine and propanolol the patient died during a fourth episode. discussion: arrhytmias are uncommon in patients with tetralogy of fallot before surgery. in our case the first diagnosis was sick sinus syndrome vs bradycardia secondary to cyanotic episodes. the incidence of cerebral abscess in children with congenital heart disease (chd) is approximately %. tetralogy of fallot is the most common associated lesion, and is unusual in children under years of age. conclusion: ) brain abscess is a rare complication of patients with cyanotic chd, but should be suggested in patients with °'apparent" sick sinus syndrome. in patients with down's syndrome, t.f.,with cyanotic episodes, and difficult neurologic exploration, a brain ct scan is recommended. guillain-ba~re syndrome (gbs) is an acute autoimmune reaction, directed primarily toward the myelin encasing the peripheral motor nerves= this reaction causes a delay or block in nerve conduction. the presentation often can be very subtle but is followed by rapid loss of neuromuscular power, leading to acute respiratory distress, resulting from weakness of muscles and aspiration pneumonia. there were boys - , , and i i years old with gbs, treated in our icu. two of them due to the respiratory distress were intubated nasotracheally and ventilated mechanically with servo- ooc (siemens-elema, sweden) ventilator. duration of ventilation was i i and days, respectively. plasma exchange was performed in all cases. the numbers of plasma exchange sessions were - in each case. mean amount of plasma exchanged per session was , ml/kg. plasma was substituted with albumin, plasma or saline. the most important aspect of the management of patients with gbs in the icu involves the airway care, prevention and treatment of aspiration pneumonia and the mechanical ventilation if respiratory distress presents. endotracheal intubation should be performed whenever there is evidence of retention of pulmonary secretions, refractory to chest physical therapy, weakness of protective reflexes of the airway, leading to aspiration pneumonia and (or) atelecr~sis. cardiac arrhithmias too, is a main threat to the circulatory stability in gbs. therapeutic plasmapharesis has been shown to be beneficial, reducing the time for weaning from the ventilator and for achieving independent ambulation. however, plasma exchange is expensive and not without significant risks for the patient. some authors find that plasmapheresis is not effective for patients with fulminant course of gbs and blocking of nerve conduction. recent studies have demonstrated that intravenous high-dose immunoglobulin can be equally effective. there were no significant complications associated with plasma exchange. all presented patients survived without residual disability. tetraparesis associated with long-term paneuronium use in an infant. paneuronium is a muscle relaxant used in ventilatory management of patients with respiratory distress in intensive care unit. after the end of sedation some patients were found to have severe tetraparesis. paresis was accompanied by complete areflexia and diffuse atrophy of alt extremity muscles. this neuromuscular complication is caused by prolonged high-dosage pancuronium treatment. in the last years, numerous reports have linked the use of pancuronium bromide with prolonged paralysis, disuse atrophy and areflexia. this side-effect is well known in adults patients but rare in a pediatric intensive care unit. we describe one pediatric observation of tetraparesis after prolonged pancuronium treatment in a -month-old girl, this female infant developed respiratory distress syndrome and was intubated and mechanically ventilated. to decrease chest wall rigidity pancuronium bromide was administered during days. (she received approximately mg of pancuronium bromide). on day the drug was discontinued and the patient had severe tetraplegia and areflexia with normal head movements. electromyograpliy showed absence of any disorder of neuromuscular transmission. this infant showed a recovely of muscles after months. the other causes of peripheral neuropathies were eliminated. electroencephalograms and head scans were normal. the recovery pattern observed in our patient correspond to the process of regeneration after axonal degeneration. it is suggested that these neuromuscular complications were caused by prolonged high-dosage pancuronium treatment (associated with cortieoid and aminoglucosides). polyneuropathy syndrome in adult lc.u. appeared in literature in and is extremely common in long stay cases. the etiology of these disorders remains elusive. it is tempting to ascribe them to administration of drugs (muscle relaxants, steroids, aminoglycosidea), plolonged immobility, malutrition, sepsis and ischemia associated with reperfusion injury. to our knowledge there is only one case report of similar condition in a children i.c.u. (pascucci ) we present a serie of previously healthy children, aged months to years, who admitted in i.c.u with respiratory failure and who following weaning from m.v, remained in profound diffuse hypotonia with proximal and distal muscle weakness for various length of time, recovery of muscle strength occured in a week or months {the longest i months), all children, except one, - days before admission developed symptoms of either respiratory or upper airway infection with fever. on admission viral and bacterial cultures were positive in cases (haemophilus influenze, herpes virus). during treatment patients became septic. muscle histological and neurophusiological investigations have not been done. considering the multifactorial nature of the aquired nmd in adult critically ill pts, is impossible to attribute the muscle weakness of our pts to any specific cause, in conclusion, our findings suggest the need for further investigation of nmd in critically ill children treated in i.c.u. a van esch, ha van steen~l-m , ir ramtal, g derksen-lubsen, idf habbema. febrile status epilepticus (fse) is a prolonged and serious febrile seizure. little is known about the outcome of fse in neurologically normal children. this survey involved patients between months and years of age who had visited due to their first fse, the sophia children's hospital during the period of january till december . patients with a history of neurologic disorders were excluded. patients were identified, % were male. the cause of the fever remained unknown in % of the cases. in all case the fse was generalized and it most frequently occurred at night ( %). the mean age at fse was t. years ( . - . ), the mean temperature . °c ( . - °c). the mean follow up time was . year. twelve children ( %) had neurologic sequelea. the neurologic sequelae varied from speech deficit ( case mild, v - year delayed; case moderate > year delayed) to severe retardation and epilepsy ( cases). speech deficit was detected after a mean period of months (range - ), age, gender, temperature, family history and time of onset were no significant risk factors for neurologic sequelae. duration of seizure [rr . ( . - . )] and more than two drugs to treat fse (rr . (t. - . ) were related to neurologic sequelae. we recommend that fse children should be followed for at least a year to detect possible speech disorders properly and start early intervention. unusual presentation of myasthenlg gra%qs ibtza e. modesto ,v~ abe~gochea a, sanch]s l all, go l varas k folgado s, garcia e. p. .c.u. la fe, valencia. spain case report: the patient was a -year-o!d gift transferred to our pic because of severe respiratory failure. the patient, convaleseem of ehiekenpox, came into contact with horse manure previous afternoon. in the morning, she was lethargy, and irritability, with poor finding, and ~ an episode of coughing, cyanosis and acute respiratory failure after mucous vomiting when she was drinking milk. on admission she had severe respiratory distress, respiratory acidosis, and the sat was %. she was mtubated without difficulty, and was transferred to our p.i.c.u. physical examination reveals stable hemodynamies, pupils equal, round, reactive to light, normal fandi, and muscle relaxation. crusted vesicles diseminats~d. rhonehi over both lungs. hepatomegaly (+) and splenomegaly (+). ~lhe urine, hematologic, and c.s.f. laboratory findings were normal. c.t. scan of the brain, e.e.g., and ekg. revealed no'abnormalities. rx chest disclosed a retrocardiac atelectasis. speci~ts of stool and blood were obtained for cultures and study of c. botul#num toxins. pending receipt of these results, a broad-speotmm antibiotic and acyctovir was begun. the initial differennal diagnosis consisted of laryngospasm associated with aspiraqlon, botulism, and postmfecfious varicella encephalitis. after hours, weatm~ was begun. the neurologic examination showed a low modified glasgow coma ~ale (mgcs), generalized hypotouia and muscle weakness. these data suggested three diagnoses, posfnfecfious encephalitis, residual neuroumsoaar blockade, and excessive doses of sedative and analgesic drugs. after hours she regained skeletal muscle poxver and ufltlcient respiratory effort, the mcgs was acceptable, and blood gases were normal. she was given n~-tigmine and atropine, and her tr~ma was extubated. an acute respiratory failure ocurrs ram. after. chest radioga'aph disclosed a left inferior lobe atelectasis. after hours weaning begun~and the same episode w~as seen. at this point her mother stated that the girl showed weakness of the eyelids or extraneular muscles. it suggested myasthenic syndrome vs ~-barr syndrome. c. botul#num toxins were negative, chotinesterase level ~as normal. edrofoinum test ~as positive. anti-acetyleholine receptor antibodies were negatives. e.m.g. confirmed myasthenia gravis (congenital vs juvenile serenegative). pyridostigmine was begun and the trachea was extubated without complications. conclusion: din the differential diagnosis of weamng failure we must consider ~c gravis~ )myasthenia gravis could resemble encephalitis, because of low ocs, overall if is triggered by viral infection. )in some diseases (this case) gcs could not he an aemuate index of mental state. a burguet*, a menget*, e monnet**, a gasca-avanzi*, c fromentin*, h allemand**, jy pauchard*, ml dalphin*. * r animation infantile potyvaiente chu st jacques besancon cedex. ** d~padement de sant publique besancon cedex, france, objective : to point out that strabism is) of one-year-old premature is a good predictor of a poor neurological outcome at two years of age. design and setting : two-year prospective cohort study and geographically defined study (region of franche-comte, france). main outcome measures : neurological assessment was performed at one and two years of age (uncorrected for gestationnal age). a mailing questionnaire was sent to the famity and fuu-filled by thefamily doctor (pediatrician or physician), or neonatologist of the icu at tertiary center, s was diagnosed at one year of age by the examinator but s was not used to diagnose cerebral palsy (cp). sample : of survivors ( %) evaluated at one and two years of age. results : correlation of one and two years neurological evaluation is weak (kappa= . ). correlation of s at one year and cp at two year is fair (kappa= , ). the goal of this paper is to review evidence related to hypothesis that the "waiting" axons and cells of the transient subplate zone may participate in the structural plasticity of the human cerebral cortex after perinatai brain damage (kostovic et al, metabot brain res : , t ) and to correlate this phenomenon with different forms and mechanisms of structural plasticity. it is our basic assumption that all lesions occuring during cortical histogenesis will lead to more or less pronounced structural reorganization. here we show that various components of the subplate zone participate in several forms of the structural "plastic" responses in the human cortex: modification of convolutional pattern, changes in size of cytoarchitecturat areas~ columnar reorganization, dendritic and synaptic plasticity. the etiological factors which induce lesions and subsequent plastic changes act via the following pathogenetic mechanisms: * disturbances of radial unit formation (rakic); * changes in ingrowth of afferent fibres; * changes in the rate of normally occuring reorganisational events, depending on the critical period for a given histogenetic event. in the present study developmental lesions (localized perlventricular leukomalacia and haemorrhages) were demonstrated by ultrasound in live-born infants ranging between to weeks of gestation. in younger infants ( - w) who died shortly after birth, examination revealed lesions of the white matter with the preservation of the subplate zone. in infants who died one week of more after the lesion, we have observed localized micropolygyria, cavities, condensed layer vi -subplate zone, and columnations of the cortical plate. these changes are less prominent if the lesion occurs after diminishment of the subplate zone (after w). since in the fetal cortex the subplate zone serves as predominant source of growing fibers, transient neurons, trophic factors and contains cellular substrata for migration, this zone is the most likely candidate for major types of structural plasticity. in conclusion, cerebral cortex of the low -birthweight infants is more susceptible to the various lesions but shows vigorous structural plasticity and conspicuous functional recovery due to the growing, transiently located neuron at elements. the mortality due to meningoccocal sepsis is high in spite of important progress in emergency and intensive care medicine. during the last decade multiple scoring-systems have been developed in order to establish a therapeutic approach and to evaluate the final outcome of a meningococcal infection. different clinical and biological data (shock, ecchymosis, peripheral wbc and platelet count, coagulopathy, acidosis, meningism, etc) are taken into consideration and the importance given to these data depends on the scoring-system used. a review of the different scoring-systems is given and a clinical case is presented. we report the case of a year old male, who was transfered to our icu hours after onset of temperature and skin rash. the parents described a fast deterioration of his condition. the boy presented wide spread ecchymosis, high temperature, no signs of meningism, circulatory insufficiency and shock, coagulopathy and low peripheral wbc and platetet count. disseminated intravascular coagulopathy developed promptly. the glasgow meningococcal septicemia prognostic score (gmss) was used and the obtained score reached the highest level ( / ). this corresponds to a % mortality. the patient required mechanical ventilation for days. at admission he received human albumine, fresh frozen plasma, dexamethason, dopamine, dobutamine and a continuous infusion of adrenaline. antibiotical treatment consisted of ceftdaxone. the evolution was favorable and the infant fully recovered. retrospectively the gmss was compared to other meningococcal scoring scales which gave the same mortality ( %). we conclude that the scoring-systems are important to evaluate the seriousness and to assess the therapeutic approach, but they should be used cautiously even when % mortality is predicted by several risk evaluations scoring-systems. the aim of this study was to assess the haemodynamic status on admission and the critical care management of children presenting with meningococcat infection. this was a retrospective study of the charts of consecutive admissions. mean age was . years (+/- . ). the average duration of symptoms prior to admission was . hours (+/- . ). on admission . % were hypotensive, . % had clinical signs of haemodynamic instability and . % of cases that had a blood gas analysis on admission had a metabolic acidosis (bases excess < - .q): the mortality rate was . %. % of patients that died were hypotensive on admission and all had a metabolic acidosis. of the survivors . % were hypotensive on admission, % had clinical signs of haemodynamic instability, % required invasive pressure monitoring and . % were ventilated and received inotropic support. this study demonstrates that at the time of presentation with meningococcal infection children had a high incidence of established haemodynamic instability. successful management of this infection is dependent on early presentation and initiation of therapy and on aggressive support of the cardiovascular and vital organ systems. dept. of intensive care medicine and dept of infectious diseases, our lady's hospital for sick children, crumlin, dublinl , ireland. jude. pediatric intensive care unit, ch&u, lille-france. more than % of children surviving sip (defined as purpura with shock) have snli. objective. to search for a specific hemostatic profile in children with snli. patients and methods. between may and march , children with sip were admitted to our picu : ( . %) died and ( . %) ranged in age from to months (mean : ) survived, of them ( . %) with snli (defined as the need of a surgical procedure). in survivors, two hemostasis studies (between h and h , and h later) included the determination of coagulation factors (routine tests), protein c (pc : amidolytic activity, biogenic), total protein s (ps : elisa, stago), c b binding protein (c bbp : laurell's technique, stago), antithrombin (at : chomogenic test, stago), and plasminogen activator inhibitorl (pail : chromogenic test, biopool). three severity scores were determined at admission : french group of pediatric intensive care, gedde-dahl, and crp. statistical analysis used the wilcoxon's test. results. at admission (lst sample) severity scores and at , pc, ps, c bbp levels were not different between the group with snli and the group without snli ; quick time ( - % vs ± % ; p = . ), vti+x ( . % vs - % ; p = . i) and pall ( - ui/m! vs . ui/ml ; p = . ) were lower in the group with snli. on the nd sample there was no difference between the two groups. kinetics of hemostatic abnormalities was not different between the two groups. conclusion. in the literature, intravascular coagulation (dic), low fibronectin and at were identified as predictors of snli, and a negative correlation was found between the mean size of the skin lesions and pc activity, at , and total ps. in this series, apart from dic, there were no specific hemostatic abnormalities that support the use of treatments such as pc, at , and pail antibodies administration to prevent snli. further studies including more children are needed. the aim of study was to investigate the efficacy of intravenous immunglobulin with enriched igm content pentaglob/n /biotest/. in our pediatric intensive care unit ten septic children /group i/-their average age , years /sd:o, /, of them with gramm negative and one with gramm positive blood cultures, and two with unindentified bacteria-were treated with basis sepsis therapy and pentaglobin. the application of pentaglobin was as follows: , ml/kg loading dose for one hour, followed by a continuous intravenous infusion , - , ml/kg/hour depending on body temperatura /lanser scheme/ for - hours. another ten septic patients /control-group ii/the mean age , years/sd:o, /, their blood cultures were gramm negative bacteria , positive , and the bacteria was not indentified in two cases -were treated with only the basis therapy. results: the duration of intensive treatment decreased from an average , days /sd: , min -max days/ to , days /sd: , min -max days/ in the group treated wit pentaglobin. the difference was significant /x p< , /. in the group i nobody died, but three in the group ii. conclusion: the pentaglobin therapy can improve the efficacy of the basis therapy of sepsis. sinus bradycardia after an episode of sepsis is a rare symptom complex decribed in children with hematologic malignancies. we present a case of postsepsis bradycardia following severe typhlitis and septic shock in a year old boy with relapse common all. blood and ascitic fluid specimen grew clostridium species and pseudomonas aeruginosa. at surgery there was a necrotic gangrenous terminal ileum and cecum, requiring ileocecal bowel resection with ileostoma. while clinically recovering from sepsis he developed bradycardia for hours. extensive diagnositic procedures was given and the heart rate slowly increased to normal range of age. postsepsis bradycardia in children with hematologic malignancies after an episode of sepsis is self-limiting and after careful differential diagnostics warrants an expectative attitude. nitrate level is known to be enhanced during sepsis. serum nitrate is the stable metabolic end-product of endogenous nitric oxide generation. nitric oxide has demonstrated to be a powerful anti microbial final mediator and also a key molecule driving to the lethality of one of the most common complication of sepsis; the endotoxic shock. such facts prompted us to investigate the possible diagnostic and/or prognostic value of monitoring serum level in high risk, presumptive and confirmed sepsis patients. additionally we have explored the usefulness of this mediator as index of therapeutic response. in our study it is demonstrated that there is an important relationship between nitrate level and the occurrence of neonatal sepsis. septic newborn group showed fold higher nitrate level than that of healthy control group. in addition, the group of patients with high risk of sepsis which finally became septics, exhibited fold higher nitrate level at - hours before the first symptoms appeared, when compare with those who did not develop sepsis. however in the presumptive sepsis group, there was no difference between the patients which finaliy ,&'ere considered septics and those which not. in all septic cases, after days of a successful therapy with antibiotics, the level of nitrate diminish fold. our results suggest the utility of monitoring nitrate as index for the diagnosis of neonatal sepsis. the potential benefits of exchange transfusion, plasma exchange, and haemofiltration have all been described in children with overwhelming sepsis. however, little hard evidence exists to prove the benefits of any of these techniques. i have treated five patients with plasma exchange (pe), having been asked to see all these patients at a point when it was felt death was inevitable. two of the patients had staphylococcal, two meningococcal and one enterococcal septicaemia. all patients showed a dramatic haemodynamic improvement following pe with improvement in blood pressure, reduction in inotrope requirement and improvement in tissue perfusion. three patients survived. one of the patients with staphylococcal sepsis and both of the patients with meningococeal sepsis had developing gangrene of the limbs which showed remarkable reperfusion with pe. in two of the patients measurements of cardiac output (co) and systemic vascular resistance (svr) showed ~a reduction in co and a rise in svr over the course of a pe despite the reduction or cessation of vasoconstricting inotropes. many believe haemofiltration is of value in septic shock. a trial with a no treatment limb is difficult to achieve. i believe we now have enough evidence to justify a controlled trial of haemofiltration versus plasma exchange in patients with septic shock and unstable haemodynamic status whilst on inotropic support. during the next several days, cough and chest pain suggested pulmonary embolism confirmed by radiologic evaluation. echocardiographic examination showed multiple thrombosis of the superior vena cava, right atrium and ventricle and pulmonary artery. estimated protein c level was . % (normal range - %); identical deficiency was found in patient's mother and elder sister. cvc was removed, and alter -month heparin therapy and supstitution of protein c with fresh frozen plasma, there was almost complete thrombolysis of the great vessels and cardiac chambers. we conclude that invasive diagnostic and therapeutic procedures in such patients may result in higher risk for severe thrombosis at unusual sites, and numeuos further complications bronchopulmonary dysptasia (bdp) is a chronic pulmonary disease of preterm and term babies treated with mechanical ventilation for respiratory problems of different origin and requiring oxygen therapy days after birth. bpd is a disease affecting the growth and development of pulmonary tissue. such pulmonary }esions heal by squamous metaplasia leading to scar formation and fibrous tkssue r~growth, the pediatric intensive care unit makes the survival of babies w~h very low birth weight ( - g) possible. with the increase in their aulyival, the number of complications in low birth weight babies increases as well. bdp is a very serious complication. therefore the importance of early diagnosis and treatment of bdp must be stressed in order to reduce the consequences. babies with bdp must be under medical suveillance for at least years as the disease needs at least that long for complete resolution. tn the icu of pediatric department at madbor teaching hospital: during the past two years ( - ) newborns were treated with mechanical ventilation. the neonatal and postnatal death rate of all newborns admitted to our icu was , %o.ln the two years from to , newborns were admitted to our icu ( %~ of all newborn babies at maribor teaching hospital), with birth weight - g. in the icu, the survival of these babies and parallel to it the number of complications is increasing. during the mentioned -year period, babies with very low birth weight ( - g) survived: in and in t . in - %, first or second stage bdp was treated,there was no case of third of fourth stage bdp. the treatment consisted of eary removal from mechanical ventilation, oxygen therapy~ intensive treatment of infection, volume and caloric intake contro}, corticosteroid treatment throught weeks with decreasing doses, diuretic end antioxydant therapy. the children are to be reevaluated at the age of and months and again at i and years. oeure j van der, markhorst do, haasnoot k department of pediatrics, pediatric intensive care unit, free university hospital, amsterdam, the netherlands. case summary a %-month . kg girl of african origin was admitted to the pedfatric irtensive care unit with pneumonia and progressive respiratory irlsuffjderey. she was intubated and ventilated by pressure regulated volume controijed ventilation (servo c, siemens, soma, sweden). maximum conditions were inspiratory minute volume . l, peep cm h~o ahd % ~. chest x-ray showed bilateral interstitial consolidation. material obtained by broncho-alveolar lavage showed preumocystis car}nil htv-serology (elisa and westerll blott) and p -antigerl were positive, confirming the diagnosis of pediatric aids. she was then treated with high dose co-tllmoxazoie, penthamldine, z{(~ovudire and steroids iv. because of thee x-ray features, high need for o ( %, pad mm hg), not responding to elevatiofi of peep (max cm h=o) and pao /fio = < (s ). m acute respiratory distress syhdrome (ards) was diagnosed. because conventional ventilation (cv) failure, hfo-v ( ooa, serisor medics,yorba linda, ca) was initiated. starting mean airway pressure (map) of cm h~o was based or map of the cv, oscillatory pressure amplitude (dp) of was, at ii~itial frequency of . hz, adjusted ur~til chest wall vibrations were visible, it was required to raise map to cm h and dp to before optimal lung volume and ventilation were achieved and need for o reduced within hours, this was monitored by frequent blood-gas analysis and chest x-rays. map and dp could slowly be reduced, after a good response the first day, gradually demand reduced and the patient could be weaned from the ventilation. map, dp, fi and oxygenation index (map x pa ~jfio ) are shown in table i. chest x-ray follow-up showed gradually improving lung features, with marked improvement of aereation. after days hf -v she could be succesfully detubated when a map of cm h was acmeved. results : sianificant increase in ventilato~ rate and mean airway pressure was noticed after the change to savi. no differences in oxygenation, co partial pressure and systolic, diastolic or mean blood pressure between imv and savi periods were noted. in infants however an improvement in pao /p .ol/ and decrease in paco was observed after the switch to savi. these babies had a lower initial a/a oxygen tension ratio and required higher initial ventilator rate /p<o. / in comparison to the rest of the studied patients. conclusion:infants with low a/a po ratio who need high initial ventilator rate are likely to obtain benefit from savi.. fumimare hatori, haruo uchida, masao katayama, and rika muto department of anesthesia and critical care medicine chiba children's hospital, chiba city, japan. purpose: this study was made to find out whether the heat and moisture exchangers (hme) was effective enough to humidify in the respiratory management for children. patients: the study population consisted of patients who were provided with artificial airway. they were divided into the two groups: ii with heated humidifier (a) and i with hme (b). methods: hme could be used for those without pulmonary complication and with less leakage around the endotracheal tube. in both a and b groups, (i) we have measured relative humidity, temperature, and absolute humidity at the endotracheal tube connector. for these measurements, humidity sensor system was utilized. ( ) any troubles and complications caused by the artificial airway were investigated. results: (i) in group a, we confirmed the mean values as the relative humidity of . %, the temperature of . ~c, and the absolute humidity of . mg/l. in group b, they were . %, . °c, and . mg/l respectively. no difference was noted between the two groups as to the relative humidity, but the temperature and absolute humidity were lower in group b with a significant difference (p< . ). ( ) in neither of the groups, any respiratory complications such as obstruction or stenosis of the artificial airway were noted. in pediatric icu, hme can be used without any problems under given conditions. titei: objective: evaluate the precision, reproductibility and aplicability of a new device to measure auto-peep in pediatric patients during mechanical ventilation. material and methods: it's an electromc-pneumatic eontolled device with an oclusion valve and a pressure monitor installed between endotracheal canulla and the ventilator circuit. the measurements were performed with a sollenoid conected with to the ventilator to detect the end of inspiration phase and the actwation of the oclusion valve. the signs of pressure and flow were moniturized using a difere~rtial transducer and it was processed using a pc computer and a pneumoviaw® software. the auto-peep also displayed in the ventilator pressure monitor. results: we submitted children with neuromuscular disease or respiratory distress in this study. the incidence of auto-peep was %, with variability between , to cmh . all the measurements were performed times to verify the precision and the reproduetibility and evaluation of the pressure and flow curves. conclusion: the device is low cost, easy to use and can be used without the pneumotacograph. the auto-peep measurement can be done during mechanical ventilation with time cycled, pressure limited and continuous flow, the most commom ventilator used in pediatric patient. resistance was modeled by endotracheal tube (ett) sizes ( . , , , ), and compliance by test lungs ( and ml/crnh ), for each resistance -compliance arrangement, we measured tidal volume (vt) at various frequencies, peak to peak (p-p) and mean airway pressures. results vt increased when ett size increased with all ventilators in accordance with fredberg results (jap, ; : - ) . the maximum vt delivered was smaller with the hfv-babyiog than with the ohf or the sm a at a given frequency (figure on the left). increasing p-p resulted in a linear increase in vt delivery in the - % range of maximum p-p (figure on the right). hfv-babylog didn't provide significant vt increase when p-p was set above % and vt deliven/decreased when mean airway pressure decreased. --i ~ sm a a number of ventilatory parameters had been analyzed with russia-produced monitor humitemp htm (servisinstrument j,-s. co) in newborn infants with rdsn of t-iv stages on cmv, cppv, imv, and cpap steps using respirators of various kinds (stephan-staxel, bear- , newport rezee, sechrist). parameters investigated were: temperature of inspired mixture (t), relative and absolute humidity (rh and ah, respectively), static compliance (c), expired tidital volume (vte), expired minute volume of ventilation (mve). the monitor read these parameters in following ranges; rh-from to %; ah-up to mg/i; mve-up to . i/mini vte-from to ml; t-accurate to %; c-from . to t ml/cm h . continuous monitoring time ranged from to hours, it was noted, that during this time the monitor and transducers had no malfunctions, and it's using didn't require additional calibration, it was found, that humidifiers mr and mr (fisher, paykel) couldn't maintain parameters t and ah continuously (these values varied from , to . c and from , to . mg/i, respectively, when heating regulator position was unchanged). changes of parameters c depended clearly on cppv conditions and rdsn severity (with rdsn of stages i-ii c value ranged from . to . ml/cm h ; and with stages iit-tv-from . to . ). tn newborn suffering from rdsn of stages ii-ltl and ill-lv with pip> mbar, fi > , , peep= - mber, c-from . to . ml/cm h , effectivity of exosurf therapy was studied. in newborns in - hours of therapy pip decreased to . - . , and c increased to , - . ml/cm h . in newborn infants with aad > mmhg and c from , to . mltcm h positive effects of exosurf on lung compliance were not observed. in newborns the monitor had revealed decreased of c (from . - . to , - . ml/cm h ), manifested clinically by pneumothorax. in general, monitor htm made possible; ), to estimate the adequacy of cmv-parameters and regimes in newborn infants; ). to select optimal t and ah values in the respiratory outline in dependence on lung damage severity and infused volume; ). to reveal rdsn severity; ), to optimize indications and adequacy of surfactaot therapy; ). to diagnostieate the air leakage syndrome; ). to effects to some agents (broncholytics, spasmolytics); ). to obtain objective indications for imv/simv and cpap regimes. albano communication is an important aspect of human development and existence, and an inability to vocalise can be a problem in ventilatordependent patients. we present our experience with speaking aids as a means of enhancing verbal communication in four ventilatordependent children in our paediatric intensive care unit. the age of the children ranged from months to years, and the period of ventilation ranged from months to months via a tracheostnmy. they require continuous flow generated pressure limited or control ventilation at rates of - bpm. the reasons for ventilation include tetraptegia following a shrapnel injury; tetraplegia following congenital cervical spine damage; tetraplegia following atlanto-axial subluxation; and critical illness polyneuropathy following adult respiratory distress syndrome from prolonged ventilation for a severe head injury. the first three patients have passy-mnir one-way speaking valves and the final patient has a bivona foam cuffed tmcheostomy tube with a talk attachment in view of recurrent aspiration. an improvement in quaiity of speech has been shown by independent assessment. we will review the present literature on this subject and discuss the advantages and disadvantages of these two types of speaking aids in the light of our experience. the prognosis of antenatally diagnosed cdh is closely related to the degree of ph. there have been attempts to correlate antenatal or postnatal criteria to mortality: none have been demonstrated to be predictive of lethal ph. the aim of this retrospective study was to determine whether antenatal or early postnatal data could correlate with the findings of post-mortem examinations. patients and methods: between july and july , cdh patients have been antenatally and postnatally managed at our institution. twentythree infants underwent a post-mortem examination. ph was assessed by using the lung weight to body weight ratio (lw/bw) and the radial alveolar count (rac). antenatal results: cdh diagnosis was made at weeks of gestation (wg) ( - ). twenty-eight patients had a left sided cdh, had a right sided cdh, and one had a bilateral cdh. herniated organs were stomach none (n= ), or liver alone (n= ), or both stomach and liver (n= the patient was a -yenr-old girl with chronic renal insufficiency see~ to renal dysptasm, two months before admission a kidney trar~ptant was performed. one morah later she showed acute graft rejection with serum ereafinine (cr) level of . mg%. the rejection was unreslxmsive to an increased steroid dosage, and okt was begun with resolution of the rejection. one week arer, new rejection episode was seen marestxmsive to an increased steroid dosage, and transp~ ~s performed five days before admission to our ptc. hemedialysis and peritoneal dialysis (p.d.) each other day, was indicated (g.r.f.< ml/rnin). four days before admission t ~ rose to °c. "lhe diagnosis of opporttmistic pneumoma was made on the basis of tach ,pr',e~ hypoxi~ and diffuse interstitial infiltrates. senma ~ was positive for cytomegaloviras (cmv), and stool culture for c albicans. pentamidine, ganciclovir (dhpg), arai-cmv gamma globulin, eritromicine and amphotericin b was administered. on admission in our picu, trachea was mmbated, (a-a) o gradient was , paofffio~: , lung injury score > with peep level of cm hzo. she had normal fiver function. during te next days she had fever and developed ards. bal was negative. p.d. was of little efficiency. we adjusted pentanfdine, and dhpg doses for severe renal failure, with supplements after hero, sis, and at~rp.d.. during ~ next days she was afebrile, and the chest became radiologlcally normal. after ten days on menhani~al ventilation (mv.), the patient was extubated. cr. level was . rag%, (a-a) oz gradient was , and paoyfioz was , the patiem was discharged with chronic ambulatory p.d. discussion: opportunistic pneumonia is a major complicalaou in imm~romised children, specially after kidney tvansplaraafion. c m.v. infection can result at~r okt administration. in the treatment dhik} dose must be adapted to the degree of renal insu~cieney, with supplements after hemedialysis, and after pd. pneu~y~tis cann# tmeumov~ is ehemeterized by ventilafion-perfusion mistmaeh, decreased pulmonary compliance, hypoxia arld elevated (a-a) oz gradient, with diffuse interstitial infiltrates. in our ease bal was negative. although we did not find the etiology the prevoclons eombh~ation of arairmcrobiat therapy, along with m.v., and supportive measures were the most effective trealme~. conclusion: ) in patients with severe renal failure and life-threatening infections, we must co~ider drug adjuslments. ) in our patient we gave dhpg supplements at~r pd. with excett~at results, although p.d. was of little effiele~. introduction: endotracheal intubation and mechanical ventilation have become an important treatmem for many diseases accompanied by respiratory failure. with the frequent use of this treatment modality, an increasing number of complications associated with endotracheal intubation have gained clinical significance. material and methods: a transversal study was realized to find the prevalence of pulmonary aspiration with endotracheat tubes in infants and children. aspiration was assessed by applying two dyes (evans blue, er)¢rosine sodic) on the tongue and searching for the dye during suctioning in the endotracheal aspirate. the factors, that potentially have influenced the aspiration, including weight, age, sex, cause of respiratory failure, main pressure airway (map), level of consciousness, presence of swallowing and body position were evaluated. all the variables studied had their association with aspiration tested by chi-square method with relative risk considering a confidence interval of %. the results were adjusted by multivariate analysis. results: the overall prevalence of aspiration was . %. among all children who aspirated, compared to those who did not, there was a statistically significant difference in the presence of swallowing (p= . ). the odds ratio to aspiration in the presence of swallowing was . (t. - c.i. %) and the relative risk . . aspiration was not significantly affected by sex, weight, age, cause of respiratory failure, map, level of consciousness and position of the body during the ventilation. conclusion: the endotracheal intubated children frequently aspirate as intubated adults and that preventive measures are ineffective. the presence of swallowing movements is the main risk factor to aspiration of oropharingeal content in intubated patients. clinical features and shortterm outcome skling, rp gie pneumonia is the second most important cause of death in young south african children. the clinical features, intensive care course and outcome of children being ventilated for pneumonia in the developing world is unreported. aim: to describe the clinical findings, aetiology and shortterm outcome of children younger than months with pneumonia requiring ventilation. the data of all babies under the age of six months with a lower respiratory tract infection admitted to the paediatric icu for ventilation were prospectively collected over a period of months. tracheal aspirates and blood specimens were submitted for viral and bacterial cultures. results: forty-seven babies aged to days were ventilated for pneumonia. twenty-six infants had been born prematurely; t had been ventilated during the neonatal period and had bpd. the median duration of symptoms was day, the most common being cough, tachypnoea, apnoea and cyanosis. five babies ( %) died. the mean duration of ventilation was days (range - days) and of ward stay after icu discharge days (range - days), blood euttures were positive in children ( %). viruses were cultured in children ( %). conclusion: ) fifty-five percent of children below months requiring ventilation for pneumonia were premature infants, of whom % had been ventilated during the neonatal period. ) the median duration of symptoms prior to admission was day. ) ninety percent of the children survived and were discharged from hospital. ) viral pneumonia was responsible for % of the admissions. mechanical ventilation and atrial natriuretic factor release ulloa santamarfa, e, p rez navero jl, ibarra de la rosa i, espino hernladez m, velasco jabalquinto mj, frfas p rez m. picu. reina sofia children's llospital. c rdoba. spain. mechanical ventilation effects on renal function decreased diuresis and natriuresis due several factors including anf. several studies have demostrated anf released due increaasing pressure in right atrium. on the other hand, mechanical ventilation, overall peep modality, inhibits peptide release althougt cvp increased is found. this study was designed to demostrate anf stimulation is due rigth atrium stretch which be higher during mechanical ventilation instead of atrium pressure. we desing a prospective study including patients, age range months- years with congenital heart disease. all of them were admitted at pediatric intensive care unit after extracorporeal surgery and were assisted by mechanical ventilation. hemodinamic state was stabilized in all patients and nor renal neither neurological diseases were found. after hours with mechanical ventilation, plasmatic levels of anf were measurement, pvc, pericardical pressure were assessment; all patient were sedated with midazolan and paralized with neuromuscular blocking agent; mechanical ventilation technique was as follow: imv between and , tidal volume and fi o enough to mantain respiratory parameters in normal range. afterwards, at least twentyfour hours in spontaneous breathing, the study was made again in each patient. atrial stretch was assesssment according to following equation: transmural pressure= cvp -pericardial pressure. cvp were significantly higher with mechanical ventilation than when the patient was breathing by himself. ( . +__ . vs . + . mm hg; p< . ). however, transmural pressure during mechanical ventilation were lower than during spontaneous breathing ( . +__ . vs . +__ . mm hg; p < . ) equal, plasmatic anf levels were lower during mechanical ventilation ( . + . vs . + . pg/rnl; p< . ). in conclusion, anf secretion decreases during mechanical ventilation, even with cvp higher. anf release would depend on atrial stretch meassured by transmural pressure, lower in patients with mechanical ventilation and it would not depend on atrial pressure. the paediatric intensive care unit shaikh zayed hospital, lahore is an acute care area devoted to the care of critically sick children upto the age of years. in a bedded unit with limited equipment, constant care is ensured by the presence of at least one nurse aed one doctor round the clock. in this setup we have the facility to ventilate - children at one time, between sep. and dec. , out of patients admitted to icu, ( . %) were below yr of age, while ( %) were below month of age. life support was discontinued in ( . %). total mortality was ( . %), major mortality was in - month age group ( . %), and month to month ( . %). majority of the patients were of sepsis ( . %), cns disorder ( , %) followed by respiratory problems ( . %). it seems therefore that the major indicatiou for ventilation was overwhelming septicemia leading to multiple organ failure, rather than purely respiratory problems. high frequency oscillation (hfo) in the therapy for ards in pediatric patients requiring aggressive conventional mechanical ventilation (cmv) -routine or experimental mode ef pre ecmo therapy. fedora m., nekvasi~ r, vobruba v., srnsky p,, zapadlo m. dpt. critical care medicine, nicu and ecmo center, university children's hospita! brne, nicu of university hospital prague, czech republic. introduction: pediatric patients ( males, female, average age . months, average body weight , kg) with severe ards ventilated with aggressive regimen of pcv or prvc were connected to hfo (sensormedics ) as the last "rescue" therapy due to uncontrollable respiratory failure before intended ecmo. in the course of hfo of them were given no in the concentrations of - p.p.m., were subjected repeatedly to surfactant replacement therapy (alveofact). results: ecmo was needed in no patient, patients survived, patient was disconnected from the ventilator because of brain death in spite of conspicuous improvement of oxygenation and other parameters, some relevant parameters hours before and hours after starting hfo are given in table ~ in all the cases, the disconnection from hfo was carried out through the simv regimen, never directly to cpap. table : the levels of blood gases, oxygenation index (oi), aado ,map,fio and pao /fio ratio hours before and hours after starting hfo. conclusion: although none of the patient had to be subjected to pediatric ecmo, hfo should be carried out only in workplaces having the immediate possibility of using this method in the case of hfo failure. speculation: should not hfo be used ir pediatric patients with ards earlier than aggressive cmv? can hfo ce considered standard, not experimental method of therapy? refractory hypoxemia in premature patients is characterized in a persistent elevation of pulmonary vascular resistance, with right to left shunt through the ductus arteriosus and or foramen oval. we report the case of a vlbw patient (ga w, bw g) who present a severe hypoxemia related to hyaline membrane disease and a pulmonary and systemic infection to group b streptococcus, refractory to conventional ventilatory support and surfactant therapy, associated to hemodynamic failure falling in ecmo criteria used for term infants. a rescue therapy with hfov (sensor medics a) is decided at h of live, the table resume the patient's evolution before and after hfov. at w of postgestational age the patient present a fio of . with a chest x ray compatible with a cld type l at discharge no oxygen requirements was needed and actually he's doing well. conclusion: hfov, using an adequate alveolar recruitment strategy, was effective in the rescue of a severe hypoxemic respiratory failure with a rapid off of ecmo criteria entry in our vlbw premature patient, during the united nmioffs embargo ~nst yugoslavia the prevalence of the ast}nnafic ~acks in c~dldren aratsed. the mo~t common causes have beem dramm~e worsening of life standard, ecom~c disaster in global community, gr~ number of refugees from the other parts of former yugodavia. it wm obviom that mcio-ecoumnical conditions took a part in the exacerbations of previously known cldldhood asthra~, ~av~ of micro-and m~mclimaflc changes, psychosocis] and emotional cryses, lack of medics-m~nts for p~ve~on and tl~rspy of acute asflanatic attacks. about % of d-dldv~ tmslod in our picu for these year~ exp~dvncod ~vcr~ attack for the flint time iu ~jzeir lifts. it has been cu~ ~%~ children in mspir~ry picu of our hos~mt. the scut~ revere attack (more ~asn ~/o of hight clinical score) was detected in % of all children admitted with respirak~ problems. from tl~ mmlysss we exclu&d: bmncldolifis, ~i anomalies, ~eve~ i~ccqions. concerning our drug supplies (which wc~e reduced), we started our therapy by administration of oxygen, ~ta -ago~dst inhalations (but sometimes we had the solution for jet nebulizcm only for o~e inhalation per p~cnt), mwinophyllin and mefl~ylpr~ini~done in/ravenously. % of ih~ asthmatics needed repea~ doses of muinophyl~n pinch.ally, tnch.,ding the fluids. the bronchodilak)r msponm was poor ~r~cl slow, hospital stay in picu was for days and for days in other units sl~rwsvds. tim ~ of their stable condifio~ was hard at borne (or refugees camps), without p~ventkm, so they came bsvk to hospital for morn than times in % of cases, dtrdng ~e je last motlfl~s file dtustion improved, concerning tim drugs supply for prevention, and we hope that these lifc~restening conditions wouldd~ introduction: the incidence of ards is increasing as survival of critically ill patients is higher. the application of new therapeutic modalities have increased the survival rates in (ards) adult patients. objective: to study the therapeutic efficacy of new tleamlents in children with ards material and methods: a retros~ctive study was conducted from to . children with severe ards, (lung severity score > , ) (r), aged days to years, were included. the diagnosis were as follows: interstitial pneumonitis, non interstitial lung infection, with lung aspiration and with clinical sepsis. patients had different tipes of cancer and to suffer inmunodeficiency disease, the first subjects (group t) were treated with conventional measures. from october of new therapeutic modalities were introduced, including: less agressive ventilatory support, postural changes (prone to supine) in subjects, administration of corticosteroids in patients, rfitric oxide in , pe~ssive hypercapnia and administration of exogeans sarfactant in one, pao /fio , d(a-a)o , oxigenation index (oi) and the score of respirator), severity disease were similar in both groups. the two groups evolntiou was compared. results: -ten patients died, from group i and from group ii ( % v.s. : %,ns). -the evolution time, either to exitus or weaning from ventilatory support was higher in group ii ( . v.s. . days in group i, ns), -the incidence of barotrauma was observed in subjects ( . %), from group i and from ii. of these patients % expired. -during the course of the disease, ( %) patients had more than one damaged organ. only in one subjet mof was considered to be the main cause of death. the majority of the patients expired because of their respiratory disease, although, % of them met criteria of mof. -fifty percent of the subjects were infected at the time of death. stmmry: a trend toward a higher survival rate is observed in the subjects receiving the new modalifies therapeutic intervention (corticosteroides, postural changes and permissive hypercapnia). our results are not significative,probably because of the small number of subjects studied. a new doubleaurae~t two-stage et-tube (dl-ett) was desig~aed and tested in the rabbits with acute king injury under conventional mechanical ~entilation_ ventilation efficiency of dl-ett was emrrpared with that of canveniionally t~sed single lumen et-tube (sl-ett). meth~s: dl-ett was specially made out of two sl-ett. vertical crosssections at the distal end of two et-tube (td _ rmn portax) were adhered with each other to form a tracheal stage lumen wifu id . mm the two remained uncut parts of the tubes corlntithted the oval s~ge with two separate imnens. dl-ett and sl-ett were randomly applied to five adult paralyzed rabbits with acute lung injury (by . nffkg oleic acid. iv). a bird inter vetffttator (bird products corporation) was used for time-cycled pressure-limited ventilation at /min of respiratory rate, ern h of peak i_~piratory pressure, l: of ire ratio, ljmin. of flow rate and . of fich. peak inspirntory pressure, mean mrway pressure, posi ve end-expiratory pressure at tip of et-mbe and bemodynamics were measured and recorded continuously. arterial blood and expired gas were measured ~by avl blood gas analyzer) after each stabilization t.~iod of minntes. _analysis w~as by prated t test. result: dl-ett acaltety improve cos removal at all amman. pa(?oz was decreased by t . +_t. (p< . l) and physiologic dead space fraction (v~zvt) reduced by % +- . % (p< . t), compared with dl-ett. there were no significant change in arterial oxygenation. conelus|on: the double-lumen two-stage et-tabe significantly increases ventilation effmiency with simple operation in rabbits v, ith acute hmg injury, lts availability may influence future clinical management of ~ennated patient~. this ~muly was fimded by the science and technology. commiuee of beijing municipality. analis of hemostasis alterations on different coagulation cascades in children with septic shock has shown that coagulation disorder character is dependent on lung affection rate. the initial manifestation of the respiratory distress-syndrome (rds) are characterized by the obvious activation of blood thrombin potential, moderate coagulopathy and not sharply marked endoteliosis, the witlebrand's factor (wf) increase tot - %. progress in the clinical picture of "shock lung" leads to chronometric and structural hypocoagulation with potential hypercoagulation in "mix-test", high level of firbin derivative, thrombocytopenia with thrombocytopaty and the wf increase to ~ %, terminal stages of the rds, as a rule, are characterized by potential hypercoaguletion absense, depletion of at-lit and plasminogen, prevalence of antithrombin and antiaggregating activity, obvious endoteliosis (the wf to increase - %). the arteriowenous difference according to index of the thromboelastography (teg) in the rds ill-iv rates was , % less than in the - rates, disorder of lung filtering ability in severe rds is confimed also by minimal arterio-venous difference of activated euglobulin lyses (ael) in children with the rds ill-iv rates is only , %, while the patients whit rds i-i rates have the ael-activity in arterial blood , times as much than in venous blood. the use of then allows to determine the potential hypercoagulation rate, the at-ill level and fibrinogen quantity during the anticoagulant therapy and also the character of the x-factor activation and thrombocytic hemostasis. the effective therapy component of septic genesis rds in children is the controled coagulation method with the use of the individual selected heparin doses in according to desagregants, kryoplasma, proteolisis inhibitors and trombolytics. it is necessary to avoid the heparintherapy for children with the rds complicated with producting coagulopaties and termal phases of blood disseminated intravascular coagulation (dic). bronchoseopy has been used for evaluation of the potential problems of the airways and for investigation the bronchial specimens for diagnostic purposes. regent technical advances result in performing this procedure at the bedside manner and in critically ill patients. we have performed hronehoaeopy during last three years on pediatric patients with respiratory problems, in % of cases the opentube hroneh seopy was performed (for diagnostic as well as for therapeutic reasons) and collected secretions or bioptic material were examined. the indieatiuns were: acute upper respiratory problems, chronic wheezing, inspiratory strider, tracheal or bronchial bleeding, chronic eongh, retractable atelectssis, severe pulmonary infections, lymph node perforation in lung tuberculosis and soquells like bronehiectssis and fibrosis. our results were: anatomical malformations in %, mueosal oedema with chronic inflammation and thick secretions in %, easuos masses in %, granulation tissue and purulent secretions in foreign bodies and bronehieetasis in %, and only % of eases were normal finding. our exlxdenees pointed that this invasive procedure in carefully selected patients has important role in establishing the diagnosis and in the- introduction: tbg has been a useful investigation in the management of ventilator-dependent infants in our experience. one ml of contrast was hand ventilated into the respiratory tree via their nasotracheal tubes and their anatomy and dynamics demonstrated on radiological screening. case descriptions: three infants who were difficult to ventilate requiring high airway pressures, high peep and a significant oxygen requirement had tbgs. the ages ranged from to months. two cases were complicated by complex cardiac lesions. in all cases there were frequent episodes of desaturation, where hand ventilation proved difficult and various intermittent lobar collapses occurred. microlaryngobronchoscopies (mlb) performed on the infants by experienced paediatric ent surgeons failed to identify the airway problems. more than one mlb was frequently done. concern about introducing contrast into the airways of infants with limited cardiorespiratory reserve combined with an uncertainty about how much extra intbrmafion would be gained often led to a delay in investigation. when performed these fears proved groundless, the anatomy and pathology of the airways were demonstrated in full and the correct therapeutic plan started. in two cases tracheostomy and peep producing patency of bronchomalacic segments allowed weaning to low levels of ventitatory support. in one case tracheal reconstruction was undertaken and in the cardiac cases the respiratory component of the ventilatory dependence was fully assessed. at the age of months, a baby boy with a history of minor respiratory problems, was admitted to hospital with an upper airway infection and severe dyspnoea. shortly after arrival at the icu he had a total airway obstruction. after intubation there were still difficulties to establish a normal gas exchange, and he was tranferred to the regional picu. ct scan and bronchoscopy verified a congenital tracheal stenosis affecting the whole trachea except the upper mm below the vocal cords. the diameter was estimated to less than ram. an unsuccessful attempt was made to dilate the extremely rigid stenosis with a balloon. after the procedure he had a respiratory and circulatory arrest, and he was put on ecmo as a bridge to surgical correction. after stable days on ecmo, surgery was performed during ecmo with a tracheal homograft transplantation. immediately after surgery, ecmo was discontinued. a silastic dumont type stcnt was inserted inside the homogra~, and a nasotracheal tube was placed inside the stent for assisted intermittent mechanical ventilation. repeated bronchoscopies were performed to remove granulation tissue and secretions. at months of age, the stem was removed with an endoscopic procedure. however, the trachea was still soft and collapsable, and another silicon stent was placed inside the trachea for another months period, after removal he had some respiratory problems and he was treated with nebulized salbutamol, mcemic epinephrine and steroids. he was discharged from the hospital at months of age and his condition is now stable. this is the first procedure of its kind in sweden. it was accomplished by international and multidisciplinary collaboration. ecmo may be a bridge to corrective surgery and long time stenting may be necessary in the postoperative period. post mtubation laryngitis ( pil ) is still a frequent complication, occurmg in l - % of intubated patients. inhaled racemic epinephrine has for long been used as an accepted therapy, but this drug is not always available. the authors undertook a randomized, double-blind, placebo-controlled trial to determine the efficacy of inhaled l-epinephrine(le) in the treatment of plu in the period between july/ and may/ , patients were submitted to endotracheal intubation for ventilatory support. atter the extubation procedure patients were considered for enrollement if they met the following criteria: clinical signs of laryngeal estridor and a downes and rafaelly score for upper respiratory obstruction equal to or higher than patients with primary upper respiratory disease were excluded all patients enrolled reeieved either inhaled l-epinephrine % or normal saline. dexametasene ( , mg/kg/day) was given to all patients in both groups. after inhalations, au patients were monitored for a period of - minutes and monitoring included cardiac and respiratory rate, mean arterial blood pressure, arterial blood gases and the dowries and rafaelly score. statistical analysis included, qui-square with the fisher correction test and the z-test for paired variables. thirty eight patients ( , % ) met the criteria for enrollment, to the le group and to the placebo group.there were no significant differences in both groups in regard to age, sex, initial score ( , x , ) and endotracheal tube diameter. the period of ventilatory support and tracheal intubation was significantly higher in the le group ( , x , , p = , ). the follow-up score showed a significant drop only at minutes after the inhalations (p = , ). re-intubation due to laryngitis, occured in patient of the le group and in of the placebo group with no statistical sxgnificance (p = , ). no difference was observed on the monitored hemodynamic variables during the minutes, except for the mean arterial pressure at minutes, being heighar on the placebo group (p = , ). we concluded that, although the l-epinephrine group showed a trend in better scores post-inhalation and fewer re-intubations due to laryngitis, the results were not statistically significant. we especulate that the period of intubation may have affected our results. similarlly there were no differences in the incidence of adverse effects between both groups. objectives:to evaluate the complications of endotracheal intubation in children with upper airway obstruction due to epiglottitis or croup. methodes: during a year period ( - ) all patients with epiglottifis or croup were reviewed to determine the complications of endotracheal intubation, especially upper airway obstruction due to granulomas. results: patients were reviewed. in children (mean age . years) with epiglottitis the mean duration of intubation was . days ( - ). no complications were seen. in patients (mean age . years) with croup the mean duration of intubation until the first extubation was . days ( - days). elective extubation was performed if an airleak was present or after days without airleak but in the absence of fever and obvious secretion. reintubation was not necessary in children ( . %). in this group the mean duration of intubation was . days ( - ). in patients ( . %) reintubation was necessary because of severe upper airway obstruction due to granulomas. mean duration of intubation until the first extubation was . days ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . there seems to be a difference in duration of intubation between these two groups with croup, however it is not significant (p > . ). all the patients with granulomas could be successfully extubated after microlaryngeal surgery, with a mean intubation period of . days ( - ). revealed no complications, where as endotracheal intubation in children suffering from croup showed a high incidence ( . %) of granulomas. however laryngeal steepsis and other serious complications were not sesn~ patients ( days averagely] was obviously seen in ~he peak =one of fl, f resonance and in the zone of high freq,-~ncy :r, ~;~e composition while cases( day~ average;y] :~bowed no abnormality both clinically and isryngoscopica!~y. / patients with catheter placement for more than week~ end p~tie,~ts for less than weeks had t;~ryngeal abnormal change in their larynges,abnormal changes of sound spectrogram were all seen in patients with placement for mope than weeks. our data suggest= ca] the complication of endotracheal intubation was increases with increasing length of time of catheter placsm. entjbut aeriuoa complication is rare i (b] the time limit of pernasal endotraoheal catheter placement is weeks within which the procedure is • comparatively safe and effective means for maintaining e tong term artificial airway. in a -year period ( ) ( ) ( ) ( ) ( ) ( ) ( ) we diagnosed tbm as an apparent dilatation of the trachea and main bronchi ih four premature infants on continued mv for respiratory distress syndrome (rds). the infants were three boys and one girl with gestational age (ga) - weeks and body weight (bw) - g. mv was provided by bourns cub time-cycled and pressure-limited ventilator to attain normal gas tensions. no jet ventilation was used. chest radiographs were reviewed for a complete evaluation, and for the evaluation of the airway. after the intial subjective diagnosis of tbm, the width of the tracheal and main bronchial air column was measured at the lower level of the first and the third thoracic vertebal body it , t ) and near the carina; the width of the main bronchi below the carina was also measured. in all infants, tbm became apparent close to the lh day, that is, after - weeks of mv. therefore, for the time period from birth to the th day the following ventilatory parameters were reviewed and analyzed: ( ) the percentage of total ventilation time when more than % o concentration was required, ( ) the peak inspiratory pressure, ( ) the positive end-expiratory pressure, and ( ) the duration of high frequency ventilation ( - breaths per minute). also noted were the apgar scores ( and min after birth), the duration of hypotension (systolic bp below mmhg) and circulatory instability, the presence of systemic or tracheal conatal or later infection, the duration of mv, and the final clinical outcome. the records were also reviewed for other possible pertinent data. rigid respiratory endoscopy in children fraga j, amant a s, piva j, nogueira a, palombini b. introduction: the respiratory endoscopy is an important procedure to diagnose and treat many airway's diseases in children. although have had advances in radiologic investigation exams and pulmonary function tests, the direct anatomic visualization of airway is important to the management of many respiratory problems. objective: evaluation the respiratory endoscopies performed with a rigid bronchoscope in a pediatric reference hospital. material and methods: we study the records of all children that were submitted to respiratory endoscopy under general anesthesia from march to march . age, sex, clinical to indicate the procedure, diagnosis and complications of endoscopy were registered. results: three hundred and fifty six respiratory endoscopies were performed. the most common indications for endoscopy were strider ( %), suspected foreign body ( %), atelectasis ( %) and difficult tracheal extubation ( %). the most frequent diagnosis were laryngomalacia ( %) and subglottic stenosis ( %) in the glottic and subglottic areas, and foreign body ( %) and tracheomalacia ( %) in the tracheobronchial area. normal endoscopy was performed in ( %) of the children. only three slight complications of the endoscopy were observed. two patients presented bradycardia during the exam, and the third need tracheal intubation due to post-endoscopic subglottic edema. conclusion: the rigid endoscopy in children is efficient and has no serious complications. near drowning; indicators of acute and long term prognosis bernardien t.mj. thunnissen t, reinoud j.b.j. gemke , loes veenhuizer?, krijn haasnoot , a.johannes van vugh department of pediatrics, ~wilhelmina children's hospital, utrecht, sophia hospital, zwolle, and ~free university hospital, amsterdam, the netherlands. in this retrospective study factors that affect short and long term prognosis after submersion were analysed. all patients that were admitted to a tertiary pediatric icu between january i, and january i, were included. of patients, aged - years, died in the icu, one after hospital discharge. survivors and non-survivors showed significant differences with respect to central temperature, pupillary reactions, arterial ph, pediatric risk of mortality (prism) score and therapeutic intervention scoring system (tiss) upon admission (p < . ). non-survivors more frequently required mechanical ventilation, bicarbonate administration and active reheating. ards was seen in patients ( %), invariably within hours after admission. no patients with cardiac arrest on" admission snrvived without sequelae. hypothermia appeared to have no protective effect on hypoxic damage. survivors with persistent sequelae _> months after discharge had significantly higher prism and t ss scores (mean and , respectively) than those with complete recovery (mean and , respectively). long term cognitive problems were present in / survivors ( %) and emotional disturbances in / ( %). in conclusion, a concise number of clinical and laboratory parameters, representing acute severity of illness, are important prognostic indicators for survival and health status of children after submersion. there were ( %) bronchoscopies, and ( %) were oesophagoscopies.the average age was , years for bronchoscopies, and years for oesophagoscopies. the outcome of the patients was good. no complications were observed. extraction is recomended in every symptomatic patient. orphenadrine is an anticholinergic drug mainly used to decrease symptoms of parkinson disease. orphenadrine has a peripheral and central effect and overdose can result in athetoid movements, convulsions, cyanosis, coma, arrhythmias, shock and cardiac arrest. physostigmine is a specific antagonist of the peripheral and central effects and can be a useful antidote. we report the case of a two and a half year old female who was transfered to our icu for general convulsions. the little girl had, three hours before admission, accidently ingested rag of orphenadrinehydrochlodde (disipal®), which was her grandmothers anti-parkinson medication. three hours after ingestion she presented neurological signs: confusion, unstable walking, and periods of aggression. generalized tonic-clonic seizures appeared who were rebel to administration of multiple anti epileptica but ceased after iv administration of diazepam and endotracheal intubation and ventilation. an episode of ventdcular tachycardia responded well to the iv administration of tidocaine. the levels of orphenaddne in the serum were high at admission ( pg/l) and were present in the blood up to hours after ingestion. high serum levels are, in the literature, associated to a high mortality rate. physostigmine was administered three times at a . mg/kg dose in the first hours. we decribe the noted effects of physostigmine on the different symptoms. the patient survived and could leave the icu after one week. in conclusion: orphenadrine poisoning is a very complicated medical problem associated with high mortality. in severe intoxication, the benefit of physostigmine more than counterbalances its side effects. objective: to define the optimal volume of dilution for endotracheal (et) administration of epinephrine (epi) design: prospective, randomized, laboratory comparison of four different volumes of dilution of endotracheal epinephrine ( . , , and ml of saline) setting large animal research facility ofa universi~ medical center subjects and interventions: epinephrine ( . mg/kg) diluted with four different volumes ( , . . and i rot) of normal saline was injected into the et tube of five anesthehzed dogs. each dog served as its own control and received all four volumes in different sequences at ieast one week apart. arterial blood samples for plasma epinephrine concentration and blood gases.were collected before and . , . . . _ . . , . . , . , . , and minutes after drug administration. heart rate and arterial blood pressure were continuously monitored. measurements and main results: higher volumes of diluent ( and i ml) caused a significant decrease of pao , from :!: tort to ±i torr, compared to the tower volumes of diluent ( and ml), from ± torr tu +_ torr (p< . ). these effects persisted for over minutes. mean plasma epinephrine concentrations significantly increased within seconds following administration for all the volumes of diluent. mean plasma epinephrine concentrations, maximal epinephrine concentration (cmax), and the coefficient of absorption (ka) were higher in the ml and ml groups. the time interval to reach maximal concentration (tmax) was shorter in the ml and ml groups. yet these results were not significantly different. heart rate. systolic and diastolic blood pressures did not differ significantly between the groups throughout the study. conclusions: dilution of endotracheal epinephrine into a ml volume with saline optimizes drug uptake and delivery, without adversely affecting oxygenation and ventilation. the aetiology and outcome of paediatric out-of-hospital cardiac arrest was studied during a -year period in southern finland served by physician staffed emergency care units. the files of prehospital patients less than years old without palpable pulse and spontaneous respiration were analysed retrospectively. fifty patients were declared dead on the scene (dos) and resuscitation (cpr) was initiated in patients. the sudden infant death syndrome was the most common cause of arrest ( %) in the dos patients as well as in patients receiving cpr ( %). asystole was the initial cardiac rhythm in % of the patients in whom cpr was attempted. eight of the hospitalised patients were discharged, of them with mild or no disability, with moderate disability and one in vegetative state. in multivariate analysis the short duration of cpr (< minutes) was the only factor significantly associated with better survival. due to various aetiologies the survival rate from prehospital paediatric cardiac arrest is quite low. on the other hand, hypothermic near-drowning victims seem to have a relatively good prognosis. duration of cpr less than minutes was the best predictor of intact survival, our study supports the previous findings of the importance of early and effective resuscitation efforts for establishing ventilation and perfusion on the scene. in our system well trained physician staffed emergency care units are able to provide immediate and effective als on the scene. on the other hand, these units also appear to be able to refrain from resuscitation when the prognosis is pessimistic. objective: to assess the normal ,gastric intramucosal ph ~hi) by tonometry in healthy children patients and methods: twelve healthy children ( males and females) with age rmaged from months to years scheduled for minor plastic or urologic surgery. children were previously medicated with midazolam ( . mg/kg) and atropine ( . mg~) both i.m.. anaesthetic induction was standardized with -n ( %) administered via facial mask and increased halotane concentrations (up to %). all patients got an endotraeheal tube after iv. administration of femanile ( mcg:jkg) and vecuronium ( . mg/kg) or suxametonio ( mg/kg), pmaesthesia was maintained with o -n ( - %) and isofluorane ( . - %). during surgery, children needed mechanical ventilation and the others maintained spontaneous breathing. ekg, heart rate, blood pressure, and pulse oximetry were moniterized. after anaesthesia, a sigmoid tenometry catheter (tonometrics, inc.) was inserted in the stomach of the patients by direct visualization with laryngoscope and magyll clamps. children were all maintained normoventilated and with normal cardiorespiratery variables. cadet's balloon was £~led with . ml of saline. thirty minutes after the insertion rrd was extracted and rejected, just afterwards the remanent . ml was extracted and immediately analyzed. simultaneously an arterial gasometry by puncture was performed. gastric phi was calculated by the henderson-hasselbalch's equation using the pco obtained from the tenometry catheter and the bicarbonate value obtained from the arterial gasometry. results: average gastric phi was . -i- . , range ( . - . ). objective: demons~ating intramucesai ph (phi) alterations during transport of patients from operative room to pediatric intensive care unit (picu), material and methods: phi measurements were performed with gastric tonometer catheter in t patients undergoing cardiac surgery with cardiopulmona d" bypass (cpb), there was mate and female, the average age = yl ra, average weight = , kg, average time of cpb = rain. the measurements were made at the end of the surged' and when the patients had arrived in the picu statistical aualysis: average and ~andart deviation and test "t" student. objetive: to asses the efficacy of gastric iatramucosad ptt (phi) and arterial lactate levels to evaluate splacalc tissular perfusion in an experimental model of intestinal ischemia. suneets ~nd methods: twelve piglets weights t - kgs. undergoing orthot~ie liver trasplantation. the intestinal ischemia was induced by aortic damping. tonometry catheter (tonometrics inc.) w~s placed in the stomach after artaesthesia and ot intubation. phi ~s determined times and lactate levels was determined fi times in stages: i) pre-ae~hepatic stage (twice: before surgery and before aortic clamping ); ii) end anhepatic stage (only phi): iii) reperfusion stage (a , , and minutes). the phi was derived from application of the henderson-hassdbach formula using the pco value from the tonometer and the arterial bic~rbonate. all pipets received raaitidiila before sttrgery. values of phi above , and lactate levels between and mg/dl were considered nortrm. the results were statistically anaj.izated with anova and bonferroni tests. results: the phi was normal on pre anhepatic stage (> , ) and lactate levels were slightly increased ( , +_ , and , ± , mg/dl ns) . in relalion to we-anhepatics values, phi decreased signncatly at the mid of anhevatic stage ( , _+ , vs , _+ , p< , ), phi remain low in stage iii, at rain ( , + , p< , ) and min(g -+o, p< , ). arterial lactate levels increased significatly in relation to levels in stage i, at rain ( , _+ , p< ,o ) arid rain ( , ± , p< , ) of reperfusion stage. there is a slight improvement on phi and lactate ievels at and t rain althought the differences did not reach significance. cnmments: phi and arterial lactate levels propperly reflect hypoperfusion on the experimental model of acute intestinal isdlemia. b~kground : the paediatrie gallbladder diseases generally described are calculous ¢hol~tstitis, cystic duct obstruction, congenital anomaly of the biliary tract, and inflammation. in the neonatal period, noulithogenie gallbladder disease could be also due to erythroblastosis or hyperalimentation. obieetive : we describe an other type of disease affecting the gallbladder in neonates thought to be related to their vascular vulnerability. methods : four patients with abnormal gallbladder ultrasound not related to classical observations were included. we have studied and reviewed the biological and clinical data, the ultrasound findings and their evolutions. results : four patients, to ~.k-old neonates ~ffth a birthweight be~,een , and , kg, were intubated and under total parenteral nutrition for to days. none of them were symptomatic on repeated clinical evaluations. one newborn developped hypotensien on umbilical bleeding at hours of life. in two cases, signs of cholestasis were discovered : the total bilirubin level has risen to mg/dl; the direct bilirubin level was , mg/dl while the urina were dark and the ~o~,ls :mcolour~. the c~mplct~ ~crology as a!! the culvare~ remained negative. the ultrasound explorations were atypical : in the four eases, an initial increasing broad and thickness of the wall of the gallbladder with an hyperecbogenie inside content, which was not sludge, was discovered. in three eases the images resolved in ten to fifteen days. in one ease, an asymptomatie thrombosis of the vena portu which remained patent was discovered. in this case, at one month, the ultrasound showed images encountered in chronic ebolecystitis and, at one year, the gallbladder appeared atrophic. none of them underwent surgery. conelusiou : the gallbladder diseases are multifactorial. besides the prematurity, the infections, the total parenteral nutrition, the premature neonate is exposed to vascular vulnerability affecting also the gallbladder and this may explain our findings. progress in prognosis of pts with b-nhl had followed the use of multimodality chemotherapy (ct). with the prolonged survival, there are comlications due to myetosupression & desease process. the syndrome of neutropenic enterocolitis (ne) is one of the ominous problems because ofpts increased susceptibility to infection & overwhelming sepsis. this material included neutropenic pts ( - years) with the stages iil& iv of b-nhl who were treated with the modifired bfm- (mtx g/m in -h inf.); males, females. seventeen episodes of ne were observed & only after the first courses of ct ( of after tst, %; of after nd, %). the symptoms existed to days. wbc ranged from to in l~tl (median, ). the first signs of ne were directly correlated to the beginning of the neutropenia & the recovery of neutrophils led to the disappearance of abdominal recovery of neutrophils led to the disappearance of abdominal pain. the conservative treatment included gastrointestinal tract decompression, broad spectrum antibiotics initially, volume & electrolyte substitution, nutritional support, correction of acid-base balance, symptomatic treatment. sixteen pts were treated nonoperatively, died. on autopsy the transmural bowel necrosis due to thrombosis of branches of a.mes.sup, was found. the bowel perforation occurred in one patient, he was undergone laparotomy & hemicolonectomy & survived. we conclude that ne is a frequent complication in neutropenic pts with the st. lii& iv of b-nhl. it occurs after the induction courses of ct. close observation by surgeons, oncologists & pediatric intensivists is mandatory. conservative treatment is effective & more preferable until leucopenia resolves. operation is necessary only for those.with perforation. near infrared spectroscopy as a tool for evaluation of intestinal perfusionpresentation of an animal model. c. scheibenpflug, p. buxbaum and a.m. rokitansky the recent development of and investigations in the so called near infrared spectroscopy ( nirs --transcutanous emission and simultaneous registration of intensity of spectralcolours depending upon modulations of tissue perfusion ) enable physicians to measure and qualify organ perfusion and nowadays is mainly used to control cerebral as well as skeleton muscular blood flow in trauma patients at intensive care units ( icu ). today intestinal perfusion, hypoperfusion , cell damage caused by reperfusion injury, bacterial and toxin translocation are serious problems in critically ill patients at an icu. paediatric intensive care physicians put major concern on intestinal perfusion, which for. instance gains more and more importance, especially in the neonatal period for example as an etiologic factor for necrotizing enterocolitis. we established an animal model, in which we measured intestinal perfusion by nirs under various invasive and noninvasive conditions. methods and results will be referred. for preliminary conclusion we propose near infrared spectroscopy ( nirs ) also as a potent diagnostic tool to determine early intestinal malperfusion in order to prevent lethal outcome. fm'ther investigations in animals as well in paediatric iritensive care patients should be done to estimate our efforts. introduction: following the acute phase of necrotising enterocolitis (nec) starvation of the gut for a period up to weeks is a generally accepted treatment modality in many centres. objective criteria to refeed these patients are hardly available. recently the double sugar test has become available as a parameter for (ab)normal gut permeability ~' . aim of the study: to evaluate the changes in permeability of the small bowel in patients with nec and controls before introduction of enteral feeding. methods: a lactulose! rbarrmose (i/r) test was performed in two groups. group was studied - times within a -week period of starvation (n= , mean gest. age , range - weeks). in group seven different control patients were studied (mean gest.age , range - weeks). the test was performed by giving a patient after at least a hour fast ml/kg bodyweight l/r solution and determination of the /r ratio in a -hour urine sample by chromatography. results: objective: to evaluate the prognostic factors in the response to nitric oxide (no) in children with acute respirator/ distress syndrome (ards) and/or pulmonary hypertension (pht). patients and methods: critically ill children received no inhaled for ands and/or pht treatment. patient before and after cardiac surgery ( cardiac transplants), with bronchopneu~onia, multiple trauma, sepsis and cardiorespiratory arrest. patients showed /j~ds and pht, in with associated ards. we analyzed age, sex, diagnosis, pao , pa /fi , oxygenation index, pht, shock, and sepsis as prognostic factors and response factors to n . results : after no administration oxygenation did not improve in patients ( . %) and pht did not diminishe in one children ( %). patients survived ( %), / ( . % with /d%ds) and / ( %) with pht. the four patients with isolated pht survived , and the patients with pht and ards dead. patients after cardiac surgery presented less mortality ( . %) than the rest of patients ( . %). patients with shock presented higher mortality ( . %) than the rest of patients ( . %). there are no differences in response to no in respect of sex, age, diagnosis, shock, and sepsis. survivors showed higher increase of pao /fi . ± . to no than non-survivors . ± . (n.s). patients with pht showed higher increase in pa /fi to no administration ( ± . ) than patients with ards ( . ± . ), (n.s), but patients with ards showed a higher increase in !, ± . , than patients with pht . ± (p < . ). patients with pa /fi < i showed less increase in pa /fi , . ± . , than the rest of patients . ± . (n.s) conclusions: i. mortality of isolated pht treated with no is less than patients with ap~s. patients with shock and those with pht and ards showed higher mortality. . we have not found any clinical or analytical factor to predict clinical response to no administration. patients showed ards, and severe pht after cardiovascular surgery, in with associated ards. we registered respiratory assistance, blood gases, pao /fi , the oxygenation index (oil, and mean pulmonary pressure/ mean systemic pressure (pap/sap) before and after no inhalation. we measured continuous concentration of no and no by electrochemical method (noxbox, bedfont, airliquide). results: no administration improved oxygenation mean pao from ± tm~g to i ± ~g (p < . ), mean pa /fi fr<xa ~ to £± (p < . )and i from ± to ± (p < . ). patients did not improved with no administration. the oxygenation improved in the first five minutes and the best oxygenation was achieved with - ppm. there is no significant change in pac . pap/sap diminished from ± % to ± % ( p< . ), in one patient there is no response. the no administration was maintained between ~ minutes to days. the effect of no on pulmonary pressure and oxygenation was maintained without change during all the time it was administrated. n concentration were always less than ppm y metahemoglobinaemia less than . %. there are no side effects secondary to no administration. patients survived ( %). concluaions:l.administration of no improves oxygenation in children with ~/eds and diminishes pht after cardiac surgery. .no effect is fast and maintained during the evolution. high frequency oscillatory ventilation in combination with inhaled nitric oxide g~thberg sylvia, md, edberg k e md, phi), dept of paediatric intensive care, children's hospital, s- g teborg, sweden background: for man), years severely sick infants with congenital heart malformations or acute lung disorders have died due to pulmonary hypertension, hypoxia and multiple organ failure. today there are several therapeutic facilities coming up for these infants. ecmo progranunes have been introduced in severul centres as well as improved venlllatory techniques, as high frequency oscillatory ventilatior~ (hfov), which provides adequate ventilation with less risk for hmg injury.. in , the endothelium derived relaxing factor was identified as nitric oxide (no), and extensive studies have sho~-a that inhaled no reduces pnimonm o vascular resistance and improves oxygenation in hypoxic infants with pulmona~ hypertension. material.. from july to january , we have treated severely hypoxic children with combined ttfov-no. seven were newborn, with cdh, one with mas, one with irds, one with paediatric ards due to rsv infection and one with poor oxygenation atler open heart surgery. were between month and years and all had ards of differentorigin but one who was hypoxic after open heart surgery. method: hfov was given by means of sensormedics a oscillatory ventilator to patients, and dr~ger babylog was used in one case. mean airway pwssures varied from - cm h . oscillatory pressures varied from - cm h and ventilatory rates from - hz. fi o varied from , - , . no was administered by nomius classic. no and no was measured in the inspiratory limb with an electrcchemieal device. noconcentration varied from to ppm and the no -concentration between - , ppm. methemoglobin was never more than , % (average , ). duration of treatment varied from to days. combined trealment with hfov and inhaled no improved oxygenation and carbon dioxide elimination in out of treated childretl patients died, due to their underlying congenital heart disease, one of asphyxia due to rsv-infection and one of cdh with progressive hypoxia and multiple organ failure. combined treatment with hfov and inhaled no improves oxygenation in severely hypoxic children. treatment should be slarted early to reduce the risk for chronic lung injury following barotranma and high oxygen concentrations. we speculate that the combined hfov-no may reduce the use of ecmo, and that it may improve outcome in ceotres where ecmo programmes are not introduced. apart from its vasodilative properties nitric oxide appears to act also in physiologic immune defense. intracetlular concentrations of no synthesized by macrophages are in the range of above those produced by vascular endothelium. we investigated the bacteriostatic effect of nitric oxide at concentrations used during inhaled therapy for pulmonary vasodilation in neonates. ten different strains of five species were used (staph. anrens, staph. epidermidis, strop. group b [gbs], e. coil and pseudomenas aeruginosa), which are ~e most often tracheally isolated bacteria in mechanically ventilated premature infants and neonates. we compared bacterial growth of cultures applying three different concentrations of nitric oxide ( ppm. ppm, ppm) m the growth of the same strains in room air for a duration of hours. no bacteriostatic effect was demonstrable at no concentratioes of ppm. e. coil showed decreased bacterial growth at ppm and ppm, however without reaching statistical significance (p= . ). at nitric oxide concentrations of i ppm staph. epidermidis and gbs grew significantly less as compared to colonies of the same strains in room air. no effects were found regarding the growth of staph. aureus and pseudomonas aneruginosa. we conclude that nitric oxide has a selective bacteriostatic effect on some of the most often trachealb, isolated bacteria in premature infants and neonates. this effect appears to be dose-dependant and occurs in the upper range of dosages used with inhaled no therapy. further research is required in order to examine the mechanisms of action as well as specific interactions between different strains of bacteria and nitric oxide. the no,box monitor (bedfont, kent, uk) is used to monitor nitric oxide (no) and nitdc dioxide (no ) during no administration in ventilated neonates. according to the manufacturers specifications the monitor can be applied in cases where airway pressures range from to mbar. we investigated in-vitro the accuracy of the no monitor in a range of ventilatory conditions. using a dr~ger babylog we ventilated an artificial lung. no was administered { ppm no in % nitrogen) with a mass flow controller to the inspiratory tube, at a distance of cm from the y-piece. the no target value was set to ppm. the ventilator was operated in both cpap and ippv modes at different settings. the no sampling tube was placed on two different locations; in the inspiratory tube dose to the y-piece; in the expiratory tube haft way between the y-piece and the ventilator. the no-monitor was ca)ibrated with ppm no at mbar. fig. and show the sensitivity of the no-monitor for respectively static pressure (cpap) and pulsatile pressure (ippv) with the sampling tube located in the inspiratory tube. fig. shows the resur for pulsatile pressure with sampling in the expiratory tube. we conclude that the accuracy of the bedfont no-monitor is dependent upon airway pressure and sampling site, the latter possibly caused by incomplete mixing, pressure and no are linearly related when sampling occured further away from the administration site. based on this study we suggest to place the sampling tube in the expiratory tube. during neonatal care the clinician should be aware of varying inaccuracies depending on the respiratory pressures. hypothesis: availability of therapy with inhaled nitric oxide (ino) decreases ecmo use in patients referred .to a tertiary care hospital for treatment of respiratory, failure unresponsive to conventional therapies. background: at children's hospital of wisconsin (chw) treatment for patients who have failed conventional treatment for respiratory failure, sometimes called "rescue" therapy, has included ecmo since and high frequency oscillatory ventilation since . ino has been in experimental use at chw since may of i and has resulted in the perception of a decreased need for ecmo therapy in those patients referred for rescue therapy. this study was designed to tbst the validity of that perception. study type: retrospective cohort. methods: the medical records of all patients referred to chw from /lj to / / for rescue therapy for severe respiratory failure were included in the chart review. data were collected regarding diagnoses, illness severity, hospital course including interventions and complications, and outcomes. exclusion criteria w~ere the finding of structural heart disease or congenital diaphragmatic hernia as the basis for hypoxemia. qualification for ino treatment included an a-ado - or oi ~- for two hours or -> for twelve hours and echocardiographic demonstration of persistent pulmonary hypertension of the newborn. patients were classified into two groups based on the availability of ino at the time of their hospitalization. results: in the time period of the study, patients were referred for possible ecmo therapy. twelve patients greater than weeks old, with congenital diaphragmatic hernia and with congenital heart disease were excluded from this analysis, leaving patients for study, ino availability reduced ecmo use from of ( %) patients in the ~ino unavailable" group to out of ( . %) patients in the "ino available" group, p=& by fisher's exact test. the fact that the two groups were composed of patients of similar severity of illness is reflected by comparable rates of ecmo and ino rescue therapy ( % vs. %). conclusion: by providing an alternative rescue therapy, ino has reduced the need for ecmo in this group of neonates referred for respiratory failure. introduction: true hepatnrenal syndrome (his) is defined an acute renal failure {arf) in the presence of severe liver disease without other known causes of renal failure. hrs is frequently seen in the course of hepatic cirrhosis• in children, cirrhosis is rare; however, arf can be seen in combination with aseites and liver dysfunction• we describe patients with hepatic dysfunction and aseites in combination with ar~ and abnormal sodium-water handling, leading to the diagnosis of hrs. pathophysiology: three factors are considered in the pathogenesis of hr~: i) hepatic dysfunction, ) deranged hemodynamics, including abnormal blood pressure, reduced effective arterial blood volume and abnormal blood flew distribution, and ) neuro-humoral dysrsgulatiom, including elevated levels of aldosteron, renin, angiotensin-ll, ade, vasodilatim nitric oxide and vasoconstrictor peptide endothelin-l. the main pathogenetic feature is decreased cortical renal blood flow, decrease of glomerulur filtration rate (gfr), vastly increased sodium retention, uliguria, and azotemia. treatment: therapy is based on counteracting sodium and fluid retention by highdose aldosteron antagonists and loop diuretics, improving renal perfusion by lowdose dopamin, and strict restriction of fluid and sodium. interventions as paracenteals of aacites or n peritoneo-systemic shunt are associated with high morbidity and poor outcome in children. reversal of hem by conservative measures can only be attained at early stages of hrl liver transplantation is the only definitive treatment that can reverse ere at advanced stages. patients: the described patients developed severe ascites with insidious renal dysfunction and abnormal sodium-water handling during admission at picu and fullfilled clinical criteria fur hrs. treated according to the cited principles, all patients showed improvement of gfr, with increased natriuresis and gradual decrease of ascites. eventually, renal function normalised completly. conclusion: ere deserves greater recogmitimn in the picu population; diagnosis can be suspected on clinical criteria. with this increased awareness, therapy tun be instituted at an early phase, with better prospects for recovery. positive outcome of hem depends on early recognition of the clinical picture, understanding of the pathophysiology, and early institution of consistent treatment. mtx is an antimetatxflite widely used as chemotherapeutic agents. high dose ivitx (i to ~m ) administered as a prolonged intravenous infusion (over - hours), is often used to treat malignant paediatric diseases. major complications of this treatment are myelosuppression, orointestinal mucositis, dermatitis and impairment of anal function. we report two cases of mtx overd~age occurred in two children ( -year-old. month-old) t~ted for acute lymphoblastic leukaemia. they were treated by cavh and the mtx bhk~d levels rapidly decreasedavoiding multisystemic involvement. establishment of alkaline diuresis and monitoring of plasma mtx levels during treatment is essential to prevent nephrotoxicity. however. leuco',cnn rescue may not prevent the development of potentially lethal toxicities in patients with mtx concentrations persistantl} exceeding t mm. in theses cases, em'ly treatment of mtx intoxication may pm~cnt myelosuppression and reducerenal damage. the goal is to lower the concentration to below mmoll, at which time rescue agents aleme would be expected to be cllcctive. respective indications of these remo',at mctny.:is are still discussed : hacmt~ialysis t~ eharc(~l haemoperfusion should be prolx',sed for massive and acute intoxication. however, rebound has been reported after combined hcmodialysis and hemoperfusion. exchange transfusion may be proposed as a treatment for prolonged and moderate intoxication. peritoneal dialysis is an incflbedve method for remo~ al of mtx. cavh was used in our icu. cavh is a simple method for blood purification and n':dy iluid control. use of cavh was never be reported in this indication to our knowledge. simplicity, rap~d application and gco.l clinical tolerance are the main advantages of this technique. the technique presents ~peclal advantages in terms of low priming volume of extracorporeal circuit, low blood flow, low rate heparinisation. our results show a decreaseof plasma mtx concentration and a rapid reduction of halfqite of elimination (t hours over the period of cavh). moreover, we didn't delec~d rebound after stopping prc,xedure. small size of the i:ratients may present sometime special problems, but these technical problems can be overcome, no severe complication (needing, inlection) were observed during filtration, in summary, aggressive intravenous fluid hydration and alkaliniaation of the urine coupled with careful monitoring of renal function and plasma mtx concentrations during and al'tcr infusion along with lem~overin rescue has reduced the inndcace of life-threatening toxicity after highdose mtx. however, some mtx inu>xication still occurred, leading to se~em toxicity, particularly nephrotoxicity. in these cases, we think that cavh (or cavhd) is a reliable, rapid method without rcix~und increase in plasma mtx concentration or important adverses effects compared to other procedure removal. gouyon jb, germain jf, semama d, pr vot a, desgres j preliminary limited data suggested that hemofiltration and hemodiafiltration may be valuable in some neonates with decompensation of maple syrup urine disease (msud). venovenous hemofiltration (vvhf) and hemodiafiltration (vvhdf) were performed with a new neonatal hemo(dia)filter (miniflow , hospal) on anesthetized rabbits infused with branched-chain amino acids (leucine, isoleucine and valine) and c~-keto-isocaproate. the bcaa and aketo-isocaproate blood levels were close to those previously observed in neonates with msud when extracorporeal blood purification was required. vvhf and vvhdf performances were assessed with two different blood flows (qb = . and . ml/min). vvhdf was performed with dialysate flow rates (qd = , , . , . and . l/h). thus, each animal was submitted to successive procedures. within each studied period, clearances of the bcaa were strictly similar. bcaa clearances obtained by vvhf were similar to ultrafiltrate rates (respectively, . - . and . - . ml/min at high and low qb ; p < . ). the ~x-keto-isocaproate clearances obtained by vvhf were . - . and . - . ml/min at low and high qb (not significantly different). whatever qd value, the vvhdf procedures always allowed higher bcaa and c~-keto-isocaproate clearances as compared with the corresponding v'~hf period with similar qb. bcaa clearances obtained by vvhdf with a . l/h dialysate flow, were . - . mljmin and . - . ml/min at iow and high qb, respectively. the concurrent a-keto-isocaproate clearances were . -,. , ml/min and . _+ , ml/min. at both qb regimens, bcaa clearances provided by vvhdf were markedly higher than values previously obtained with peritoneal dialysis in human neonates with msud. the management of renal failure in the newborn is difficult. when dialysis is instituted peritoneal dialysis (pd) is usually the technique of choice. this is can be problematic and impossible in some patients with pre-existing intra-abdominal pathology. continuous arterio-venous haemofiltration (cavh) has been described in infants but sick preterm infants are not able to support the circuit. i have devised a means of having pumped haemofiltration in small/preterm infants (phis/pi) and describe its use in nine patients ranging in size from to gms for periods of to days. vascular access was achieved through or guage cannulae in either a peripheral artery and a central vein or through two central veins. blood was pumped out using an ivac infusion pump and through a gambro fh haemofilter. a second ivac pump was used to remove haemofiltrate from the filter and a third to infuse replacement solution. removal rate was set to give a clearance of mls/min/ . sq.m and blood flow rate set to between and times the removal rate. heparin was infused into the circuit to prevent clotting of the filter. biochemical and fluid balance control was achieved in all infants. guaranteed fluid removal allowed the administration of full nutritional support. four patients died when treatment was withdrawn because of an untreatable underlying problem. one recovered renal function but died some weeks later from unrelated problems, three survived and recovered renal function and one patient is still on treatment. this system allows a secure means of achieving fluid and electrolyte control in the preterm infant. the use of this technique may allow haemofiltration to become as applicable to preterm infants as it is to older children and adults. unibrtunately, children often receive no treatment, or inadequate treatment for pain and painful procedures. this prospective, multicentric study focuses on the efficacy, safety and side effects of novalgin (metamizol sodium) for this indication. patients and method: novalgin was administered to children, aged between - years, with acute, postoperative or procedural pain. novalgin ( - mg/kg) was given - hourly iv or im respectively, in some cases ( ) in combination with opioids (tramadol , piritramid , butorphanol ). the pain relief was assessed by six-step verbal rating scale (vrs) from to , vital signs were monitored, the side effects, that occured were recorded. results: pain relief was good (vrs less ) in children - . % of study patients. novalgin was very well tolerated, only one patient had adverse reaction -hyperpyrexia following intravenous application of the drug. discussion: novalgin (metamizol sodium) is safe and effective drug in the management of acute pain in children with low incidence of side effects. obie~qve: a prostx~tive study comparing simultaneous, indepeadent ratings conducted by intensi~ sts using an american (comfort) and an european chartwig) sedation scale for mechanically ventilated pediatric patients. measurements and results: the study comprised observations in mechanically ventilated pediatric patients (aged days to years) in a pediatric intensive care unit (from march to january . each patient was sedated by his/her managing physician with opiates, benzodiazepines, barbiturates, used isolated or in combination. each observation consisted of a -mid period of oly~ervatien of the patient in his or her pediatric icu bed, after each observation, the comfort (analyses dimensional physiologic and behavioral subscores -range to paints) and hartwig (analyses dimensional behavioral subsenres -range to points) were performed by the intensivist. we established the comfort scores ~ correspanding to adequate (range to ), excessive (range to ), and inadequate (range to ) sedation; and, hartwlg scores z correslxmding to adequate (range to t ), excessive (range to , and inadequate (range to ). statistical mmlysisj: agreement rate (kappa) and p <. was considere d s!l~nificant. comfort ( . %) ( , %/ ( . %) hap, twig , ( . %) ( . %) ( . %) to the comfort score, the average for adequately sedated, inadequately sedated, and too sedated was . +- . , z _+ . , and a.+_l , respectively. and to the ha~twig scorn, the average for adequately sedated, inadequately sedated, and too sedated was . :k-' . , . -&l , and . l- . , respectively. conclnsion: in our study there were no significantly statistical difference when you apply a more complex scale (conff'ort) or a less complex scale (hartwig) to assess the sedation of mechanically vemilated pediatric patients. the application of local and intravenous morphine infusion after surgery of urinary tract eva nemeth , m.d. semmelweis medical university , first oepartment of paediatrics , budapest , hungary in±roduction:continuous analgesia with morphine may be ~egaroed as a safe and effective method of pain relief during postoperative period. subjects and methods: children /mean age . years/ underwent elective ureteroneoimplanta±ion were randomly selected to receive either morphin intravenously of lo ug/kg/h /group one/ or bladder morphineinfusion ug/kg/h /group two/ after surgery. all patients were prospectively evaluated during their s±ay in the postanaesthetic care unit. cardiac and respirafory rates,blood pressure,sa ~,degree of alertness,pain perception and complaints of the paticnto ~cr~ recorded hourly. pruritus,nausea and vomiting,voiding difficul-±ies,sedation,dysphoria were systematically sough and quoted. statistical analysis was performed by chi square test. results:postoperative analgesia was the same in the two groups,but side effects were less in the bladder morphine group,because of the lower se morphine concentration.the differentes weren't significant in two groups. conclusions:the administration of bladder morphine infusion is a safe and effective method in children. objetive: compare the evaluations of sedation level made by physicians and nurses with the visual analog scale (vas) and the comfort scale (cs) in pediatrics patients receiving difforents modes of intravenous sedation. material ~ method." file evaluations were made by an attending physician and nurse with the vas and by another physician (always the same) using the cs. the observations were divided following the sedation mode: one drug (fentanyl or midazolan), two continuous drugs, one continuous and one intermi~ent drug and two intermittent drug (fentanyl and midazolan). the groups were compared using the t-student test. the groups also were compared between the percentual of agreement of the evaluations of sedation level made by physicians and nurses with the cs and vas using the x . results: we didnk find any statistical difference between the observations made by physicians and nurses with the vas in the differmts modes of intravenous sedation, the average of the observations using the cs betwom one drug and two drugs modes didnk exhibit also statistical difference. the observations made by physicians mad nurses using the the vas when compared with the cs didn't show statistical difference between the sedation level. we found statistical difference only in percentual of concordance of sedation level between physicians and nurses when compared the one and two drugs modes of sedation. conclusion: we didn't find differences in the observations made by physicians and nurses in the sedation level, only in concordance pereentua/ of observations when compared two modes of sedation. the observations using the cs (more complex) didnk show differences when compared with the vas. effects of age, concurrent administration of other pharmacologic agents, and disease [cardiac(n= ) & pulmonary(n= )] on the pk & pd of b were evaluated in volume overloaded infants aged days- mo (n= ). single doses of . , . , . , . , . , , , . , . & . mg/kg iv were given over - min after baseline evaluation. age was used as a continuous vadable to determine its effects on the variability in the pk & pd of b. values for pk parameters were compared between patients in cardiac and pulmonary disease groups. hierarchical multiple regression analyses were used to determine the effects of age, disease and other pharmacologic agents on the variability of bumetanide excretion rate (ber) and pd responses, e.g. urine flow rate (ufr) & electrolyte excretion. cit, cir & cinr increased with age (p< . ) while t, decreased markedly in the first monthe of life (p< . ). ber normalized for dose increased with increasing age. patients with pulmonary disease exhibited significantly greater clearance and shorter t~= (p< . ) than those with cardiac disease whereas vd~ was similar in both groups. the administered dose of b was the primary determinant of ber but increasing age also contributed. penicillin antibiotics decreased ber. dose response curves for ufr and electrolyte excretion were similar between disease groups. more of the variability in ber and pd responses could be accounted for in the pulmonary group than the cardiac group but this was not statistically significant. conclusion: the pk of bumetanide were influenced significantly by age and disease. differences in pk between patients with pulmonary and cardiac disease were primarily due to differences in total clearance. age and the administered dose of b were positive determinants of ber and pd responses while penicillin antibiotics had a negative impact on both, once b reached its site of action, no differences in pd responses were detected between disease groups. the pharmacodynamic effects of bumetanide were evaluated in volume overloaded infants (n= ) aged days- months. single doses of . , . , . , . , , , . , . , . & . mg/kg iv were given over - rain. bumetanide concentration in blood (n=l ) & urine (n= ) samples were quantified by hplc. baseline urine samples were collected over - hours prior to drug administration. determinations of urine volume, electrolytes (na ", k +, ci, ca ++ and mg++), creatinine and osmolality were performed before and at - , - , - , - , - and - hours after bumetanide dosing. changes in urine flow rate and electrolyte excretion were plotted as a function of bumetanide excretion rate which was considered the effective dose of the drug. peak bumetanide excretion rate increased linearly with increasing doses of drug and showed no evidence of approaching a maximum. time course patterns for urine flow rate and electrolyte excretion were similar for all dosage groups. urine flow rate and electrolyte excretion increased lineady up to a bumetanide excretion rate of approximately #g/kg/hr and either plateaued (urine flow rate) or declined at bumetanide excretion rates > #g/kg/hr. bumetanide had no detectable effect on serum electrolyte concentrations, conclusion: maximal diuretic responses occurred at a bumetanide excretion rate of about ;~g/kg/hr. higher bumetanide excretion rates produced no increased diuretic effect. peak bumetanide excretion rate of about #g/kg/hr corresponded to bumetanide doses of . - . mg/kg. neonates using an electrical syringe-pump. authors: tr~luyer j.m., sertin a., bastard v., settegrana, c., bourget p., hubert p. background and objective: many problems can be observed with drug administration by i.v. route, especially in neonates. so we evaluate different protocols of teico delivery using an electrical syringe-pump. methods: we simulate infusion of teico with a syrlnge-pump (pilot c, becton & dickinson lab.) trough d standart neonatal i.v. system. for weights ( or kg) we used doses of teico ( mg and mg/kg) and a dose volume _< . ml. our goal was to perform a complete infusion in minutes. the infusion system consisted of an life care infusion pump (abbott lab.) with its lv. set for maintenance intravenous fluid (flow _< ml/h) connected to a -way stopcock. an meter extension tubing was placed between the stopcock and a neonatal catheter. an another meter tubing (injection tubing) connected the teicoplanine syringe to the stopc, ock. the volume of the injection circuit (from the syringe to the distal part of the catheter was . ml methods of injections were assessed: a: injection of the predetermined volume of teico in minutes with no wash out. b: idem as a but the teico was injected in minutes, followed by a wash out ( ml / minutes). c: twice the required volume was introduced in the syringe and the volume to infuse was programed in minutes, followed by a wash out ( ml/ minutes). d: ]dem as c but a priming was performed before connecting theteico syringe to the tubing. during each run, serial samples were collected every ten minutes over a one hour period. the samples were assessed using hplc method. results: the amount of drug delivred at minutes were calculated. the results are a mean of to runs and expressed as the percentage of the total amount of teico prescribed. a , % , % b % , % c a % , % d , % % conclusiom for accurate and reliable intermittent drug infusion with a syringe pump it is mandatory to use a precise protocol of administration and to take in account ) a priming (for immediate starting of infusion), ) a drug volume greater than the dose prescribed and a programmed volume injected, ) a wash out of the tubing (with a volume ~ , x volume of tubing injection) caz is an antibiotic with activity against the major pathogens responsible for neonatal bacterial infections. we previously reported the pharmacokinetics of caz in preterm infants on day of life which showed that the clearance of caz increased with increasing gestationat age (ga). mean serum half-life of infants with gas < wks was . h. we wanted to investigate the effect of postnatal age on caz pharmacokinetics, we therefore studied caz pharmacokinetics on day - of life in preterm infants with gas < wks. caz ( mg/kg) was administered as an intravenous bolus injection. blood samples were coilected before (t = ), and . , , , , and h after the caz dose and analyzed by hplcassay, the pharmacokinetics of caz followed a one-compartment open model. during newborns with complex congenital heart defects requiering either htx or palliative staged single ventricle repair were admitted to our hospital: hlh n= , unbalanced cavsd, tga with hypopl. rv and hypoplastic aoa. tga with hypopl. rv, sas and dextrocardia. /i children had been admitted with cardiogenic shuck and mukiorgan failure due to intermittend closure ofductus arteriosus; in / stabilization failed. parents were informed about the known and unknown risks of the always palliative surgery; in cases parents denied further therapy. one pafiem with hlh underwent orthotopic htx at the age of month after the ducms art. had been stunted in the newborn period. month later he is still in favourable condition and without any sign of acute organ rejection. / underwent first stage of palliative single ventricle repair: norwood -op. ( ) ( n= ), damus-kaye-stansel -procedure ( ). the clue to adequate postoperative management was to archieve a balanced distribution of flow to systemic and pulm circulation, that is to protect the single ventricle from volume overload and to guarantee sufficient oxygenation and pulmonary development as well. with the centralvenous sato at about % provided maintaining the arterial sato at about _+ % is corresponding with a qp/qs of : . using modified bt-shunts of . mm resp. a central anrtopulm, shunt of mm in one case l severe puim. hypertension, surgery at weeks of age ) there was no excessive pulm. blood flow and no need to increase pvr with inspired co . one child ( norwood at weeks, preexisting pnim_ edema ) developed severe pulur hypertension and parenchymal pulm. dysfunction after prolonged bypass and multiple transfusions due to intraoperative bleeding: hypoxemia could be managed successfully by implanting a second shunt of mm hh later and temporarily using prostacyclin and no; at sternum closure dd later the second shtmt was banded to ram. follow-up ranges - month: all children are at home being assigned for second stage operation at about month of age. establishing clinical practice guidelines has become increasingly important in the current health care environment. significant effort has been focused upon development of post-operetive critical care pathways. however, benchmark data upon which such pathways should be based has not been well reported. length of mechanical ventilation (lmv) and length of stay (los) for children following cardiac surgery, for example, is poorly described. we prospectively recorded the lmv and los in patients who underwent cardiothoracic surgery between / / to / / . only patients who belonged in any one of five categories of congenital heart disease (ventricular septal defect _+ other septal defects (vsd), atrioventricular (av) canal, tetralogy of fallot (tof), transposition of great arteries (tga), and single ventricle physiology (fontan)) were included. eight non-survivors were excluded from the analysis. all patients were admitted to an intensive care unit cu) post-operatively where mechanical ventilation was managed by pediatric intensivists. lmv was defined as the period from post-operative admission to planned extubation. length of stay (los) was defined to be from le from the icu. cytokine patterns during and after cardiac surgery in young children. especially in children, cardiac surgery with cardiopulmonary bypass (cpb) can cause a systemic inflammatory response. this process is thought to be mainly a result of inflammation induced by surgery and exposure of blood to an artificial surface, and of reperfusion injury during weaning of bypass. complement activation, degranulation of granulocytes, induction of free oxygen radicals, endotoxemia and release of cytokines, are important contributing factors. we studied cytokine patterns before, during and after cpb in young children admitted for complex surgery or for septal defect correction. in the first group, significant amounts of il- and il-lra could be detected preoperatively. these findings could reflect the already existing hemodynamic dysregutation. in both groups, cpb procedure upregulated the circulating pro-inflammatory cytokines il- / , but not il- b. at the same time, il-lra became detectable. therefore, we suggest that in these patients the production of the anti-inflammatory cytokine il-ira was not induced by the preceding acnvity ot pro-inflammatory cytoidnes. during cpb, we noticed a sharp decline in the capacity of the leucocytes to secrete il- / . the ex-vivo production of il-lra however, was only slightly attenuated. we conclude that there is a differential regulatory pathway for the induction of il- / and il-lra. in addition, we studied the influence of dexamethasone administration on the cytokine pattern. administration delayed the appearance of il- / and il-ira in the plasma, interestingly, it did only interfere with the ex-vivo production of pro-inflammatory cytokines. the latter supports our hypothesis that production of il- / and of il-lra is regulated by two independent pathways, ( %) of pts. % ofpts < months of age developed metabolic alkalosis as compared with % ofpts > months of age.the infants with metabolic alkalosis received more citrated blood products and furosemide. following cardiac pulmonary bypass the highest ph-values and be-values were observed - hours and - hours, respectively. ii. prospective study: metabolic alkalosis was registerd in t children ( %), of those < month ( %) developed metabolic alkalosis and % of those elder than monms.durmg the postoperative course patients younger than months developed the highest ph-and base excess values after and t hours, in the subset of the older patients maximum ph and base excess was found after and hours, respectively. in one case the top level ofph-value exceeded . , the base excess + mvalb. conclusion: children undergoing cardiac surgery with cardiopulmonary bypass often develop metabolic alkalosis.in contrast to previous reports, we did not observe an association between metabolic alkalosis and mortality, nor greater frequency of cardiac arrythmias or prolonged mechanical ventilation. in context with decreasing serum lactate levels, our data show positive correlation of metabolic alkalosis with postoperative improvement of liver function. respirator, mechanics and weaning outcome in children undergoing cardipvascular surgery. vassallo j., cernadas c., saporiti a., landry l., rivello g., buamsha d., rufach d., magliola r. mechanical ventilation (mv) and acute respiratory failure are common events in children unergolg cardiovascular surgery (cvs), the development of new techniques helped to measure some of the main respiratory mechanics (rm) in a non invasive fashion. our goal was to evaluate the predictive value of these measurements in weaning (w) outcome in these patients, patients and methods: we prospectively evaluated children considered clinically to be ready for w with < kg and > hs mv. patients with diaphragm paralysis and those who failed w because of upper airway obstruction were excluded. before patient extubation the following measurements were recorded during spontaneous ventilation (cpap/t piece) using the cp neonatal pulmonary monitor bicore (lrvine, ca): total respiratory system static compliance (cssr) and resistance (rts), rapid shallow breathing index (rsbi). maximal inspiratory negative pressure (pi max) was measured using an unidirectional expiratory valve. threshold values predicting w success (ws) were: cssr > . ml/cm h , rts < cm h /l/sec, rsbi and pi max > - cm t . w failures (wf) -patient reintubation within the following hs, these values were compared between w success and failures using fisher exact test. an apriori level of statistical significance was chosen at p < . . considered, an increase in tnf-a levels is observed after cardiac surgery (p< . ) with a return to previous values after hours (p< . ). hours after cpb, similar values are observed in groups ii and ill, but there is a further increase in serum tnf-a levels in group i when compared with both other groups (p< . ). we found no statistically differences in any other moment. there was a significant correlation between serum tnf-o levels determined hours after surgery and cpb duration (p< , ). conclusions: cpb in childhood provokes a significant increase in serum tnfa levels, in newborns the inflammatory response is maintained hours after surgery. this enhancement of serum tnf-e levels indicates the existence of a relevant inflammatory response in these patients. introduction: cardiac surgery appears to induce a systemic inflammatory response. we have investigated the behaviour of il- i~ and il- before and after cardiac surgery. patients and methods: we studied serum il- and il- levels from children with congenital heart disease ( boys and girls), aged from days to years, undergoing open heart surgery, before cpb (d we found no statistically differences in the il-i levels in the different groups and moments. there is a significant increase in il- immediately after surgery (p< , ) with similar levels hours after cpb and a significant decrease (p< . ) hours after cpb. preoperatory il- levels were higher in the groups i and tl than in group i (p< . ). hours after cpb serum il- levels in group were significantly higher when compared with group (p< . ). conclusions: cpb in childhood induces a significant transient increase in serum il- levels, strongly relevant in newborns. cpb was not associated to a significant modification in serum il- levels. thus, cpb in childhood induces a dissociated behaviour in the proinflammatory il- and il- & pathways. obiective, to evaluate the effects of amg receipt on the clinical condition during the first hours after birth (t ), the morbidity and mortality in immature outborn neonates. methods. we studied outborn neonates with ga to wks, admitted during the years to . eighteen neonates exposed to amg (ga: , +lwks, bw: _+ g) and neonates did not (ga: , _+ wks, bw: _+ g). results. amg-exposed neonates compared to those not exposed had lower incidence of apgar score at min _< ( % vs %, p<. ), lower incidence of ph t < . ( % vs %, p<. ), decrease need of bicarbonate ( % vs %, p<. ), lower fio (fio min> : % vs %, p<. and fio max > : % vs %, p<. ), lower incidence of intubation ( % vs %, p<. ), lower requirements of surfactant ( % vs %, p<. ) and lower mortality ( % vs % p<. ). there were no differences between the two groups for the following parameters: type of delivery, hypothermia hypoglycemia and anemia during admission, hypernatremia, hypotension (map< mmhg), need of dopamine and or plasma , incidences of ptx pda sepsis nec severe rop major ivh (plus pvl) and bpd and duration of intubation. conclusions. the main beneficial effects of amg receipt on the immature outborn neonates were the decrease of mortality and the decrease of surfactant need. there was no effect of amg receipt upon other severe morbidity in this high risk group of neonates. premature babies are very sensitive on homeostatic disturbances, and often develope intracranial haemorrhage (ich). ultrasound scan of the bram shows four grades of ich: -grade i -only periventricular hyperechogenic areas -grade ii -haemorrhage ham the lateral ventricles -grade ili-dilated lateral ventricles -gtrade iv -intracerebral haemorrhage. the purposes of this study were: to show the incidence of ich in premature babies and its correlation with the gestational age, . to determine the severity of ich . to present the outcome &those babies. in the study were included premature babies successively-born at the department of gynecology and obstetrics before gestational week (g.w.) and grouped in three groups: less than g.w., - g.w., - g.w. to all of them was performed ultrasound scan of the brain. results : . the incidence of ich hi premature babies is % and there is ingh level of correlation with the gestational age: -babies born before t~ g.w. have % incidence of ich and graduated : i grade - %, ii grade - %, iii grade - %, iv grade - % -babies old between - g.w. have incidence of % : i grade - %, i[ grade - %, iii grade - %. -babies older than g.w. have incidence of %: i grade - %, ii grade %, iii grade - % . sixty of premature babies have died and it is . % lethality. in all died ilffant was confirmed the grade of ich diagnosed by ultrasotmd scan of the brain. d. maksimo~ c. z.braiko~ic, n.vunjak. p. ivanovski ( ~iversi~, children's hospital. belgrade, yugosla~, ia infantile intracranial hemorrhage is the most frequent and serious manifestation of late hemorrhagic disease of the newborn caused by ,,~tamm k deficiency in earl?,, ti~fancy. in the last two years, we recorded five cases of infantile intracranial hemorrhage due to "dtamin k deficiency, despite routine prophylax~s (intramuscular vitamin k, mg) , with bpieal clinical presentation: age was - days (average days): vomiting, poor feeding, lethar~'irritabiljty, palor, bulging t ntanelle and convatsiones were present in most cases.two patients developed signs of hemorrhagic shock, with hemoglobin level less than g. . in ~ f \qi level was less than % of predicted value. there was no evidence of head trauma or liver disease in none of patients. four inlants were breast fed, while one, who had diarrheal disea.se, was on adapted milk formula. routine therapy wa.s given (including vitamin k and fresh frozen plasma). two patients were discharged with no sequellae, one developed posthemon'hagic hydrocephalus as a complication and two patients died. late hemorrhagic diseo.se of the newborn is sill/ a significant cause of morbidib' and mortality in earl ' infancy, despite different approaches to prophylaxis developed in recent years. background: neonatal hearing screening in at risk newborns can detect % of the children with a congenital hearing loss. automated abr hearing screening (algo- ) has been introduced for healthy newborns. the aim of this study is to test the validity of this algo- screener in at risk newborns in a neonatal intensive care unit. subjects: at risk newborns (median gest.age: . wks, median birthweight g) selected according to the criteria of the american joint committee on infant hearing. interventions: algo-i automated abr-hearing screening at a level of db was performed in the neonatal intensive care unit. when bilaterally referred, further audiologic screening and/or therapeutic intervention took place. when passed uni-or bilaterally, children enrolled in a) a nation wide screening programme (ewlng) at the age of months and b) in a half yearly follow-up programme in which hearing and speech-and language development were observed according to egan an illingworth. results: screening without disturbance from ambient noise or from routine technical equipment was possible in the incubator, even during nasal cpap therapy. ( %) newborns passed algo- screening. ( %) did not pass bilaterally. of with a congenital rubella died shortly after screening.in of bilateral congenital hearing loss of -> db was confirmed. of the newborns passed were still alive at the age of year. ewing screening was performed in of ( , %). / passed, of had passagere conductive hearing loss, in / no further investigation was performed. all children enrolled in the i/ yearly follow-up programme had normal speech-and language development. in this study all at risk newborns with bilateral congeni "tai hearing loss were detected with algo- screening. screening results showed no false negatives at follow-up. the algo- infant hearing screener can be used as an valid automated abr-screener to detect hearing loss in at risk newborns in a neonatal intensive care unit. gancia gp, bruschi l pnlito e, ferrari g, rondini g -divisione di patologia nc~matate e turapia intensiva -irccs policlinico s. mattco -pavia, italy latrogenic esophageal perforations (iep) in preterm and term infants are seldom reported in litteraturc, in association with difficult endotracheal (et) intubation (with or without stylets), insertion of gastric tube, and pharyngeal suctioning with stiff catheters. crieopharyngeal muscle spasm caused by instrumentation may also lead m a narrowing of lumen, with increased risk of local injury. we report iep observed in intubatcd, mechanically ventilated newborn infants ( male, female, all outborn). a common feature of iep was inability to pass a nasogastric (ng) tube into the stomach, mimicking e~)phageal atresia.~se : birth weight (bw) (i g, gestational age (ga) wk, sepsis. before admission to n cu, the baby underwent multiple et inmbations, because of inappropriate securing of et robe. bloody secretions in pharynx were observed. the endoscopy showed a large lesion at the end of proximal third of the esophagus, case : bw g, ga wk, rds. chest x-ray (cxr) showed a retrostcrnal air leak: the ng tube was stopped }~etwcen d and d and soluble contrast was seen in upper mediastinum.case : bw (/g, ga wk, rds. the endo~opy showed an esophageal lesion. cxr showed a paravertebral route of ng tube and a right pneumothorax.case : bw (i g, cz ,.v!:. rd c. ~!,'.::;;: ::':'_'rvt!~' s l" ~k':.rvrx. cwr, d,,,,vs ~,,mr~e, ~n rhe upper mediastinum and abnormal route of ng tube through a false passage. surgical intervention is needed in case of mediastinitis or mediastinal abscess: conservative management included broad spectrum antibiotics, total parenteral nutrition, antireflux therapy and, if necessary, drainage of air leaks. enteral feeding has been stopped lor days and cautiously resumed after radiographic study. [x~cal sequelae and death are uncommon, but iep occur in newborns with high risk of death due to prematurity and other diseases. in our patients, et intubation has been performed by experienced personnel: therefore the lack of skills in resu~itative procedures is not always the main factor of iep. prevention of iep requires appropriate materials (et tubes, laryngoscope blades, suction catheters), and procedures (positioning of the infant with correct neck estension, firm et placement). sedation and pain control may help to prevent the muscle spasm. aggressive treatment has improved the tong-term outcome of extremely low birth weight neonates (elbw) but it has also increased the chances of iatrogenic lesions. reviewing the charts of our neonates we observed a high number of vascular injuries. from to , neonates were admitted to the neonatal intensive care unit (nicu); of them were elbw ( . %). studying the charts of these elbw we observed cases ( m - f) with vascular lesions ( . %). mean gestational age of these patients was . weeks (rain -max ). mean weight at birth was g . mean weight at diagnosis was g ( - ). in the same period patients with vascular injuries were reported in the neonates over g ( . %). the injuries observed in elbw group were: arteriovenous fistula ( bilateral) at femoral,level, carotid lesion and limb ischemic lesions. aetiology was in cases by venipuncture, in one case umbilical catheter and in the case of carotid lesion a wrong surgical maneuver. no general simptoms were observed. the vessels were repaired with microsurgical technique in six cases: the carotid lesion and five arteriovenous fistula; one case was solved with thrombolitic drugs; an amputation at knee level was required in one case after a long period of medical treatment. the last neonate with an arteriovenous fistula was only observed for parent's will. at follow-up (clinical and by ecodoppler) out of neonates presented normal vascular function without sequelae. from our experience elbw neonates have more chances than older neonates to develop iatrogenic vascular lesions. we advocate an aggressive microsurgery and/or medical treatment to obtain good results and prevent late sequelae. a retrospective comparison between natural surfactants l.j.i.zimmermang m.c.m,van oosten. dept. pediatrics, div. neonatofogy, sophia children's hospital/erasmus university, rotterdam, the netherlands. aim: retrospective comparison of alvofact (in ) versus survanta (in ) as rescue treatment for neonatal respiratory distress syndrome (rds). methods: both surfactants were given at an initial dose of mg/kg (except for alvofact mg/kg for mild rds grade mi). repeat doses were attowed (survanta mg/kg, alvofact mg/kg) up to a maximum of mg/kg, all parameters and outcome criteria were strictly defined beforehand. the initial response (good,mild,no response,relapse) to surfactam therapy was defined on the basis of the decrease in fio . results: there were no signif. differences in patient population and initial parameters: ga ( . +_ . vs a _+ , wks), birth weight (t _+ vs -+ g), severity of rds (grade ill-iv: . % vs . %), apgar scores, cord blood gases, initial ventilatory settings. in ' however, the initial surfactant dose was administered earlier than in ' ( . -+ . vs . _+ . hrs postpartum, p= . ). although the average total cumulative dose was equal in ' and ' ( . -+ , vs . _+ . mg/kg), more doses of alvofact were given compared to survanta { . _+ . vs . _+ . , p=o.o ) and more patients in ' received more than two doses than in ' ( % vs % of patients). there was no difference in the incidence of non-putmonarycomplications. aivofact ( there was a better initial response to survanta and a better respiratory outcome in : in the group < g the duration of ventilation was half in , and in the group >~ og the duration of extra o need was half in as compared to . we speculate that the main reason for this difference is the earlier and initially higher dosing used with survanta compared to that used with alvofact which was given in the same total cumulative dose but over a larger time span. background: e×ogerlous sur&ct~t raplacem~t treatmem has become rou~ne k~ the t~eatme~t of respira~"¢ dim'~ syndrome (i~ds) of pr~e~tur~, wh~eas its effica w th odi~ respiratory diseoses is sdi being wader mvesugatio~. objective: "eac~ mt ereat isto report ottr results of prospect/re, non-randomized "re~-o.e" study oe suffact~t replacement in outhom premamae infa~t~ with rds reruirmg me~aical ventilatioa (nfv). p~tien~ and metho .s: from j-aly to june , / ; ( %) out~ ~¢ infaats, at a mesa age of z , horn's ( boys, ~rls; ~ gestafioan age -+ . weeks, mera~ birth weight _+ g, ~ . i" at minutes) with rds, requiring mv, received bov~e-suff~amt (survanta, ros~/aboti, laboratotie~ columbus, ohio) eadotracheally, as was recomm~aded by maaufacturer. as the c,~:ttrol group o~bom premature infants (ot~ of ; %, admitted with rds from euiy to eune ) were saelected ~d who did not receive surfaaam, compared with ~hctant ~'oup they were admitted for treatmeat e~'li~" aft~" daliv~:y (at the age . :: . hours vs. . +- hours), but they did not diff~ in othe~ baseline dam'a~eri~cs at ~ti~ion. entry crkeda for ~¢fa~aut ~hcadou were fractional i~firat o~ oxtgem r~emeats -fio > . - . , ratio au-lerlal to alveolar oxygea pre~are~ao ~ao < , ~ad oxyge~at,~ i~.dex -ol > . primary o~comes were deter~caned by ~hanges m exs'ge~ab, c~ ~r~d vmtilatic~ ~ the following variable~; ( ) fi'aaic~ of i~spired oxtge~ (fio ); ( ) mesa nnvay presmzre (map) ( ) pag ~ao ratio, ( ) oxyge~ion index (oi). commo~ comphcadces of prem,musty ~d con~ol mechamcal v~ati]al~on (pater dumas merios.s, intracr~nlal haemcrn:hage, air leak, br onchop ulmrmm'y dy~pl~a ~d death) were reg~ded as sec~d,~y outcomes. r~suas: in warfactaat group we observed slg~ .c~t improve~aeat (p< . ) in oxygea~thia md veaatilation at hours all~ e~try k~to the m~dy in compari~ion to nons~fa~m" group. compa~on of secondao' outcomes in ~ts with p,.ds showes table l we did not observe ~y major acute hfe fl:u-eattming complicatlola,s m sxlrlhct~mt grou~ tr/lmediately after stu'~actsmt rcplacemev_t therapy. the duramm of mechmucal ven~ation ~ad oxygen lreau~ent m survivals of both groups did not dafter gmficautl y a-ore ead~ other. condusion: l!a premature mthats with rds treated with surfaaaat replacemeaat therapy we observed decrease m mc~de~ce of tme'~m~o~oraces add de~th (p< . and p< . ), whe~e~s m othe~ observed variables thee was uo ,igmfi~t d~=ecce infectious complications during the therapy of respiratory insufficiency in neonates with birth weight less than g in the course of yearsretrospective study. zitek infants on cmv, cppv, and imv were administered exosurf in dose of - mg/kg twice endotracheally (see table) . in newborns ( . %) hours after surfactant admin fi value decreased by . %, and after hours -by . % compared with initial value; pip and peep values decreased by - cm h and - cm h after hours, and by - cm h and - cm h after day, respectively accompanied by mean decrease of aado from , to . mmhg, qs/qt decrease from . to . % (see table) . mean time of cmv, cppv was . days, imv- - hours, cpap - - hours. respiratory therapy in newborns ( . %) was complicated by pneumothorax (bilateral -in infants chorioangioma is a rela~ively rare placentai malformation associated with considerable mortality and morbidity. a chorioangioma can be regarded as an arterio-venous shunt in the circulatory system of the fetus. this causes volume loading eventually resulting in cardiomegaly and high output cardiac failure. a female neonate (gest age wk, birth weight g, - . sd) was born with an apgar score of and after and rain respectively. the placenta showed multiple chorioangioma. ultrasound of the heart showed a hypertrophic cardiomyopathy. she developed severe hypertension ( / mm hg), treated with nitroglycerine and nitropruside. finally blood pressure decreased when enalaprillic acid was given ( . mg.kg ). we measuered the activity of the renin-angiotensinsystem. an elevation in renin-angiotensin system is shown probably to compensate for the low resistance circulation before birth, hypothesis: the instantaneous cut off of a large arteriovenous shunt did not result in a fast downregulation of the renin-angiotensin system resulting in hypertension. hypertension should be added to the list of complications of chorioangioma of the placenta. the authors studied cases of children's septicemia with blood culture yielding staphylocucetts aurens. the age of patients varied from months to years ( , % from years downward), % of the children caught their disease in the hot season (may to october). the deaths also occured in this season: , % ( / ). following were the anatomo-dinical lesions. -skin %, muscle , %, bone , %, joint . %. -viscera : lung %, heart . %, cerebrum . %, kidney . %, fiver , %. -simple lesion skin-muscle-bone joint: %, no death in this group. the concomitant lesions of the soft tissue,bone-joint and viscera : % with one viscera, % with two viscera, % with three viscera and % with four viscera. -bone lesion : mainly on the long bones ( % on the tibia, % on the femur, the remainder being the mandible ( ) and the humerus), inflammation of' the hip joint was the main one. -i,ung lesion had forms pneumatocele ( cases), bronchopneumonia ( cases), pleural effusion ( cases), multimicroabcess bursting into the pleura ( cases), most multimicroabcesses were lethal : / ( , %), -heart: all thethreelay~rs got le@~r~, % had or layers alrected and death ensued. -cerebrum : the meninges had three forms of lesions purulent meningitis ( cases), obturafing embolns of brain vessels ( cases) and cerebral abcess (one case). the characteristic clinical sign was paralysis and meningismus, phlebothrombosis of eavcrnous finus ( cases)was mually ther~sultofalxil vdfi:h burst there were cases of death with lesion of the meninges and cases of obturating embolns of brain vessels. -the main sign of lesion of the kidney was a change in the components of urine: % got proteinuria, % had leucocytes in their urine, % had erythrocytes in their urine, the urea in their blood increased (over rag%) in . % of cases.the lesion of the kidney seemingly had little relation to death. seven cases of ictertts due to an increase of direct bilirubinemia and a decrease of blood-albumin. -the biological characteristics of the pathogen staphylococci showed that all the isolated specimens had positive coagulaza ; the specimens from the dead patients were less semiti~e to, mad ~t to mali~ overag death rate was . % ( / ). the fungal infection to fusariun species in immunocompromissed child have been reported in the literature with a rare, severe and high, mortality rate in spite.of the use of antifungal drugs. we report a case of successful treatment of a severe disseminated fusariun infection in a ll-year-old boy with acute lymphocytic leukemia (lla-l ), after use a chemotherapy followed by absolute granuloeytopenia. the patient developed fever, skin lesions, pneumonia and fungaemia. fusariun species was cultured from the blood, necrotic skin lesions and lung secretion. the child developed multiple organ system disfunctiou in spite of use broad spectrum antibiotcs and antimycotic therapy needing. uci during days. the patient receive suport treatment (mechanical ventilation, inotropie d~.ugs, diuretics, imunestimulants, blood components, a broad spectrum antibiotes and antifungal agents). we absorved a gradual recovery in the white blood cell count and regression on the sites of infection. the association of preeoce diagnostic and the terapentic with increase in the white blood cell count was the most important in a successful treatment. a year old african-american child suffered a severe pulmonary injury in a house fire. initial survey revealed % total body surface burns, soot on the face, and bloody endotracheal secretions. initial chest radiograph revealed diffuse, bilateral infiltrates. severe respiratory failure with an oxygenation ratio of rapidly developed. he developed a pneumomediastinum and subcutaneous emphysema. although transient improvement occurred with inverse i:e ventilation and surfactant, he became more hypoxic (sac as low as %) and acidotic. on day post injury, he was placed on venc~venous extracorporeal life support (ecls). on ecls day he was decannulated. chest radiograph on ecls day showed an opacity in the left chest. ultrasound of the left chest was consistent with atelectasis rather than pleural fluid. flexible bronchoseopy failed to reveal any obstruction in the left lung. a computed tomography (ct) seen of the chest, which was performed after decannulation, revealed a large loculated collection of fluid in the left, anterior chest. under ct guidance, a f cope loop catheter was inserted and cc of thick blood was removed, follow-up ct performed immediately after this procedure revealed minimal change in the size of the fluid cavity. over the next hr, we instilled urokinase , units over minutes every two hours. a minute dwell time was allowed before draining the fluid. repeat ct scan done at the end of the urokinase infusion showed a marked decrease in the size of the fluid cavity. act scan was not performed prior to decarmulation because the ecls circuit tubing was too short to allow appropriate positioning of the child in the ct scanner. after a ct scan revealed loculated pleural fluid, a simple drainage procedure was diagnostic but inadequate treatment. we were able to successfully dissolve the thrombus after hr of urokinase therapy even though the thrombus was > days old. we suggest that large loculated plenral thrombi which develop as a complication of ecls therapy may be successfully managed with urokinase infusion. introduction: haemorrhages, particularly intracranial, are major complications experienced in - % of neonates treated with extracorporeal circulation. an induced thrombocytopenia and impaired platelet function play a key role in the increased bleeding tendency observed in these patients. the aim of the present study was to establish a dose-respons curve for the effect of a synthetic protease inhibiting agent, nafamostat mesilate (fut- ), on platelet membrane glycoprotein density and platelet activation during experimental perfusion. methods: two identical extracorporeal life support (ecls) circuits were primed with fresh, heparinized human blood and circulated for h. four different concentrations of fut- ( . mg/l blood/h; . mg/l/h; . mg/l/h+ % bolus at the start of the perfusion and & mg/l/h+ % bolus) were used in different perfusion experiments. a total of eight paired experiments were performed. platelet count, plasma betathromboglobulin levels and platelet membrane density of glycoprotein ib and lib/ilia were followed as well as plasma concentration of haemoglobin. results: a protective effect of the agent on platelet count, plasma concentration of btg and platelet membrane gpib could be observed during the first hours of the perfusion when a bolus dose was added. no positive effect could be recorded with the two lower doses used. plasma concentration of haemoglobin was higher in all the fut-circuits compared to the control circuits. conclusion: the addition of a bolus dose of fut- at the start of the perfusion seem to induce a protective effect on platelets during the first hours of perfusion. extracorporeal membrane oxygenation (emco) is a form of invasive cardiopulmonary support that can provide imporary physiologic stabilisation in reversible circulatory failure and or respiratory failure. we reviewed our expierence with extra corporeal membrane oxygenetion in children aged day to year between and . two neonates was succesfully decanulated, but died - well after decanulation due to septic complictions. one child years old, one neonates died on day and day" respectively while still on emco. complication which were and encountered were heavy bleeding in case (child), (neonate) and raceway rupture in case (neonate). problems which are specific developing countries like indonisia are: high cost ( . us for days) difficulty in transportation (transporting intubated baby) from the orgin hospital, lack of knowledge and understanding of the primary physician and nm-ses and difficulty organizing in hours emco team. resnratory mon tor/ng in picu z,zjvkovic, s. mihailovic, o, tosev respiratory monitoring in pediatric intensive care unit picu) provide the importartt informations for understanding of the pathophysiology of the clinical signs, aid with the diagnosis, and assist in therapeutic management and predicting prognosis. pien in children's hospital for ~flmonary diseases and tuberentosis remained for the t~s't two end a half years relatively limited for diagnomic tools and therapeutic regimens, mostly because of the poor fmnaeial suptx~rt. the number of children admitted for aurae asthmatic at.lzek~ severe pneumonias, bronehiolitis, complicated pulmonary tuberculosis, foreign bodies and exacerbations of ehronit'. pulatonary diseases was t . for all patients the respirator' monitoring system means: physie~d examination, ehe~ x rays, capillary bltxxl gas mmlyses (vevv few ehiktren experienced itwasive arterial blt~.~'i gases), noninvasive oxyntctry, measuring of the vital capacity in coopo-able patients, as~d capnography. later on, after the imtial critical illness, a complete hmg fimction tests was performed, as well ,~s bronehoscopy in selected eases, (~lr experience revealed that abotrt % of ehil&en heos suecessthl outcome, without s~lllens , instead they had been tremted in limited conditions. ']'he rest of our patients were previously diagnosed ~s ettronie pulmonary patients, with high risk score system ibr having seqnells 'llae mortality rate were , %. the continuous blood gas monitor, pasatrend (biomedical sensors, ltd., high wycombe, bucks, england) has the capability of measuring ph, pco , and po via an indwelling optical absorption optodelclark electrode sensor that is placed through an intra-arterial catheter. we evaluated the accuracy of the sensor in radial and femoral locations in critically ill pediatric patients. methods: the simultaneous values of ph, pcoz, and po recorded from the paratrend monitor were compared to values measured by standard arterial blood gas analyzer (coming , ciba-corning diagnostics, medfield, ma). criteria for the elimination of data points included a core vs. sensor temp. gradient, and sensor pulled back beyond accepted insertion distance. mean time of monitoring per sensor was hours (range . - . hrs). mean time of radial monitoring was hrs (range . - . hrs) and of femoral monitoring was . hrs (range . - . hrs.). linear regression and bland-altman analysis for bias and precision for each parameter were calculated. results: a total of patients (age range weeks to years) had paired samples of ph, pens, and poz made by the sensor and blood g&s analyzer. the range of measurements were ph . - . , pco, . -i . t(n r, and po - torr. the paratrend monitor demonstrated accuracy that is comparable to the accepted standard of blood gas analysis in a group of critically ill pediatric patients manifesting wide variation in ph, pen , and poz..this technique appears m be very useful especially in the extreme values of the parameters measured. funding provided by biomedical sensors. understanding of pulse oximetry d.semple, l.e.wilson. royal hospital for sick children, edinburgh, eh lf, scotland, uk. pulse oximetry is a useful, non-invasive monitor, routinely used on the itu and increasingly often on the general wards. we used a questionnaire incorporating questions on the theory and clinical uses of the pulse oximeter to assess understanding of pulse oximetry in medical and paramedical staff doctors indicated grade, speciality, pulse oximetry tuition and neonatology experience. doctors, itu nurses, t medical students and physiotherapists completed the questionnaire. some confusion existed between the principles of pulse oximetry and transcutaneous oxygen measurement. wide variations in the lowest acceptable saturation in fit children were seen ( - %), with around % of respondents in all groups accepting values of % or less. some potentially serious mistakes were made in the evaluation of oxygen saturations in the clinical scenarios. there were widespread variations in correct responses at all grades of medical staffing. nurses scored well on more clinically-orientated questions but relatively poorly on theory. only % of doctors (mostly senior grades) had received tuition in putse oximetry. neonatology rotations appeared to confer little additional knowledge on pulse oximetry. few doctors and nurses receive tuition in the use of pulse oximetry a significant proportion of nurses and doctors, of all grades, exhibited a lack o{" understanding of the principles of pulse oximetry. this may result in unsafe use of the equipment and put patients at risk. one can see from the table that blood composition in uv and ua differens in some characteristics, and similar in sgp magnitude. venous-asterlal gradients "gas functiomals" between uv and ua represent the measure of difference in this characteristics. the gradient cari be positive, zero -order or negative and change both in value and in sign but not reach apo (positive) and apco (negative) in absolute significance.minimization of "gas functionals" deviations atom the zero is achieved due to"mutual replacement acts" between po and pco in uv and ua blood. we suggest that presented tests can be useful in full evaluation of gas exchange in newborns. (pap) in the context of pulmonary hypertension is oft desired but rarely achieved. inhaled nitric oxide (no) has been shown to produce this desirable effect, but is relatively difficult to administer or monitor. we wondered whether np, chemicaily related to no but more stable in solution, would produce similar physiologic effects when administered in the convenient modality of nebulization. methods: piglets were anesthetized, mechanically ventilated, and surgically instrumented. systemic blood pressure (bp), pap, and cardiac output (co) were monitored continuously. after postoperative stabilization, . % nac} nebulization was begun, and pulmonary hypertensiorr was induced by reducing fio from . to . . the piglets were monitored for minutes during this hypoxic phase, next, without altering fio or ventilator settings, np ( mg/ml, dissolved in . % nacl, flow ipm) was substitued for . % nacl in the nebulizer circuit. np was nebulized for mins. results: during hypoxia, pao fell from to mm hg. pap rose during hypoxia from to torr (p< . ). ,^fhile bp and co did not change significantly. pap fell during nebulized np in each piglet, (mean apap = to torr; p< . ; mean reduction of hypoxia-induced rise in pap = %; range: to %; p < . ). pvr/svr fell by % during np nebulization (p< . ), while bp and co did not fall significantly ( to tort; to mllkg-min), the reduction in pap began within minutes of the onset of nebulized np, and appeared to reach a plateau by minutes. no tachyphylaxis to nebulized np was noted. nebulized np did not significantly affect pap, bp, or co under normoxic conditions. conclusions: ) like no, np selectively reduced hypoxia-induced pulmonary hypertension without altering systemic bp, ) unlike no, np can be administered by nebulizer, a technique familiar to virtually all health-care providers, and potentially adaptable to both intubated and non-intubated patients. } nebulized np may be beneficial in clinical contexts where inhaled no is impractical. dang phuong kiet and nguyen xuan thu examining cases of purulent pericarditis with various clinical forms treated by surgery, the authors drew the following experiences for their diagnosis. t. clinical factors. purulent pericarditis appeared like a cardiac tamponade in a septicemia due to staphylococci with dassieal symptoms: severe dyspnea, tachycardia, faint heartsound, big liver, prominent cervical vein ; rentgenography of the chest showing enlargement of the cardiac silhouette, a diminution of ventricular pulsations, ~i clear lung field. by an emergency operation, ml of diluted blood were drained. purulent pericarditis and pleural effusion appeared at the same time but at first tile symptoms of purulent pericarditis were masked by the predominant symptoms of plearal efihsion. after the pleura was drained, its pus was no more, the general state was relatively stabilized but there still were big liver, dyspnea, enlargement of the cardiac silhouette while central venous pressure increased. purulent pericarditis appeared late. in the first stage (about weeks) there was no suspected sign. later on gradually appeared such symptoms as dyspnea (during serum transfusion for instance). central veinous pressure also raised. the heart chest diametre increased at first (up to - %) then decreased (down to below % ) but the liver kept on swelling together with the particular changes of electroeaediegramme. now the pericardium had no more pus but get fibrous (up to ram) thus constricting the heart and its main arteries ike pick syndrome). . diagnostic values of electrocardiograms : common signs of ecg related of these purulent pericarditis were: a diminution of voltage, a widespread elevation of the st segment, the tf wave flattened and inverted. however, what should be stressed was : the diagnostic values of an electrocardiogram for purulent pericarditis was mainly in the dynamics of their signs: in the first week, the voltage diminished corresponding to a pericardium containing pus, while the st segment went up then seemed parallel to the fibrosis of the epicardium, the liver swelled, the central velnous pressure increased, the heart/chest dimension ratio decreased, the st segment went down, the t wave became more flat and inverted. between and neonates, aged - days (median ), weight , - kg (median , ) with critical valvar pulmonary stenosis were scheduled for balloon dilation (psvp), children ( %) were on pge and ( %) needed mechanical ventilation. after stepwise dilation a final balloon : pulmonary valve (pav) ratio of % ( - ) was achieved, there was a significant correlation (p< , ) between an adequately sized balloon and freedom of reintervention. two valves could not be passed, four neonates underwent surgical procedures (brock n = , commissurotomy n = ), two children ( %) died of sepsis. / patients ( %) were successfully palliated by psvp in the first month of life. the rv : systemic pressure value fell from % ( - ) to % ( - ), complications included transient dysrhythmias, transient hypoxia, vessel occlusions;- right ventricular outflow tract perforation. in / patients follow up data is available. the residual systolic peak doppler gradient over the pav on the last out patient visit ( - months after psvp) was - mmhg (median ). four children needed repea.ted psvp to months after the initial intervention. conclusion: psvp of critically ill newborns is possible. the risk of mortality is relatively low. psvp in neonates with an adequately sized balloon is a challenging alternative to surgical treatment. post hypoxic-ischemic (hi) reperfusion induces the formation of non protein bound iron (npbi), leading to production of the reactive hydroxyl radical. it was investigated if the ironchelator deferoxamine (dfo) could reduce free radical production and improve neonatal myocardial performance after hi. severe hi was produced in newborn lambs and changes from pre-hl values were measured at , and min post-hi for (mean) aortic pressure (mean pao), cardiac output (co) and stroke work (sw). left ventricular (lv) contractility and co were assessed by measuring lv pressure (tip-manometer) and volume (conductance catheter), using inferior caval vein occlusion to obtain slope (ees) and intercept of the end systolic pv relationship (v ). npbi, reduced and oxidized vitamine c ratio (vcred/ox) and lipid peroxidation (mda) were measured from sinus coronarius blood. lambs received dfo ( mg/kg i.v.) immediately post-hi, control lambs (cont) received a placebo. results: mean pao was stable, co and sw decreased up to and % respectively in cont as compared to pre-hi. in both dfo-groups co and sw remained within the normal range. ees and v decreased in all groups post hi, but did not differ between groups. npbi and mda were higher at min post hi (p<o. ), vcred/ox was lower at min post-hi (p<o.o ) in cont as compared to dfo-group, indicating more oxidative stress in cont. conclusions: dfo reduced the oxidative stress of the heart and prevented co and sw to drop, suggesting a positive effect of iron chelation on the myocardium after h{. from we published reports with analysis of performance of the heart as a whole organ.while analysing the heart performance as a whole, we recognize three block in it located intrapericardially: l."atrial"block -left (la) and rigth (ra) atriums; ."aorta-pulmonary" block-aorta 'bulb (a) and pulmonary artery(pa); .ventricular "three-chambered" block (vb) consisting of: -left (lv) and right (rv) myocardial chambers, both frith blood outflow into "aorta-pulmonary" block vessels, and -spongy (venous) myocardial chamber with the blood outflow through coronary sinus (cs) and thebesiau vein (tv) into "atrial" block. the following conceps are introduced for vb functions assessment "common" systole and "common" diastole of "three-chambered" vb. the process of normal "common" vb systole: -begins with blood ejection from vb spongy chamber into the "atrial" block; -continues with blood outflow from rv and lv into "aorta-pulmonary" block wi~ venous minimums -x-coiiapses -~btmation iu "a~riai" block; -completes with "three-chambered" vb general emptying. at this period the following blood volumes are transferring: -two-from rv and lv(their stroke volumes)into"aor~o-pulmonary"block; -two-from "spongy" chamber .into "atrial" block; -two-from systemic and pulmonary veins into "atrial" block during the process of so called "systolic" membrane suction (at pulling atrio-ventricular valves into rv and lv chambers as blood ejects out of them). it appears to be the regulation of blood inflow to "atrial" block by blond outflow from "three-chambered" vb into "aorto-pulmonary" block. objective: to evaluate the efficacy and complications of transpyloric enteral nutrition(ten) in critically ill children patients and methods~ from june to january children (i boys and girls), aged from days to years (mean l. ± . years) were treated with ten. patients were included after cardiac surgery, with acute respiratory failure, and after cerebral surgery. the indication of ten was mechanical ventilation in patients and failure of nasogastric nutrition in patient. children ( %) had previously received parenteral nutrition. and i fg enteral tubes was inserted into duodenus through nasogastric intubation en patients and by endoscopy in patients. rx and ph determination were used as methods to control tube situation. children received adapted formula, caseine hidrolisate, and enteral formulas. (in patients the nutrition formula was changed during ten). patients were supported with mechanical ventilation during ten, received midazolamperfussion(range to mcg/kg/min), fentanyl (i - . mcg/kg/h), and vecuronium ( . - . mg/kg/h). resulte: ten was used during to days (mean . ± . days). mean maximmnvolume ad~iniateredwas ± ml/kg/day (range ) .patients with midazolam, fentanyl and vecuronitml tolerated ten similar than children without sedatives. complications were diarrhoea patient, abdominal distension and/or important gastric fed residuals patients. pulmonary aspiration or respiratory complications were no registered. ten was ended in one patient due to diarrhoea, in due to change to oral or nasogastric nutrition, children continue yet with ten and was discharged of picuwith ten. conolusion: ten is an useful method of nutritional support in critically ill children with mechanical ventilation and/or nasogastric intolerance. introduction: the enteral nutritional support of the critically ill child may reduce the morbidity and mortality by attenuating, the hypermetabolic and catabolic response, decreasing the gut translocation and the septic complications, improving the bowel mucosa integrity and the wound healing. the aim of this study was to show the safety and tolerance of en. methods: we enrolled consecutive patients admitted to our unit from may to december , mean age . years (range imonth to i years). the prism, mechanical ventilation, inotropic use, opiates and other drugs were recorded. the en was delivered by continuous infusion across a polyuretane tube.we kept the patients head elevated °, and we used cisapride routinely. the gap betweeen admission and the en beginning, average time to meet the nutritional objective, en duration, selected formula, patients outcome and complications were recorded. results: prism groups were - = patients; - = ; - = ; - = ; - = . two patients with mof (multiorgan failure) died. % needed mechanical ventilation.the average time elapsed from picu admission to initiation of en was . days (range to ) and the average duration of the en was . days (range to t ). the average time to meet the nutritional objective was days (range to ). the selected formula were: lactose free infant formula in patients, elemental diet in , pediasure in and jevity in . we recorded vomiting in patients, diarrhea in and constipation in . none of them forced the en suspension. we didn't recorded any case of ~spiration or pneumonia. we found no relation between drugs, selected formula and complications. we think that en may be a safe and well tolerated procedure at the picu setting. generation of free radicals in iipid emulsions used in parenterai nutrition (pn) has been described recently. this seems to be due to high polyunsaturated fatty acid content. we studied lipoperoxidation status and antioxidant parameters presurgically and through the postoperative period ( days) in a group of twelve children (ranging from months to years of age), who underwent heart surgery and received nutritional support with pn. malondiaidehyde (mda), an end product of lipid peroxidation was used as a marker of oxidative stress. it was determined by the yagi spectrofluorometric method. antioxidant activity was measured with an assay based on the inhibition of spontaneous autooxidation of a brain homogenate, and vitamin e in serum by the technique of hansen and warick. erythrocyte mda release fonowing incubation in hzo in vitro was used as a functional measure of tissue vitamin e levels. data were compared using one-way variance analysis. there was not significant differences in the plasma mda production among the values obtained before surgery ( :t: . nmol/ml), postoperative pre-pn ( . +_ . nmol/ml) and through the course of pn ( . : . nmol/ml). levels of vitamin e previous to surgery were , : . #g/ml and decreased to . _+ . #g/ml at hours post-surgery before starting pn. while receiving pn, concentrations of vitamin e increased progressively up to + . l #g/ml (p = . ). pre-surgicai plasma antioxidaut activity values ( :i: %) dimiuished in the first postoperative day ( _+ . %) and then showed a clear increasing tendency, directly correlated with that observed in vitamin e levels. the measurements of the maximal percentage of mda release of erythrocytes in vitro, showed an inverse relationship with plasma vitamin e levels and with the plasma antioxidant capacity: preoperative . + . % and postoperative descending from . + % to _+ . %. our results indicate that during surgery and within hours postsurgery, a decline in antioxidant defenses occurs, in the postoperative period, while patients are on pn and taldng into account our obtained data: the unaltered mda values, the rise of plasma vitamin e levels, the recovery of both functional erythrocyte membrane antioxidant protection and the plasma antioxidant activity, they do reflect that during the intravenous administration of lipid infusions, oxidative damage does not occur, probably due to the addition of vitamin e to pn solutions in order to avoid autooxidation of lipid emulsions. garnitine is one of the essential nutrient in the diet of newborn infants. mother's milk represents the natural eamitine source for the newborn infants. we studied enteral earnitine intake in premature infants eutrophic (eu) and hypotrophic (hy), fed with mothers milk trough consecutive days-from fifth to tenth day of life. mean gestational age was (range - ) weeks for eu and (range - ) weeks for hy. birth weight was ranged - g for eu and - g for hy (p < , ). breast milk carnitine concentration was , i.tmol/i in mothers milk delivered eutrophic babies, and singnificant higher mean coenetration , l.tmol/i, of hypotrophic premature infants (p< , ). breast milk earnitine concentration was ranged between , - , ~moi,'l, and daily carnitine excretion was between , - , p.mol/i, mean , +- , i p.molll. the daily mean carnitine intake was from - , mgkg/d, in eutrophic and , - , mg/kg/d, in hypotrophic premature infants, in milk volumene intake from - mllkgld., the results of this study suggest that premature infants should be fed with own mothers mitk in eady neonatal pedod in attempt to prevent camitine deficiency. keto-acids and parenteral lipid admlnistration. castorina m., antuzzi d., ricci r., rendeli c., polidori g. pediatric intensive care unit -catholic university -roma (italy). some metabolic studies have suggested that the administration of mediumchain tltgticerides (mct) might induce a marked increase in ketone body concentrations; the aim of this study is to evaluate the effective ketogenetic risk of infusing mct emulsions in total parenteral nutrition (tpn). two groups of seriously infected children were examined: a- septic infants were given to tpn with a % of long-chain tryglicerides ( lct). b- septic infants, in wich the lipid source in tpn consisted in a - mixture of lct and mct. the children in the different groups showed similar severity and therapeutic scores (prism, tiss, ~, f~ , f~ ); the lipid intake was the same, corresponding to - . g, ckg bodyweight/day. in all patients a sample of urine was taken six times a day for the whole time of tpn the urine samples were screened by the diphenylhydrazone test, and the ketoacids excretion was confirmed by a chromatography. the urinary excretion of -keto-acids equivalent to p-ohphenyl-pymvate (mcg/ml urine) is summarized in the no significant increases in frequence and/or in severity of acid-base disturbances were found in all studied patients in consequence of the lipid infusion. the patients of b-group showed a lower excretion of -ketoacids. the c~-keto-acids excretion was ever unsignificant and seemed to be correlated with the illness severity more than with the lenght of the tryglicerides chain. this observations let us suppose that the mct, at employed doses, can be safely administred also in childen with metabolic acidosis and/or serious illness. reye syndrome cayetano heredia hospital, lima, peru five patients with reye syndrome were studied; included were those diagnosed from january to march and treated at cayetano heredia hospital. the age of presentation varied from z. to months. the syndrome occurred more frequently in males ( / ); the time of illness at the presentation varied from to lo days, and the following features were found: fever ( / ), akeration in mental status ( / ), seizures ( / ), gastrointestinal illnesses (diarrhea) ( / ), and respiratory insufficiency ( / ) --in this case ventilator)-support was needed, in all cases hepatomegaly, intracranial hypertension, hypokalemia hyponatremia, metabolic acidosis, hyperammonemia and elevation of hepatic enzymes were found. coagulation blood tests were abnormal in three patients. cerebrospinal fluid (csf) showed hypocellularity in all cases (less than cells/ram ). the cultives were negative, and the final diagnosis was confirmed by hepatic biopsy. no deaths were due to reye syndrome. a contribuhon to the study of reye's syndrome the aulhors analysed records of patients meeting all the clinical, biochemical and anatomic criteria put forward by hutrealoch~ and samaha ( ) who were ireated at the emergency and resusdlation depammlt tithe ipch in years. the average age of the children was years (the ~umgest monks and the driest year~ most of lhon ( / ) came from lhe countryside to lhe lmfia~ willfin hours of file oulbreak. a few days previously the patients w¢~ld have fever (some of ahem wilh diarrhea or cough) ,,rod vomit then rapidly fidl into coma and c~nvuksien. they ~me to the imtia~ with the signs of typical increased inlracranial pressure ( stages - according to lovejiy for most cases) but the liver was unllkely big. the main biocheafical dmnge of the blood was acute hepatic thilure -the bllod glucose decreased : (ha the aver~ , n~% ( ) ~ not measured in five cases (glucose a"aces). -the blood ammonia ino~ased riven to microg/dl fin lhe average , n = ) -the percentage of p~in was low, in the average , % (n-- ), % at the lowest (a case ofmekaa lasling five days was ctwed with vitamin k). -got and gpt enzymes increased - times (n= ) besides, tht~re was decompcmated addt~is with lhe average valnes ph = . ; pcoz = ~ mmhg and eli = - ~ ~, (n~). the charadaislic dumge in the ¢~'ebrospinal fluid was a low glucose tx~acenwalien h the average of k mg/dl ( cases of glucose it'aces) ; meanwhile protein decreased in the average of mg/dl ( cases below mg/dl). besides ~here was no inflammatory lesion ~tion) : very serious brain oedona and fatty degeneration of live~ scatteredly or parllally.microso~ically (biopsy and necropsy) in file brain appeared bright space around blood arteries and glioma extended v'lrdmw -robin space, with no inflammatory reaction. liver.-the sa~dure was intact there was no " "mtlammal~s ' cell, the kupffer cells did not show hyperplasia but show a heterogenous deg~nerafion~eart: also showed a fatty degencralion in some areas of the myocard eellkidney: the ihtty degenerati~l scattetedly in the tubular cell. pano~as:fatty degeneration scattetedly in file pancreatic cell. two cases of liver biopsy for lhe rid lime (check before leaving file inslitute) found that fatty degeneration was r~tta~ nearty ff~pletely. the exact muse remained unknown. objectives: to analyse whether there exists serum and urine electrolyte disorder in children with respiratory failure, methods: we have made prospective study of children in our icu during a month period, electrolyte concentrations were measured in serum and urine collected during hours (sodium-na, potassium k, chloride-ci, calcium-ca, magnesium-mg and phosphorus-p). results: average values in serum were: na . +/- , (rv: - ) mmol/i, k . +/- . ( . - . )mmol/i), cl . +/- . ( - ) mmol/l, ca .t +/- . ( . . )mm /i), mg . +/- . ( . - . )mmol/i. average electrolyte levels in hours urine were: na . ~/~ . ( - )mmol/day. k . +/- . ( - )mmol/day, ci . +/- . ( - )mmol/day, ca . +/- . ( . - . )mmol/day and p . +/- . ( . - )mmol/day. conclusions: average serum ci and ca levels were decreased in children with respiratory failure. in urine, average k, ci, ca and mg levels were decreased and urine p concentration was increased. we concluded that serum and urine electrolite disbalance may be expected in children with respiratory failure. the nurse is going to kill me! (icu syndrome) anita duvndam ~, sonia v.d,sluis ~dpt. of pediatrics, div. pediatric intensive care, sophia children's hospital, rotterdam dpt. of pediatrics, div. pediatric tntensive care, juliana children's hospital, the hague. content description: this presentation will provide the critical nurse with background information concerning the identification, the prevention and management of the critical ill child who has developed the icu syndrome. the results of a questionnaire concerning the occurrence of icu syndrome on a picu will be presented. behavioral objectives: at the end of this session the participant will be able to: . describe four clinical symptoms of the tcu syndrome, . identify major sources of picu environmental stress, . discuss nursing's unique role in helping the child to cope with the icu syndrome, . discuss nursing's unique role in preventing icu syndrome. content outline: the icu syndrome is a situation in which the individual is not able to deal with changes in observation and interpretation of both quantity and of patterns of sensory perceptions. in other words the borders between the inner and outer world becomes unclear for the person. the exact incidence of the icu syndrome in the critical care setting is unknown, we have limited knowledge of the occurrence of the icu syndrome in critical ill children. yet critical ill children in our hospitals sometimes show the symptoms of icu syndromes. to be able to identify the occurrence of icu syndrome we developed a questionnaire. the thesis of the questionnaire was: does the icu syndrome occur in critical ill children in the age between three and ten, who have been intubated and/or ventilated in a picu? eighteen questionnaires have been send out to parents of critical ill children in two hospitals. the response was percent. conclusions: . most of the children were anxious (n= ), showed regression (n= ) or had sleeping problems (n-lo) during the stay on the picu. . children showed the same problems after discharge. . there is no relationship between environmental factors and symptoms of the tcu syndrome during the stay. . there is no relationship between preexistential behavior and symptoms of the icu syndrome during the stay. because the lack of a scientific definition for the icu syndrome in children and good criteria for diagnosis, we were not able to get an answer to our thesis. more research is needed. continuous veno venous haemofiltration (cvvh) was adopted as a first line renal replacement therapy in our paediatric intensive care unit (picu) some three years ago. ~/he process of introduction and development of cvvh proved to be an exciting challenge for nurses as indeed it was for the whole multidisciplinary team involved. we have successfully used cvvh in the treatment of over infants and children, weighing between - kg. the range of conditions treated is ever increasing. to date we have not only used cvvh for patients in renal failure and fluid overload, but also to gain biochemical control in tumour lysis syndrome-and metabolic disorders. other distinct patient benefits in comparison with more conventional means of renal replacement therapies are that, it is a continuous controlled treatment being extremely effective in creating 'space' for nutrition, which is especially important in the fluid restricted, catabolic patient. of paramount importance in providing this level of service is the education and training necessary to impart the skills and knowledge for nurses to become expert practitioners in this field. we now have a teem of highly advanced practitioners who have developed their nursing skills to provide even more holistic care for their patients. this group are embracing new challenges and responding positively to the developing service whilst expanding their knowledge base. this paper will discuss the advantages and disadvantages of cvvh as we have experienced, relating i t to the wide variety of patients we have treated. we wish to share how cvvh has become an accepted role of the picu nurse and how the service has been successfully implemented. integrated care pathways (icp's) define the optimum course of events in the care of a patient with a specific condition, within a set timescale. aims and methods: icp's and case management (cm) were introduced to evaluate and improve clinical practice. icp's were developed for patients undergoing surgery for atrial septal defect (asd) ventricular septal defect (vsd) and fallots tetralogy. they were based on the median experiences of the last patients. patients have been managed using icp's. results: analysis of variation from the icp's shows a reduction in the following potentially avoidable causes of variation: delay in extubation has been reduced from % to %, and patients ( %) were extubated earlier than the median. prolonged stay on the intensive care unit (icu) has decreased from % to %, and patients ( %) were discharged to the ward a day earlier than the median. the number of patients receiving inadequate post operative analgesia has decreased from % to %. delayed feeding after operation has been reduced from % to %. unavoidable delays in extubation and discharge from icu occurred in patients ( %) because of haemodynamic instability" or lung problems. cm utilising individualised pathways for these patients has reduced variation in care which can improve patient outcomes. pathways have been developed for other conditions and it is anticipated that approximately, % of patients will be treated using an icp revision of icp's results in continuous evaluation and improvement of the care provided, conclusions: the use of icp's and cm has improved patient care and decreased avoidable delays and variations. the future development of other icp's, combined with a case managed model for selected patients, is seen as a viable method of continuously improving patient care. this paper is a retrospective audit of children who received continuous veno venous haemofiltration (cwh) whilst on the paediatrie intensive care unit (picu), data was collected over a month period from may to january . the girls and boys were aged from day to ½ years (median ½ years) and weighed between , and kg (median . kg). length of admission to picu was to days (median days). sepsis syndrome was diagnosed in cases: of these had bowel necrosis and meningoceccal disease. eight patients had an underlying haematological, oncolngical or immune disorder. the remaining children had inborn errors of metabolism ( ), nephrotic syndrome ( ), or cardiac failure ( ). cvvh was instituted for a variety of reasons. muttiorgan dysfunction was present in % of patients, % had acute renal failure, . % tumour lysis syndrome, . % uncontrollable metabolic acidosis with hyperammoneemia and % required fluid management. treatment was continued for between hours and days (median ½ days). haemofiltration was successful in achieving its desired effect in all except one patient. . % of children with haematological, ontological or immune disorders died despite successful haemofiltration. survival was higher in cases of septicaemia ( . %), nephrotic syndrome ( %) and errors of metabolism ( %). overall mortality was . %. this was attributed to the severity of the underlying disease process rather than the effectiveness of cvvh as a renal replacement therapy. asynchronous defibrillation and synchronized cardioversion deliver direct currents of high amplitude through the chest, to stop ventricular fibrillation or to convert life-threatening arrhythmias. since the human body is partially conductant, it should be possible that after a brief delay of time, a derivation of this monophasic defibrillation pulse is measurable in regions of the defibrillated body, other than the traject between anodal ("sternum") and cathodai ("ape~;") paddles. one could also estimate that health care personnel in the immediate environment of defibrillatoty shocks, are always at risk to possible electric injury. accidents might occur when health care personnel do not stand clear from the patient during firing and create an additional electric path from this patient through their own body. most likely, the nonisolated parts creating a transversal eiectric path, will be the hands of the caretaker. since defibrillation generates touch voltages up to v and - a in ved, short delay's ( to millisec), the total body resistance during skin to skin or skin to paddle contact, is calculated to be as [ow as ohm (percentile tank for the entire population). an experimental protocol was developed to evaluate the conducted currents during defibrillation. mature pigs with a weight of up to kg. were used as animal models. the:,, had barbiturate anaesthesia, inotropic support with dobutamine at l micmgrams/kg bodyweight and all had a sinasat rhythm at the begining of the test-rounds. synchronized defibrillation at j (approx. . j/kg) and asynchronized -at j (whenever ventricular fibrillation or ventricular tachycardia occurred), were attempted, through latero-lateral placed and firmly pressed defibrillation paddles and/or adhesive multi-function electrodes. gel pads were choosen as contactmedium to cross the skinpaddle barrier. subdermal electro-myography needle electrodes were put at different parts and regions of the pig's body, but not between the paddles. these electrodes were coupled to a resistance device of ohm and a measuring computer. we measured monophasic and low current pulses up to , a during the defibrillation burst. thus conducted currents are high enough to produce accidental electric shocks when additional electric paths are created by the caretaker. "llaese are usually harmless, but in some "worst case" (wet or transpiring hands, rings,...) or in association with defectuous equipment, peak currents might be harmfull, and can produce pain, bums, apnae or cardiac (transient or persistant) arrhythmias. maria skogbv. karin mellgren, lars-g ran friberg, g sta mellgren, hans wadenvik. g teborg, sweden. bleeding complications in extracorporeal circulation is partly due to perfusion-induced platelet dysfunction. the aim of this study was to evaluate an in vitro model for investigation of platelet function parameters in an extracorporeat system. study: two complete extracorporeal life support systems were perfused with fresh heparinized human blood for hours. piatetet membrane glycoprotein changes, platelet granule release and platelet count were followed. eight paired experiments were performed. one of the circuits was perfused by a roller pump (polystan) and the other by a centrifugal pump (biomecicus), other components were identical. results: a continuous increase in glycoprotein (gp) ib-negative platelets was seen in both circuits. a marked increase of plasma beta-thromboglobulin concentration and a decrease of the intracellular pool of serotonin was observed, indicating a marked release of alpha as well as of dense granules. the plasma concentration of glycocalioin increased in parallel with a reduced platelet surface expression of gpib, suggesting that the loss of gpib is caused by proteolysis rather than by a downregulation of this receptor protein. conclusion: hours of ecls perfusion induces pronounced platelet degranutation and causes changes of important membrane receptors. this is in accordance with clinical studies, suggesting that our in vitro model does mirror the platelet exhaustion observed in a clinical reality. no significant difference was observed between the two pumps. the purpose of this study was to examine signs of distress exhibited by several patients following the discontinuation of opioids and benzodiazepines (o & b) in a -bed pediatric intensive care unit. the authors compiled a list of all cases of possible withdrawal reactions reported by nurses in the study setting over a one-year period. five cases were selected for study in terms of their wide diversity of relevant circumstances. the cases were examined through a retrospective chart review. data pertaining to analgesic and sedative administration, along with nurses' reports of behavioral distress, were transcribed and coded. a remarkable pattern of behavioral distress was clearly associated with discontinuation of o & b. cessation of benzodiazepines was associated with severe distress. these reactions were characterized by various combinations of crying, irritability, tremors, grimacing, gagging, vomiting, or feeding problems. some of these persisted in spite of large amounts of bolus drug administration. these signs were manifested for to days following cessation of o & b. these findings demonstrate that the rapid weaning of o & b infusions, sometimes following short-term therapy, can cause severe withdrawal reactions. the particular course that a specific patient will follow will likely be modulated by underlying manifestations of "icu psychosis" and unresolved pain and emotional distress. hermana tezano mt, pilaf orive j, latorre garcia j, lizarraga azparren ma, lopez bayon j ucip hospital de cruces. bilbao. spain a female child, one year old, was referred to our unit from another hospital because she had ischemic syntomsjn her right forearm wich started three days before. she had ~i downs syndrome and fallot's tetralogy with a systemic-pulmonary by-pass. nine days before the admission in our hospital she was undergone to a cardiac catheterization by arterial and venous femoral punctures with no incidences. on the admission to the picu her forearm was cold and pale with absence of cubital and radial pulses and slow capilary filling. treatment with heparine was started unsucessfully in the firsts hours, so uroquinase was added that later was changed to rtpa plus brachial plexus blockade. by the time me arterial tlux became more and more evident (doppi~) il bccuute aiat) evident a regional aedema witch made necessary a fasciectomy &the wrist. she also developped necrotic delimitted areas that included her first and fifth fingers. she was discharged from the picu after days and a week later she went to the operating room where an amputation of the distal phalange of the fifth finger was made. the etiolgy of this ischemic picture can't be attributed to the catheterization itself, but probably it should be due to some attempts in arterial puncture in its course, attempts that could be guess by the little marks noticed at the elbow flexure when she was admitted. long maintenance of cardiovascular funtion by assisted ventricular device with membrane oxigenator hermana tezanos mt, pilar orive j. latorre garcia j, lizarraga azparren ma lopez bay n j. ucip. hospital de cruces. bilbao. spain a little girl of three and a half years came to the intensive care unit from the operating room. affected of great vessels transposition with a pulmona~' banding corrected vith rastelli technique, it became impossible to discharge her from the by-pass circulation. it was assumed that this factt was due to a transient myocardial disfunction dfter the surgeon x~ent through the technique and found no wrong step in it. at the picu admission she had an ejection fraction belox~ % (echncardiografy), and was manteined on veutricular assistance with a circuit consisting ofa plate's membrane oxigenator, a bio-medicus pump and pvc rubber tubes, with monitoring of pulmonary', right and left atrial pressures. initially she needed a ventncular support of . l/min with dopamine. dobutamine and adrenaline; gradually the ejection fraction rose to % aliowing a decrease in the mechanical and inotropic support. the organic function ,.'.-as preserved but x-ray of the thorax showed images of pulmonary aedema with an increase in the needs until a fi of . . even though she was placed in a transplant program, she was remouved when the ejection fraction rose to % on the lth da of the evolution. there were no signs of infection (profilactic coverage). on the fifhteen postsurgery day. with a good cardiac sound and an ejection fraction of at least %. it was considered the withdrawal of the ventficular support, but unsucessfully despite the increase ofinotropic drugs, so she died. we think that our patient is a good example of the posibiti~ to mantain x~ith ecmo during a prolonged period of time a potential transplant receiver without major complicatios. joan wierema; ma )an mourik. sophia children's hospital, department of pediatric surger,j, rotterdam, the netherlands. the success of an ecmo program is heavily determined by the organizational structure and the daily availability of both well trained nurses and physicians. until now there is no general guideline to determine the optimal training schedule to set up an ecmo program. objective: to evaluate the set-up of an ecmo program in an area without direct availability of perfusionists of cardiothoracic surgeons. description of the progress: during the preparation of the ecmo program which started november , different phases are identified. first phase, acquisition of experience by training abroad of one physician, staff member of the icu, physician acting as a fellow and one icu nurse with theprimary task to serve as an ecmo coordinator and train the nursing start. in the second phase animal experiments were performed by paediatric surgeons, staff physicians, fellows and nurses. third phase, start of the actual ecmo program, priming by one trained nurse and ecmo fellow, daily care of the patient by nurses, icu nurse taking care of the patient related activities, ecmo trained nurse taking care of the circuit. fourth phase, transition of the tasks for priming the system from the ecmo fellow to the ecmo nurse and changing daily care for the ecmo patient now including the circuit by trained icu nurse with special legislation for ecmo. ongoing training of to nurses working for at least half a year in the icu. during an ecmo run direct contact between the ecmo nurse and one of the staff members; patient data: in a year period neonatal ecmo cases were performed with an overall mortality of %, ranging from over % in meconium aspiration to % in congenital diaphragmatic hernia. in other patients pediatric ecmo (age range month - months) mortality was %. conclusions: ecmo can be established without perfusionists or cardio surgical back up once a predetermined training schedule is available, resulting in comparable results with the international ecmo registry both for neonatal as well as for pediatric cases. mechanical ventilation at home makes normal development of the child with chronic respiratory failure possible. the parents must be encouraged to accept the treatment at home; therefore, they should be enabled to take care of their children all by themselves and the continuous professional support has to be offered to them. it must be stressed that the nurse is the link between the diseased child and his family introducing them into the process of the medical care at home. all the equipment needed for the mechanical ventilation of the child at home (ventilator, suction device, pulse oxymeter, oxygen tanks) has to be provided before the child is discharged. it is paid by the national insurance company. the maintainance service workers will make the necessary installations available and they will renew oxygen and compressed air weekly. the ventilation assembly must not interfere with the childs activities. the related dispensary and nursing service should be notified of the child's condition and the list of the required material (suction tubes, cannulas, desinfectants etc.) has to be made. our experience is so far -from the nursing and medical point of view -satisfactory, however the social status of the parents needs further regulations, omphalocele. mourik m, sophia children's hospital, department of pediatric surgery, rotterdam, the netherlands. in patients with (giant) omphalocde operative closure is impossible due to the lack of intra-abdominal space and a variable degree of pulmonary hypoplasia. consequently conservative treatment allowing epithelialization is the treatment of choice with a high risk of infection and/or sepsis due to a sometimes very long stay in the icu. this conservative treatment consists of daily application of an antibiotic containing powder on the cele which is covered by sterile gauzes. furthermore the enteral intake is started as soon possible. objective: to evaluate whether this treatment renders a high risk for serious infections. patient characteristics: in a year period patients with a giant omphalocele were admitted; mean birth weight g ( - g) mean gestational age . ( - weeks). overall mortality ( %). patients had to be ventilated for a median duration of t days ( , - ). results: patients were discharged at the mean of days following birth ( - ) following complete epithelialization of the omphalocele. following the initial stabilisation period parents were involved in daily care of all patients even during artificial ventialation. enteral intake was started at a median of days ( - ). colonization of the omphalocele was observed within - days in all patients. although infectious periods were observed at regular intervals, sepsis was the primarily cause of death in only of the patients who died, in conclusion: nursing care to prevent sepsis in patients with (giant) omphalocele is feasible for a long period of time and can be performed with simple measures, since june , we have been using nasal cpap in the treatment of respiratory disorders in newborn infants (severe apnea syndrom, tachypnea, hyaline membrane disease). in newborns presenting hyaline membrane disease, we have used cpap and administered surfactant therapy to premature babies out of . clinical data were : mean ga : . + weeks. mean bw : + g. mean administration time was hours. before administration, mean fio was . + . , mean pao . + . kpa and mean a/ao ratio . . all babies were intubated for administration of surfactant (curosurf) and were extubated after half an hour to hours after administration. this treatment failed for seven babies and they were ventilated by oscillation ventilation ; all babies survived. complications : we have observed pneumothorax in cases and cerebral hemorrage. mean duration time of nasal cpap was hours for the babies without assisted ventilation. nurses in charge of babies with nasal cpap should be aware of neonatal care, of the possibility of surfactant administration and of complications during this type of treatment. therefore nurses should know very well the use of nasal cpap, the monitoring during this treatment, the fixation of the system on the baby, nursing of the babies during this treatment and finally should take care of the baby physical and psychological well being. as a conclusion, nurses in intensive care neonatal units should know the possibility of treatment of rds by nasal cpap and should be aware of baby-nursing. emco in the postcardiotomy infan_t : a simplified circuit and reduced management grillet lsabelle, bosson christa, goelz andrea, helmer pascale, icu nurses, tschaut rudy perfusianist aldo castaneda institute, clinique de genolier, suisse to improve postoperative survival of infants with repaired cardiac tessions but severe residual cardiopnlmonary dysfunction, emco was used in infants, ranging in age from l days to months and in weight from , kg to , kg. diagnosis included tga / intact ventricular septum ( ) (both subiected to rapid arterial switch operation because of unsuitable lv function) and tga +vsd ( ). "['he emco circuit included a sarns roller ump, a medtronic minimax plus hollow fiber oxygenator and tubing ( / - / inch) with bioactive surfaces, an arterial canula fr., venous canulas : right atrial fr, left atrial fr., an air oxygen mixer and a sarns heater. the act was kept between - sec. after the infant was connected to emco by the surgical team and the perfusion technologist, the patients were being cared for and controlled exclusively by the cu nurse assigned to the patient and the physician on call for the icu, pump flows of - ml / kg / mln were targeted to insure an adequate urine output, a brisk capillary fill and physiologic left and right atrial pressures. sodium nitroprusside was used to control systemic hypertension while all other inotropic drugs were discontinued. inspired oxygen fraction on the ventilator was reduced to , at a respiratory rate of - breaths per minute. body temperature was maintened close to normal. when the heart started to eject again, inspired oxygen was increased to about , . the preparation for weaning from emco was done by the icu nurse: including oxygen fraction and ventilator rate. weaning from emcowas coordinated by the medical / surgical team and the perfusion technologist. the duration of emco in the patients ranged from to days. three patients ( %) were saccessflly weaned from emco. one of the three died within hours from neurologic complications, unrelated to the emco. the other patients ( both with tga + vsd ) are long term survivors and are doing well. neither the survivors nor the patients who died bad hemorrhagic complications, despite the fact that of the were placed on emco because of failure to wean from cardiopulmonary bypass. the simplified emco circuit worked reliably throughout this experience and no complications occurred nor could any shortcoming be ascribed to the use of this reduced, cost effective management team. years of praciical experience wiih exiracorporeal membrane oxygenation (echo) -the viewpoint of the nursing siaef monika schindler in a training program was started for the introduction of echo in the kinderklinik of mannheim/germsny. years later ( ) the first european echo patient, a days old newborn baby, ~as treated successfully in our institution. with echo, more treatment modalities ~ere available after failure of conventional therapy in eases of severe lung diseases like meeonium aspiration syndrome, congenital diaphragmatic hernia, sepsis/pneumonia, primary persistent pulmonary hypertension of the newborn and -in infants and children -ards (acquired respiratory distress syndrome) due to multiple underlying illnesses. in the first ards patient, a years old girl, was treated successfully ~ith ecmo in mannheim. until now patients (age: i day - years) were treated with echo in our institution; additional patients were transferred to us from other institutions for therapy with echo, but could be treated ~ieh alternative moda]ities of therapy like improved conventional ventilatory support, high frequency oscillatory ventilation (hfov) and inhaled nieric oxide (no), unexpectedly the more ~ide-spread use of hfov and no -also in other ~ntensive care units -did not lead to a reduction of the echo frequency in our hospital. due to this long ecmo practice, which is helpful in many critical situations, a certain routine came up in the nursing staff of our intensive care units; but ecmo continues to be a maximum strain for all co-workers and implies an high personnel and material/financial-expense. but in our opinion the improved survival rates in severe respiratory failure-( % in neonates and % in infants and children, estimated survival rates under continued conventional management: < %) by the use of echo justifies this expense, even in times of less financial support for public health. • analyse the differences between each urdt. method : ni criteria were defined by the rt~a ped group according to cdc criteria. all data were collected by a nursing and medical team. data vere dealed with epi info software. all u-dections data were validated by an external mvestigatol : patients who stay more than hours in an icu were included over a months period. % were newborns, % infants and , % children. ni were idenlified among sll patients, the overall ni incidence rates was , % and , person day, septicemia was the first cause of ni ( %). staphylococcus epidermidis were isolated in % of septicemia cases. pneumonia was the other main ni ( %) with gram negatwe bacille isolated m % ( % of them beei,ng fseudomonas) accordmg to age, the septicemia mcidence rate varied from , ~., to . %,, catheter/day and pneumonia incidence rate from , %, to , % ventilation/daywith the lowest rate for newborns. : this survey was possible thanks to the collaboration between laboratories, bacteriologist, physicians and nurses, and allowed each concerned unit to work together in~tead of every one in his own field. 'lhe results of this survey bring changes in attlhades and empower the different team work the next step is to analyse the ditferent nurses procedures of indwelling central venous catheter in each unit ar)d implement a ni quatity care. program in all nicu anti picu. v~,~jl~j~'v~y : new born in~ra-v~ nous drugs ~md lheir pr~taration mcounter a lot of risk ~)f utters, due to inadapted conditionn~ng to podia)eric and minimal amount injec~.~d, th~ study t~ed to evaluate degrees of do.~age er~o~ dye tt) dru~ preparaek, n i~ nc,:,~atol~gy and to come out w~th ~ method of dihttion able to reduce risk to minima (errors _< %) at the ~eme t~me, ¢o~,~x-tueaces on cost were studioj methnd~ : amikacine was choosen because this antibiotic is regularly ~,~l in neonatology unit and its dosage is easily used by standardiaud method, amikacine doses ( , mg doses and , mg doses) were prepared" by nurse~ form mg for m) vial. each dose w'a~ diluted to obtain mi volume, tallowing usual ward method, for each dilution, amikacine con<entrati('m and it,s volume was measured and used equipment for preparation wa~ regi~tred. "first s utj(m" were prepared diluting ~t'~ mg amikacine pouder inn ) ml glucnse solution bag (viaflex baxter bag with transfer cap), from tho~e "first solution" doses at mg and doses of ~ rng were prepared, amik~cine coc, cenir,)tmn for each preparabon and their volume were measured, ai,~n ntttl~,ber of seringe were tounted. first soiuhon staeiiity was conrroieo aher one week conservation at + '>c. amjkacine concentration were measured by fluorescence process (tdx abbott) after sample dilurion. on a mg/l sample, tovhnical reliahility show~ > ~ % of result mpmductlon and < % of variation due to dilutions. results : when amikacine injection werv pro.pared from araikacme /) mg for mt vial > % do~ge, ermr~ were found in / cases ; ~ % in ,t ,to cases. if preparation is done from amikacine "~it'st soltltion", les.--concenvr~tcd, it i~ more preci,,,e and only one dosage error ~ % ( , %} is found in eli studied doses. in add)inn to )hal if doses were wep,m-'d from one "first soiatiol~' bag, the cost economy sl~ouid b~" of fr~, and ii dos~$ were prepared tram the same bag the saving mtmey should be o{ i its .cencluslon : .ur survey shows th~t h' ntu)nato|ogy the u~ of a "first sohation which can be kept fi~r one week is enable to reduce dosage erroes and i~ co,~tsavmg, regarding [,v. admimst'rahon method the survey is still on, introduction: so-called vein of galen m~iformations ale rare in~racranial embryologycal anomalies, repl~senti~g tess than of symptomatic intracranied artefiovenoas l~alform~tions. the spontlneous prognosis is ~s~u~lly fatal, because of cardiac frilure due to left-to-right shunt thrq~ugh the fistula. recent developments of new techniques of treatment of the malformation and its cardiac consequence have led to a revolution in the practical approach of children w~th galen malformation. our fukfose is to contribute, with our persoaal series of s newborns and infal~ts admitted in our unit after endov~,scular embolization, to a better management of these children. such a management requ!res a rnultidisciplinary approach. intensive care are required prior to embollzation for patients with cardiac failure or cardiogenic shock and after cmbolization in order to insure cardiac and cerebral hemodyna.mic stabilities. this overlooking suppose for the nursing team to understand: prior to embolization : heart failure and cardiogenic shock. after cmbolization : evaluation of neurological and hemodynamic consequences of this proccdure, without forgetting the nursing and psychologic aspects, in concl'iision, this last ten yerrs, these new approaches give to the patients and their famitiy a good reason to hope a total recovew, in our exl)erience, the global mortality is % aad % of children #j-e neurologically normal after embolizafion, ii ii~ i ~ii i ii i i l i iiii~ i ~i iii i background: venous oxygen saturation (svo z) reflects the residuai oxygen after tissue oxygen extraction and represents the relation between tissue oxygen supply and demand. we studied svo and arterial lactate during progressive isovolemic anemia to assess the relation between svo and tissue hypoxia. subjects: ten - day old anesthetized ventilated piglets sao and svq were measured continuously by a fiberoptic catheter (oximetrix, abbott lab.) in the carotid and pulmonary a~epy tissue hypoxia was confirmed by a reduced vo, and an increase in lactate. conclusion: svo reflects better a reduced dp obtained by progressive anemia surfactant replacement improves gas exchange in early-stage adult respiratory syndrome (ards) [ , ], but not in late-stage ards [ ] . we report the first case of successfull treatment of ards after repeated instillation of surfactant.a ten year old boy, weighing kg, presented with hemorragic shock. biphasic-positive-airways-pressure ventilation was performed (evita ii, dr~ger, germany). he had recieved nine units of packed red blood cells and underwent surgical exeresis of two bleeding gastric ulcus. post-operatively, a cardiac arrest required cardiopulmonary resuscitation for three minutes. hemodynamic status was subsequently stabilised. the chest-radiograph showed infiltrates of both lungs without signs of cardiac failure. on the third day, the patient became severely hypoxic with a pao /fio ratio of . gas exchange was not improved by high ventilator settings. peak inspiratory pressure (pip) and ventilatory rates were cmh~o and breaths/min respectively. inspiratory:expiratory time was : and the positive end expiratory pressure (peep) cmh . after increasing the peep level to cmh , we instilled over minutes, mg/kg of porcine surfactant (curosurf, serene france), in two equal volumes in both main bronchus,the spo~ rose to % within rain, the fie could be reduced to . . twenty four hours later, gas exchange worsened again (pao /fio ratio ). we increased the peep from to cmh , and instilled a second dose of surfactant ( mg/kg). again, fie could be reduced within minutes (spo ; fie . .). the patient was weaned from the ventilator and extubated on the tenth day. follow-up at four month showed normal lung function.we demonstrate improvement in oxygenation after repeated exogenous surfactant administrations. we assume that in early-stage ards, surfactant may potentiate shunt-reducing effect of peep as it has been demonstrated in experimental model of ards [ ] , and allow decrease in fie . in case of secondary deterioration, we think that a second dose of surfactant should be administered. . weg jg, balk ra, tharratt rs, et al. ,lama : : - . . spragg rg, gillard n, pdchman p, et al. chest t : : - . . haslam pl, hughes da, mcnaughton pd. et al. lancet : - . . huang yc, caimulti sp, fawcett ta, et al. jappl physiol : - % (ref) . the aim of this study was to verify these data: patients/~lethods: all pts admitted to our multidisciplinary nicu/picu in were included if they were in respiratory failure recruiting conventional mechanical ventilation (cmv) with peep >_ and 'fig -: % or high-frequency oscillation ventilation (hfo) with mean airway pressure _> t cm h for or more houm. diagnosis, maximal ventilatory parameters, barotrauma, organ/ system failures, mechanism of death and glasgow oulcome scale (gos) and months after study entry were prospectively collected. results: patients were admitted to the unit, o whom required mechanical ventilation for a mean duration of . days. overall mortality was %, patients fulfilled study criteria. survivors had gos , pts with preexisting neurological impairment survived with gos . neonatal diseases included hyaline membrane disease ( ), meconium aspiration syndrome ( ) and cardiovascular surgery ( ), pediatric diseases included bacterial ( ) and viral ( ) pneumonia, aspiration ( ) and cardiovascular surgery beyond the neonatal period ( ). - ) . patients and methods: cefotaxim was used as a prophylactic agent in patients in life threatening situations (e.g. multitrauma, neurosurgery atc.). more than % children required cefotaxim for the treatment of severe infections (epiglotitis, meningitis, sepsis, pneumonia mainly in immunodeficient and neutropenic patients) in monotherapy or in the combination with the other antimicrobial agents. results: cefotaxim as a prophylactic drug was successful in all cases ( %). the effectivity of treatment of infections was . % ( patients). the change of antibiotic therapy required patients ( . %). patients ( . %) died, but only in of them ( . %) the obduction confirmed infection. conclusion: we conclude that cefotaxim is very effective and safe antibiotic and represents "golden standard" in the treatment of severe infections in childhood. in order to improve nursing quality, we recently adapted nursing care to the "five nursing functions" (activities of daily living, accompagnment in crisis, treatment, prevention and research) as described by the swiss red cross in accordance to the new educational guidelines of the european community, the aim of this study was to document complications of "treatment nursing function".methods: all treatment complications were prospectively collected by the nursing and medical staff. the nursing staff included patient (pt) name, time of occurence and exact description of complication, proposal for prevention and information of parents. the medical staff reported type of complication together with pt information, diagnosis, medication, treatment and interventions, outcome and referral, all complications were discussed in monthly meetings including nursing and medical staff.results: from january until december , pts were admitted to the picu/nicu for nursing days ( % of total bed occupancy). pts needed endotracheal intubation for an average of . days and pts required nasal cpap. complications in pts were noted ( per pi): inadequate check-up of equipment ; accidental extubation ( in intubated pts); bedsores ; false drug dosing ; wrong drug ; umbilical bleeding ; wrong transfusion setup ; nasal septal necrosis ). there was no mortality due to these complications. exact documention of treatment complications and their meticulous discussion within the medical and nursing staff may improve "treatment nursing function". however, documentation and evaluation of nursing within all "five nursing functions" will be nessecary in order to achieve optimal nursing care. cardiac output determination by thermodilution, using iced injectate has been shown to be valid and reliable in pediatric patients. it has been demonstrated in adult patients that there is no difference in cardiac output values when using room temperature injectate as compared to iced temperature injectate. the purpose of this study is to examine the effect of injectate temperature on cardiac output values in pediatric patients. our study consisted of sixteen pediatric patients who had oximetric thermodilution catheters in place after cardiac surgery and who had cardiac output determined using both iced and room temperature injectate. with each patient, cardiac output was measured once on the day of surgery and again the following day. in each case cardiac output was measured using both iced and room temperature injectate. statistical analysis included a two-way, repeated measures analysis of variance for each individual injectate administered and no significant differences were found in cardiac output. no statistically significant differences were found between groups with regard to the order of injectate administration or volume of injectate used (i,e., or cc's). the correlation coefficients between groups for cardiac output measurements at each injectate administration time, and for the average measurements across times, ranged between . to . (p < . ). preliminary data analysis suggests that cardiac output measurements for children are not effected by the temperature of injectate. a lenghty stay at a paediatric intensive care unit will always have sideeffects on a child's well-being and will put a high strain on the parents. in order to minimize the side-effects longterm intensive care unit opened in at the childrens' hospital. admitted children are all ~ongterm-ill and technically-dependent and the ventilatory support can alter from a tracheostoma to cpap or portable volume ventilator. nutritional support is applied by gastrostomies. a homelike atmosphere surrounds the children, they share a dormitory, a living-room and a dining-room the main purpose is to send the child home with or without technical equipment. this can only be implemented by giving structured education (theory and practice) to all categories involved. the multi-disciplinary team consists of one anaesthesiologist, head nurse, clinical specialist, rn nurses, nurses, one habilitation doctor, one social worker and therapists. twenty-four patients have been admitted to licu during these six years. length of stay was from one day to four years. four are presently staying at the trait. the assessment of pain in children ( - yrs) is still difficult, because children of this age have limited language and cognitive skills. to standardize the assessment of postoperative pain and distress in the intensive care unit an observational mstrument was needed that met several criteria. it should be easy to use in daily routine care. be suitable for the i.c. situation, and in children of - hrs of age. the comfort scale, an observational instrument designed to assess distress in infants in i.c. units, met these criteria. to accommodate the use of the comfort scale in the i.c. units and in research, nurses should be trained to use the scale. an additional requirement was that the inter-rater reliability should be sufficiently high, (cohen's kappa > . ). objectives: ) to introduce the comfort scale in the i.c.u.; ) to examine whether this instrument can easily, be incorporated into routine care; ) to investigate the inter-rater retiabtlity. methods: the comfort scale is an -item instrument specifically designed for use in pediatric i,c, units and contains both physiological items (heart rate, blood pressure) and behavioral items (e.g., alertness behavior, calmness/agitation, body movement, facial expression respiratory response, muscle tension). the observation period is minutes. the scale is supplemented with an item on crying tbr children who are not mechanically ventilated. groups of t.c. nurses were trained by means of video's and observations at the wards. after the training, each nurse completed scores with other nurses, after which the cohen's kappa was computed. when the kappa's for the items met or exceeded our . criterium, a new group of nurses was trained. results: to date, nurses have been trained. nurses find the comfort scale easy" to administer and a valuable addition to routine care in the i.c. unit. the cohen's kappa's were higher than . for all items that the inter-rater reliability was high. the comfort scale is feasible in postoperative care in the i.v. and is considered a valuable instrument to improve and maintain high postoperative quality of care in the i.c. unit. introduction:children with neuro-muscular disease are believed to have a higher resting energy expenditure (ree), because of their increasedwork of breathing.the influence of nocturnal nasal mask ventilation on energy metabolism and nutritional state of these children has not been studied so far.objective:l,ls the ree inereased? . s there an influence of nasal mask ventilation on the ree? .what is the nutritional state? .what is the estimated total energy expenditure(ete) in relation to the caloric intake? methods:a pilot study of patients( - years) .the following measurements were performed:l.anthropometry. .bioelectric impedance- .ree was measured by indirect calorimetry during the day (in bed) with and without nasal mask ventuation,ree was compared with predicted ree according to schofield(pee), .caloric-intake and activities were recorded during hour before measurement. .total energy expenditure was calculated as follows:measured ree x estimated activity factor. results:tin all children weight for height was too low, <p . .aii children had a low % of fat. .according to the healthy group, ree is not increased. ,nasal mask ventilation lowered ree in patients, in patient we measured a clear decrease.conclueion:l.anthropometry and indirect calorimetry were helpful in evaluating the nutritional state. .there is no increased ree,in t patient there is clearly effect of nasal mask ventilation. .when caloric intake was related to ree and level of activity patients had too low caloric intake. .the influence of nocturnal nasal mask ventilation and nutritional assessment should be evaluated longitudinally. increasing awareness that the neonate can percept pain and suffer following surgical procedures, has major impact on pain management. in our unit continuous morphine infusion (t /~g/kg/hr) was introduced from january as a standard medicatton following surgical procedures. before morphine was given as a bolus (i.m., i.v., rectal) of /~g/kg up till four times a day. repeated gifts of morphine were only gwen following observation by the nurse that the child experienced pain. aim of the study: to evaluate pain management before and tbllowing introduction of a continuous morphine infusion for postoperative pain in our icu. patients and methods: term born babies with isolated oesophageal atresia and tracheo-oesophageal fistula were included in the study. the total amount of morphine (/~£/kg/ hours); number of days morphine was given; duration of artificial ventilation was evaluated in a case control study. res ults: - - central venous catheters (cvc) have been increasingly used also at our picu, university medical centre ljubljana. besides advantages, the cvc has brought numerous risks of complications; the catheter sepsis occurs most often and can be fatal for the patient. for example, in we dealt with children aged - years or in other words . cvp per a child treated at our picu. a specialized unit for preparing parenteral natrition started to function in the university medical centre pharmacy in . after that we equipped a special room for preparing other necessary infusion mixtures and organized a team of nurses -"catheter nurses".their tasks comprise: managing cvcs, replacing and installing infusion systems and mixtures made at pharmacy, preparing drugs and some necessary infusion mixtures in special room, permanent training and co-operation with related disciplines, pharmacy, epidemiology, microbiology, etc. all registered nurses at picu have been trained for this work. the routine is as follows: in the morningafter the doctor's visit, we first check the cvc and change the dressings; according to the nurse's observations, the doctor decides whether there is a need to remove or replace the cvc. if the catheter site is inflamed, swollen, or purulent, a swab is taken for microbiological culture and the area cleaned. in case of systemic infection signs, cvc has to be replaced, blood culture taken and the infection treated with antibiotics. the dressings are always changed by two tmcatheter nurses together. afterwards the infusion mixtures for the next hours are prepared for each child. the infusion mixtures which will not be used immediately are kept on + °c; before application they are, of course, warmed to at least room temperature. finally the "catheter nurse" supplies the cvc with new infusion mixtures made at our pharmacy. our observations show that such work requires responsibility and is demanding; but our pemanent care lessens the number of complications. thus the described scheme fulfilled our expectations. perl- perative management of neonates with hypoplastic left heart syndrome muir warren, blondel maryse, foltz rhonda, farine martine, icu nurses, the aldo castaneda institute, clinique de genolier, switzerland both the pre-operative and post-operative physiology of patient with hypoplastic left heart syndrome is critically determined by the puhnonary-tosystemic resistance ratio. a high ratio results in minimum volume work for the right ventricle but is at the expense of important hypoxemia. a low ratio is reflected in minimum hypoxemia but an unfavorable increase in excess ventricular volume work. a resistance ratio of approximately results in an arterial oxygen saturation of % - %, a qp/qs of , and ventricular volume work only twice normal. this latter physiology appears best for oxygen delivery and systemic perfusion with tolerable volume work. carbon dioxide (co ) in the inspired gas is known to increase pulmonary vascular resistance without significantly influencing systemic resistance in the range of to tort partial pressure. hyperventilation with no co in inspired gas decreases pulmonary vascular resistance. thus, managing co may be useful to modulate the pulmonary-to-systemic resistance ratio, in the tast months, patients with hypoplastic left heart syndrome were managed by a norwood procedur e. patients with endotracheal entubation pre-operatively had co added to inspired gas to modulate the pulmonary-to-systemic flow ratio. patients had co added to inspired gas post-operatively. all but patient had characteristically large pulmonaary-to-systemic flow ratio with oxygen saturation mean % preoperatively, even in those patients receiving co , co was accompanied by elimination of metabolic acidosis pre-operatively. the most comlnon partial pressure of co added post-operatively was torr. po ranged from mmhg to mmhg. all surviving patients developed a metabolic alkalosis ( mean) when treated with co . there were deaths, one from drug toxicity, one from excessive post-operative hemorrhage, two from sepsis or mocardial insufficiency. only one patient benefited from catecholamine support which can increase systemic resistance unfaborably. in this experience co was felt to be an important adjunct in some patients pre-operatively and most patients postoperatively to faborably modulate the critically important systemic-topulmonary flow ratio, to maximize oxygen delivery and systemic perfusion, while minimizing excessive right vantricular volume work. key: cord- -i cahnxd authors: wendt, f. r.; de lillo, a.; pathak, g. a.; de angelis, f.; covid- host genetics initiative,; polimanti, r. title: host genetic liability for severe covid- overlaps with alcohol drinking behavior and diabetic outcomes and in over million participants date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: i cahnxd to distinguish correlation from causation, we performed in-silico analyses of three covid- outcomes (n> , , ). we show genetic correlation and putative causality with depressive symptoms, metformin use, and alcohol use. covid- risk loci associated with several hematologic biomarkers. comprehensive findings inform genetic contributions to covid- epidemiology, molecular mechanisms, and risk factors. host genetic liability and epidemiologic risk for severe covid- (coronavirus disease ) following sars-cov- (severe acute respiratory syndrome coronavirus ) infection is of immediate clinical interest [ ] . genome-wide association studies (gwas) of covid- outcomes identified several risk loci and provided evidence of the pleiotropic effects (i.e., a single locus confers risk to several similar or disparately related traits) shared with other human diseases and traits [ ] . to understand better the association of biological measurements, lifestyle indicators, biomarkers, and health and medical records with covid- susceptibility, we performed analyses to distinguish genetic correlation (e.g., shared genetic liability) from genetically-informative putative causal effects. these results provide insights into the mechanisms underlying several associations linking covid- susceptibility to human diseases and traits. genome-wide association statistics were accessed from the covid- (table s ) . for two covid- phenotypes with h z-scores > , severe respiratory covid- and hospitalized covid- , we then estimated their genetic correlation (rg) with , phenotypes from the uk biobank (ukb, see http://www.nealelab.is/uk-biobank). ldsc analyses were based on linkage disequilibrium information from the genomes project ( kgp) european reference population. for continuous traits, we restricted our analyses to genome-wide association statistics generated from inverse-rank normalized phenotypes. to distinguish between genetic correlation and causative effects, we applied the latent causal variable (lcv) approach [ ] . under the assumption of a single effect-size distribution in per-trait gwas, lcv tests for the presence of a single latent trait connecting covid- outcomes to ukb phenotypes. lcv was performed in r using the kgp european reference ld panel and genome-wide association statistics for snps with minor allele frequencies > %. variants in the major histocompatibility complex region of the genome were excluded because of its complex ld structure. lcv gĉp estimates were only interpreted for trait pairs where both traits exhibit lcv-calculated h z-scores ≥ . the gĉp estimate ranges from - with values near zero indicating partial causality and values approaching indicating full causality. lcv developers recommend that gĉp> . is evidence of a fully causal relationship between trait pairs [ ]. phenome-wide association studies (phewas) were performed for loci associated with one of the three heritable covid- outcomes (table s ) using the pan-ancestry uk biobank resource (available at https://pan.ukbb.broadinstitute.org/downloads). we analyzed genome-wide association statistics generated from the analysis of , phenotypes in six ancestries: european (n= , ), centra/south asian (n= , ), african (n= . ), east asian (n= , ), middle eastern (n= , ), and admixed american (n= ). pan-ukb traits were analyzed if they had cases in european ancestry or cases in all other ancestries. association statistics were covaried with sex, age, age , sex´age, sex´age , and the first ten within-ancestry principle components. a detailed description of the methods used to generate these data is available at https://pan.ukbb.broadinstitute.org/. multiple testing correction was performed for the number of phenotypes and ancestry groups using the p.adjust(method= 'fdr') function of r. assuming a population prevalence of %, the h of severe respiratory covid- (a h = . , se= . , p= . x - ) and hospitalized covid- (b h = . , se= . , p= . x - ) were significantly different from zero. the h for covid- versus population was significantly different from zero (c h = . , se= . , p= . ) but not accurate enough for genetic correlation. severe respiratory covid- and hospitalized covid- were genetically correlated with and phenotypes, respectively, after multiple testing correction ( figure a ). the most significant genetic correlate of each phenotype was waist circumference (severe respiratory covid- rg= . , se= . , p= . x - and hospitalized covid- rg= . , se= . , p= . x - ). five phenotypes were significantly genetically correlated with severe respiratory covid- after multiple testing correction and not correlated (unadjusted p-value > . ) with hospitalized covid- (table s ) the strongest of these was the genetic correlation between ukb field id _ "lack of maternal history of heart disease, stroke, high blood pressure, chronic bronchitis/emphysema, alzheimer's disease/dementia, diabetes" and severe respiratory covid- (rg=- . , se= . , p= . ; versus hospitalized covid- rg=- . , se= . , p= . ). with traits genetically correlated with either covid- outcome after multiple testing correction (table s ) , we tested for causality among ukb, severe respiratory covid- , hospitalized covid- . after multiple testing correction we detected and latent causal genetic relationships with severe respiratory covid- and hospitalized covid- , respectively (table s ) . severe respiratory covid- showed a partial casual effect for manifestations of mania or irritability (gĉp= . , se= . , p= . . x - ) and candesartan cilexetil use (gĉp= . , se= . , p= . x - ). hospitalized covid- was fully genetically causal for diabetes (gĉp= . , se= . , p= . x - ) and alcohol drinking status (gĉp= . , se= . , p= . x - ). twelve phenotypes had significant causal estimates with both severe respiratory covid- and hospitalized covid- ( figure b) . these included smoking and drinking behaviors, diabetes, and heart attack. after multiple testing correction there were no significant differences between genetic causality proportions estimated for severe respiratory covid- and hospitalized covid- . phenome-wide assessment of covid- liability loci (across three severity outcomes: severe respiratory, hospitalized covid- , and all covid- ) identified significant (fdr q< . , figure c ) out of , phenotypes across six ancestries (table s ) . after multiple testing correction, alkaline phosphatase was significantly negatively associated with rs (abo locus) in all six ancestries. the abo locus exhibited significant effect size heterogeneity ( figure s ) with respect to low-density lipoprotein cholesterol concentration (cross-ancestry meta-analysis β= . , p= . x - ) and hemoglobin concentration (crossancestry meta-analysis β= . , p= . x - ). outside the abo locus, we detected significant effect size heterogeneity at rs (cchcr locus, lymphocyte count meta-analysis β= . , p= . x - , phet= . x - ) and rs (keap locus, heel bone mineral density t-score meta-analysis β=- . , p= . x - , phet< . x - ). to date, millions of people worldwide have been infected with sars-cov- , which is the causative agent responsible for global lockdowns and heavily restricted interpersonal contact after widespread covid- outbreak. in light of the covid- pandemic, host genetic susceptibility to severe responses to sars-cov- infection is critical. until recently, genetic epidemiologic approaches to understand host susceptibility have been limited due to relatively small sample sizes and reduced statistical power. in this investigation we use genome-wide all rights reserved. no reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this this version posted november , . ; data to uncover overlap and putative causal relationships between genetic liability to covid- severity, preclinical risk factors (e.g., alcohol consumption) [ ], and long-term consequences of infection (e.g., diabetes) [ ] . these data reflect potential measures to refine, and/or improve accuracy and generalizability of, covid- severity outcomes with epidemiological and selfreport information [ ] . our most notable findings reflect ( ) causal consequences of cigarette smoke exposure on covid- , ( ) causal consequences of covid- on diabetes, and ( ) abo blood type effects on covid- severity across ancestry [ ] . alcohol and diabetes relationships with covid- severity demonstrate that epidemiologic relationships between them are due to putative causal effects [ ] . persons with diabetes have been identified as some of the most high risk individuals and there are several instances of spontaneous diabetes onset following covid- recovery [ ]. to our knowledge, this is the first indication that genetic liability to covid- severity also contributes to diabetes at levels suggesting a "fully causal" relationship. with single-snp measures we recapitulate the relationship between covid- severity and diabetes outcomes by detecting consistent negative association between rs (abo locus) and alkaline phosphatase, an enzyme with evidence of protective effects against diabetes when present in sufficient concentrations [ ] . given the extremely large number of people infected with sars-cov- globally, these risk factors and potential chronic outcomes have critical public health consequences on the long-term economic burden of the covid- pandemic [ ] . all rights reserved. no reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this this version posted november , . ; https://doi.org/ . / . . . doi: medrxiv preprint supplementary information table s . heritability (h ) estimates for seven covid- outcomes in the covid- host genetics initiative (https://www.covid hg.org/; september , release date). table s . genome-wide significant snps associated with covid- outcomes. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this this version posted november , . ; https://doi.org/ . / . . . doi: medrxiv preprint indicate significance after multiple testing correction (fdr %) while single asterisks indicate nominal significance (p< . ). all rights reserved. no reuse allowed without permission. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this this version posted november , . ; https://doi.org/ . / . . . doi: medrxiv preprint prediction for progression risk in patients with covid- pneumonia: the call score the major genetic risk factor for severe covid- is inherited from key: cord- -hnf vayd authors: ford, richard b.; mazzaferro, elisa m. title: emergency care date: - - journal: kirk and bistner's handbook of veterinary procedures and emergency treatment doi: . /b - - - / - sha: doc_id: cord_uid: hnf vayd nan in the event that you suspect peritonitis and have a negative tap with abdominal paracentesis, a diagnostic peritoneal lavage can be performed. to perform abdominal paracentesis, follow this procedure: . place the patient in left lateral recumbency and clip a -to -inch square with the umbilicus in the center. . aseptically scrub the clipped area with antimicrobial scrub solution. . wearing gloves, insert a -or -gauge needle or over-the-needle catheter in four quadrants: cranial and to the right, cranial and to the left, caudal and to the right, and caudal and to the left of the umbilicus. as you insert the needle or catheter, gently twist the needle to push any abdominal organs away from the tip of the needle. local anesthesia typically is not required for this procedure, although a light sedative or analgesic may be necessary if severe abdominal pain is present. in some cases, fluid will flow freely from one or more of the needles. if not, gently aspirate with a -to -ml syringe or aspirate with the patient in a standing position. avoid changing positions with needles in place because iatrogenic puncture of intraabdominal organs may occur. . save any fluid collected in sterile red-and lavender-topped tubes for cytologic and biochemical analyses and bacterial culture. monitor hemorrhagic fluid carefully for the presence of clots. normally, hemorrhagic effusions rapidly become defibrinated and do not clot. clot formation can occur in the presence of ongoing active hemorrhage or may be due to the iatrogenic puncture of organs such as the spleen or liver. if abdominal paracentesis is negative, a diagnostic peritoneal lavage can be performed. peritoneal dialysis kits are commercially available but are fairly expensive and often impractical. to perform a diagnostic peritoneal lavage, follow this procedure: . clip and aseptically scrub the ventral abdomen as described previously. . wearing sterile gloves, cut multiple side ports in a -or -gauge over-the needle catheter. use care to not cut more than % of the circumference of the catheter, or else the catheter will become weakened and potentially can break off in the patient's abdomen. . insert the catheter into the peritoneal cavity caudal and to the right of the umbilicus, directing the catheter dorsally and caudally. . infuse to ml of sterile lactated ringer's solution or . % saline solution that has been warmed to the patient's body temperature. during the instillation of fluid into the peritoneal cavity, watch closely for signs of respiratory distress because an increase in intraabdominal pressure can impair diaphragmatic excursions and respiratory function. . remove the catheter. . in ambulatory patients, walk the patient around while massaging the abdomen to distribute the fluid throughout the abdominal cavity. in nonambulatory patients, gently roll the patient from side to side. . next, aseptically scrub the patient's ventral abdomen again, and perform an abdominal paracentesis as described previously. save collected fluid for culture and cytologic analyses; however, biochemical analyses may be artifactually decreased because of dilution. remember that you likely will retrieve only a small portion of the fluid that you instilled. during the early stage of repair, granulation tissue, some exudate, and minor epithelialization is observed. place a nonadherent bandage with some antibacterial properties (petroleum or nitrofurazone-impregnated gauze) or absorbent material (foam sponge, hydrogel, or hydrocolloid dressing) in direct contact with the wound to minimize disruption of the granulation tissue bed. next, place an absorbent intermediate layer, followed by a porous outer layer, as previously described. granulation tissue can grow through gauze mesh or adhere to foam sponges and can be ripped away at the time of bandage removal. hemorrhage and disruption of the granulation tissue bed can occur. later in the repair process, granulation tissue can exude sanguineous drainage and have some epithelialization. a late nonadherent bandage is required. the contact layer should be some form of nonadherent dressing, foam sponge, hydrogel, or hydrocolloid substance. the intermediate layer and outer layers should be absorbent material and porous tape, respectively. with nonadherent dressings, wounds with viscous exudates may not be absorbed well. this may be advantageous and enhance epithelialization, provided that complications do not occur. infection, exuberant granulation tissue, or adherence of absorbent materials to the wound may occur and delay the healing process. moist healing is a newer concept of wound management in which wound exudates are allowed to stay in contact with the wound. in the absence of infection a moist wound heals faster and has enzymatic activity as a result of macrophage and polymorphonuclear cell breakdown. enzymatic degradation or "autolytic debridement" of the wound occurs. moist wounds tend to promote neutrophil and macrophage chemotaxis and bacterial phagocytosis better than use of wet-to-dry bandages. a potential complication and disadvantage of moist healing, however, is the development of bacterial colonization, folliculitis, and trauma to wound edges that can occur because of the continuously moist environment. use surfactant-type solutions (constant clens; kendall, mansfield, massachusetts) for initial wound cleansing and debridement. use occlusive dressings for rapid enzymatic debridement with bactericidal properties to aid in wound healing. bandage wet necrotic wounds with a dressing premoistened with hypertonic saline (curasalt [kendall] , % saline) to clean and debride the wounds. hypertonic saline functions to desiccate necrotic tissue and bacteria to debride the infected wound. remove and replace the hypertonic saline bandage every to hours. next, place gauze impregnated with antibacterial agents (kerlix amd [kendall] ) over the wound in the bandage layer to act as a barrier to bacterial colonization. if the wound is initially dry or has minimal exudate and is not obviously contaminated or infected, place amorphous gels of water, glycerin, and a polymer (curafil [kendall] ) over the wound to promote moisture and proteolytic healing. discontinue moisture gels such as curafil once the dry wound has become moist. finally, the final stage of moist healing helps to promote the development of a healthy granulation tissue bed. use calcium alginate dressings (curasorb or curasorb zn with zinc [kendall] ) in noninfected wounds with a moderate amount of drainage. alginate gels promote rapid development of a granulation tissue bed and epithelialization. foam dressings also can be applied to exudative wounds after a healthy granulation bed has formed. change foam dressings at least once every to days. for closed wounds without any drainage, such as a laceration that has been repaired surgically, a simple bandage with a nonadherent contact layer (telfa pad [kendall] , for example), intermediate layer of absorbent material, and an outer porous layer (elastikon, vetrap) can be placed to prevent wound contamination during healing. the nonadherent pad will not stick to the wound and cause patient discomfort. because there usually is minimal drainage from the wound, the function of the intermediate layer is more protective than absorptive. any small amount will be absorbed into the intermediate layer of the bandage. it is important in any bandage to place the tape strips or "stirrups" on the patient's limb and then overlap in the bandage, to prevent the bandage from slipping. place the intermediate and tertiary layers loosely around the limb, starting distally and working proximally, with some overlap with each consecutive layer. this method prevents excessive pressure and potential to impair venous drainage. leave the toenails of the third and fourth digits exposed, whenever possible, to allow daily examination of the bandage to determine whether the bandage is impairing venous drainage. if the bandage is too tight and constricting or impeding vascular flow, the toes will become swollen and spread apart. when placed and maintained properly (e.g., the bandage does not get wet), there usually are relatively few complications observed with this type of bandage. in some cases, it is necessary to cover a wound in which a penrose drain has been placed to allow drainage. in many cases, there is a considerable amount of drainage from the drain and underlying soft tissues. the function of the bandage is to help obliterate dead space created by the wound itself, absorb the fluid that drains from the wound and that will contaminate the environment, and prevent external wicking of material from the external environment into the wound. when the bandage is removed, the clinician can examine the amount and type of material that has drained from the wound in order to determine when the drain should be removed. when placing a bandage over a draining wound, the contact layer should be a commercially available nonadherent dressing and several layers of absorbent wide-mesh gauze placed directly over the drain at the distal end of the incision. overlay the layers of gauze with a thick layer of absorbent intermediate dressing to absorb fluid that drains from the wound. if the gauze and intermediate layers are not thick or absorbent enough, there is a potential for the drainage fluid to reach the outer layer of the bandage and provide a source of wicking of bacteria from the external environment into the wound, leading to infection. some wounds such as lacerations have minor bleeding or hemorrhage that require an immediate bandage until definitive care can be provided. to create a pressure bandage, place a nonadherent dressing immediately in contact with the wound, followed by a thick layer of absorbent material, topped by a layer of elastic bandage material such as elastikon or vetrap. unlike the bandage for a closed wound, the top tertiary outer layer should be wrapped with some tension and even pressure around the limb, starting from the distal extremity (toes) and working proximally. the pressure bandage serves to control hemorrhage but should not be left on for long periods. pressure bandages that have been left on for too long can impair nerve function and lead to tissue necrosis and slough. therefore, pressure bandages should be used in the hospital only, so that the patient can be observed closely. if hemorrhage through the bandage occurs, place another bandage over the first until the wound can be repaired definitively. removal of the first bandage will only disrupt any clot that has formed and cause additional hemorrhage to occur. fractures require immediate immobilization to prevent additional patient discomfort and further trauma to the soft tissues of the affected limb. as with all bandages, a contact layer, intermediate layer, and outer layer should be used. place the contact layer in accordance with any type of wound present. the intermediate layer should be thick absorbent material, followed by a top layer of elastic bandage material. an example is to place a telfa pad over a wound in an open distal radius-ulna fracture, followed by a thick layer of cotton gauze cast padding, followed by an elastic layer of kling (johnson & johnson medical, arlington, texas) , pulling each layer tightly over the previous layer with some overlap until the resultant bandage can be "thumped" with the clinician's thumb and forefinger and sound like a ripe watermelon. the bandage should be smooth with consecutive layers of even pressure on the limb, starting distally and working proximally. leave the toenails of the third and fourth digits exposed to monitor for impaired venous drainage that would suggest that the bandage is too tight and needs to be replaced. finally, place a top layer of vetrap or elastikon over the intermediary layer to protect it from becoming contaminated. if the bandage is used with a compound or open fracture, drainage may be impaired and actually lead to enhanced risk of wound infection. bandages placed for initial fracture immobilization are temporary until definitive fracture repair can be performed once the patient's cardiovascular and respiratory status are stable. wounds with exuberant granulation tissue must be handled carefully so as to not disrupt the healing process but to keep an overabundance of tissue from forming that will impair epithelialization. to bandage a wound with exuberant granulation tissue, place a corticosteroid-containing ointment on the wound, followed by a nonadherent contact layer. the corticosteroid will help control the exuberant growth of granulation tissue. next, carefully wrap an absorbent material over the contact layer, followed by careful placement of and overlay of elastic bandage material to place some pressure on the wound. leave the toenails of the third and fourth digits exposed so that circulation can be monitored several times daily. bandages that are too tight must be removed immediately to prevent damage to neuronal tissue and impaired vascularization, tissue necrosis, and slough. because wound drainage may be impaired, there is a risk of infection. gaping wounds or those that have undermined in between layers of subcutaneous tissue and fascia should be bandaged with a pressure bandage to help obliterate dead space and prevent seroma formation. an example of a wound that may require this type of bandage is removal of an infiltrative lipoma on the lateral or ventral thorax. use caution when placing pressure bandages around the thorax or cervical region because bandages placed too tightly may impair adequate ventilation. to place a pressure bandage and obliterate dead space, place a nonadherent contact layer over the wound. usually, a drain is placed in the wound, so place a large amount of wide-mesh gauze at the distal end of the drain to absorb any wound exudate or drainage. place several layers of absorbent material over the site to further absorb any drainage. place a layer of elastic cotton such as kling carefully but firmly over the dead space to cause enough pressure to control drainage. place at least two fingers in between the animal's thorax and the bandage to ensure that the bandage is not too tight. in many cases, the bandage should be placed once the animal has recovered from surgery and is able to stand. if the bandage is placed while the animal is still anesthetized and recumbent, there is a tendency for the bandage to be too tight. finally, the tertiary layer should be an elastic material such as elastikon or vetrap. many wounds require a pressure relief bandage to prevent contact with the external environment. wounds that may require pressure relief for healing include decubitus ulcers, pressure bandage or cast ulcers, impending ulcer areas (such as the ileum or ischium of recumbent or cachexic patients), and surgical repair sites of ulcerated areas. pressure relief bandages can be of two basic varieties: modified doughnut bandage and doughnut-shaped bandage. to create a cup or clamshell splint, follow this procedure (figures - to - ): . place a nonadherent contact layer directly over the wound. . place stirrups of tape in contact with the skin of the dog, to be placed over the intermediate layer and prevent the bandage from slipping. . place a fairly thick layer of absorbent intermediate bandage material over the contact layer such that the bandage is well-padded. pull the tape stirrups and secure them to the intermediate layer. . place a length of cast material that has been rolled to the appropriate length, such that the cast material is cupped around the patient's paw, and lies adjacent to the caudal aspect of the limb to the level of the carpus or tarsus. in the case of a clamshell splint, place a layer of cast material on the cranial and caudal aspect of the paw and conform it in place. . take the length of cast padding and soak it in warm water after it has been rolled to the appropriate length. wring out the pad, and secure/conform it to the caudal (or cranial and caudal, in the case of a clamshell splint) aspect of the distal limb and paw. . secure the cast material in place with a layer of elastic cotton gauze (kling). . secure the bandage in place with a snug layer of elastikon or vetrap. short or long splints made of cast material can be incorporated into a soft padded bandage to provide extra support of a limb above and below a fracture site. for a caudal or lateral splint to be effective, it must be incorporated for at least one joint above any fracture site to prevent a fulcrum effect and further disruption or damage to underlying soft tissue structures. a short lateral or caudal splint is used for fractures and luxations of the distal metacarpus, metatarsus, carpus, and tarsus. to place a short lateral or caudal splint, follow this procedure: . secure a contact layer as determined by the presence or absence of any wound in the area. . place tape stirrups on the distal extremity to be secured later to the intermediate bandage layer and to prevent slipping of the bandage distally. . place layers of roll cotton from the toes to the level of the mid tibia/fibula or mid radius/ulna. place the layers with even tension, with some overlap of each consecutive layer, moving distally to proximally on the limb. . secure the short caudal or lateral splint and conform it to the distal extremity to the level of the toes and proximally to the level of the mid tibia/fibula or mid radius/ulna. . secure the lateral or caudal splint to the limb with another outer layer of elastic cotton (kling). . cover the entire bandage and splint with an outer tertiary layer of vetrap or elastikon. make sure that the toenails of the third and fourth digits remain visible to allow daily evaluation of circulation. long lateral or caudal splints are used to immobilize fractures of the tibia/fibula and radius/ulna. the splints are fashioned as directed for short splints but extend proximally to the level of the axilla and inguinal regions to immobilize above the fracture site. • packed cell volume drops rapidly to less than % in the dog and less than % to % in the cat • acute loss of more than % of blood volume ( ml/kg in dog, ml/kg in cat) • clinical signs of lethargy, collapse, hypotension, tachycardia, tachypnea (acute or chronic blood loss) • ongoing hemorrhage is present • poor response to crystalloid and colloid infusion • life-threatening hemorrhage caused by thrombocytopenia or thrombocytopathia • surgical intervention is necessary in a patient with severe thrombocytopenia or thrombocytopathia plasma support • life-threatening hemorrhage with decreased coagulation factor activity • severe inflammation (pancreatitis, systemic inflammatory response syndrome) • replenish antithrombin (disseminated intravascular coagulation, protein-losing enteropathy or nephropathy) • surgery is necessary in a patient with decreased coagulation factor activity • severe hypoproteinemia is present; to partially replenish albumin, globulin, and clotting factors type a cats typically possess weak anti-b antibodies of igg and igm subtypes. transfusion of type b blood into a type a cat will result in milder clinical signs of reaction and a markedly decreased survival half-life of the infused rbcs to just days. because type ab cats possess both moieties on their cell surface, they lack naturally occurring alloantibodies; transfusion of type a blood into a type ab cat can be performed safely if a type ab donor is not available. the life span of an rbc from a type-specific transfusion into a cat is approximately days. . indications for fresh whole blood transfusion include disorders of hemostasis and coagulopathies including disseminated intravascular coagulation, von willebrand's disease, and hemophilia. fresh whole blood and platelet-rich plasma also can be administered in cases of severe thrombocytopenia and thrombocytopathia. stored whole blood and packed rbcs can be administered in patients with anemia. if pcv drops to below % or if rapid hemorrhage causes the pcv to drop below % in the dog or less than % to *indicates that this must be done for each donor being tested. minor crossmatch* . obtain a crossmatch segment from blood bank refrigerator for each donor to be crossmatched, or use an edta tube of donor's blood. make sure tubes are labeled prop-erly. . collect ml of blood from recipient and place in an edta tube. centrifuge blood for minutes. . extract blood from donor tubing. centrifuge blood for minutes. use a separate pipette for each transfer because cross-contamination can occur. . pipette plasma off of donor and recipient cells and place in tubes labeled dp and rp, respectively. . place µl of donor and recipient cells in tubes labeled dr and rr, respectively. . add . ml . % sodium chloride solution from wash bottle to each red blood cell (rbc) tube, using some force to cause cells to mix. . centrifuge rbc suspension for minutes. . discard supernatant and resuspend rbcs with . % sodium chloride from wash bottle. . repeat steps and for a total of three washes. . place drops of donor rbc suspension and drops of recipient plasma in tube labeled ma (this is the major crossmatch). . place drops of donor plasma and drops recipient rbc suspension in tube labeled mi (this is the minor crossmatch). . prepare control tubes by placing drops donor plasma with drops donor rbc suspension (this is the donor control); and place drops recipient plasma with drops recipient rbc suspension (this is the recipient control). . incubate major and minor crossmatches and control tubes at room temperature for minutes. . centrifuge all tubes for minute. . read tubes using an agglutination viewer. . check for agglutination and/or hemolysis. . score agglutination with the following scoring scale: + one solid clump of cells + several large clumps of cells + medium-sized clumps of cells with a clear background + hemolysis, no clumping of cells neg = negative for hemolysis; negative for clumping of red blood cells fresh whole blood coagulopathy with active hemorrhage (disseminated intravascular coagulation, thrombocytopenia; massive acute hemorrhage; no stored blood available) stored whole blood massive acute or ongoing hemorrhage; hypovolemic shock caused by hemorrhage that is unresponsive to conventional crystalloid and colloid fluid therapy; unavailability of equipment required to prepare blood components packed red blood cells nonregenerative anemia, immune-mediated hemolytic anemia, correction of anemia before surgery, acute or chronic blood loss fresh frozen plasma factor depletion associated with active hemorrhage (congenital: von willebrand's factor, hemophilia a, hemophilia b; acquired: vitamin k antagonist, rodenticide intoxication, dic); acute or chronic hypoproteinemia (burns, wound exudates, body cavity effusion; hepatic, renal, or gastrointestinal loss); colostrum replacement in neonates frozen plasma acute plasma or protein loss; chronic hypoproteinemia; (contains stable colostrum replacement in neonates; hemophilia b and clotting factors) selected clotting factor deficiencies platelet-rich plasma* thrombocytopenia with active hemorrhage (immune-mediated thrombocytopenia, dic); platelet function abnormality (congenital: thrombasthenia in bassett hounds; acquired: nsaids, other drugs) cryoprecipitate congenital factor deficiencies (routine or before surgery): (concentration of factor hemophilia a, hemophilia b, von willebrand's disease, viii, von willebrand's hypofibrinogenemia; acquired factor deficiencies factor, and fibrinogen) *must be purchased because logistically one cannot obtain enough blood simultaneously to provide a significant amount of platelets; platelets infused have a very short (< hours) half-life. dic, disseminated intravascular coagulation; nsaids, nonsteroidal antiinflammatory drugs. universal donor (e.g., should be administered whenever possible. because there is no universal donor in the cat and because cats possess naturally occurring alloantibodies, all cat blood should be typed and crossmatched before any transfusion. if fresh whole blood is not available, a hemoglobin-based oxygen carrier (oxyglobin, to ml/kg iv) can be administered until blood products become available. table - indicates blood component dose and administration rates. blood products should be warmed slowly to °c before administering them to the patient. blood warmer units are available for use in veterinary medicine to facilitate rapid transfusion without decreasing patient body temperature (thermal angel; enstill medical technologies, inc., dallas, texas). red blood cell and plasma products should be administered in a blood administration set containing a -µm in-line filter. smaller in-line filters ( µm) also can be used in cases in which extremely small volumes are to be administered. blood products should be administered over a period of hours, whenever possible, according to guidelines set by the american association of blood banks. the volume of blood components required to achieve a specific increment in the patient's pcv depends largely on whether whole blood or packed rbcs are transfused and whole blood ml/kg will increase max rate: ml/kg/ max: ml/kg/ volume by % hours hour packed red ml/kg will increase critically ill blood cells volume by % patients (e.g., cardiac failure or renal failure): - ml/kg/hour fresh frozen ml/kg body mass (repeat - ml/minute or use rates as for plasma in - days or in - days whole blood (infuse within - hours) or until bleeding stops); monitor act, aptt, and pt before and hour after transfusion cryoprecipitate general: unit/ kg/ hours - ml/minute or use rates as for whole or until bleeding stops blood (infuse within - hours) hemophilia a: - units factor viii/kg; unit of cryoprecipitate contains approximately units of factor viii platelet-rich unit/ kg ( unit of ml/minute plasma platelet-rich plasma will check platelet count before and hour increase platelet count after transfusion hour after transfusion by , /µl) whether there is ongoing hemorrhage or rbc destruction. because the pcv of packed rbcs is unusually high ( % for greyhound blood), a smaller total volume is required than whole blood to achieve a comparable increase in the patient's pcv. in general, ml/kg of packed rbcs or ml/kg whole blood will raise the recipient's pcv by %. the "rule of ones" states that ml per lb of whole blood will raise the pcv by %. if the patient's pcv does not raise by the amount anticipated by the foregoing calculation(s), causes of ongoing hemorrhage or destruction should be considered. the goal of red blood component therapy is to raise the pcv to % to % in dogs and % to % in cats. if an animal is hypovolemic and whole blood is administered, the fluid is redistributed into the extravascular compartment within hours of transfusion. this will result in a secondary rise in the pcv hours after the transfusion in addition to the initial rise to hours after the rbc transfusion is complete. the volume of plasma transfused depends largely on the patient's need. in general, plasma transfusion should not exceed more than ml/kg during a -hour period for normovolemic animals. thaw plasma at room temperature, or place it in a ziplock freezer bag and run under cool (not warm) water until thawed. then administer the plasma through a blood administration set that contains an in-line blood filter or through a standard driptype administration set with a detachable in-line blood administration filter. the average rate of plasma infusion in a normovolemic patient should not exceed ml/kg/hour. in acute need situations, plasma can be delivered at rates up to to ml/kg/minute. for patients with cardiac insufficiency or other circulatory problems, plasma infusion rates should not exceed ml/kg/hour. plasma or other blood products should not be mixed with or used in the same infusion line as calcium-containing fluids, including lactated ringer's solution, calcium chloride, or calcium gluconate. the safest fluid to mix with any blood product is . % sodium chloride. administer fresh frozen plasma, frozen plasma, and cryoprecipitate at a volume of ml/kg until bleeding is controlled or source of ongoing albumin loss ceases. the goal of plasma transfusion therapy is to raise the albumin to a minimum of . g/dl or until bleeding stops as in the case of coagulopathies. monitor the patient to ensure that bleeding has stopped, coagulation profiles (act, aptt, and pt) have normalized, hypovolemia has stabilized, and/or total protein is normalizing, which are indications for discontinuing ongoing transfusion therapy. plasma cryoprecipitate can be purchased or manufactured through the partial thawing and then centrifugation of fresh frozen plasma. cryoprecipitate contains concentrated quantities of vwf, factor viii, and fibrinogen and is indicated in severe forms of von willebrand's disease and hemophilia a (factor viii deficiency). platelet-rich plasma must be purchased from a commercial source. one unit of fresh whole blood contains to platelets. the viability of the platelets contained in the fresh whole blood is short-lived, just to hours after transfusion into the recipient. because platelet-rich plasma is difficult to obtain, animals with severe thrombocytopenia or thrombocytopathia should be treated with immunomodulating therapies and the administration of fresh frozen plasma. in dogs, blood and plasma transfusions can be administered intravenously or intraosseously. the cephalic, lateral saphenous, medial saphenous, and jugular veins are used most commonly. fill the recipient set so that the blood in the drip chamber covers the filter (normal -µm filter). with small amounts of blood ( ml) or critically ill patients, use a -µm filter. avoid latex filters for plasma and cryoprecipitate administration. blood can emergency care be administered at variable rates, but the routine figure of to ml/minute often is used. normovolemic animals can receive blood at ml/kg/day. dogs in heart failure should receive infusions at no more than ml/kg/hour. volume is given as needed. to calculate the approximate volume of blood needed to raise hematocrit levels, use the following formula for the dog: anticoagulated blood volume (ml) = body mass (kg) × × pcv desired − pcv of recipient pcv of donor in anticoagulant an alternative formula is the following: . × recipient body mass (kg) × (dog) × pcv desired − pcv of recipient pcv of donor in anticoagulant surgical emergencies and shock may require several times this volume within a short period. if greater than % of the patient's blood volume is lost, supplementation with colloids, crystalloids, and blood products is indicated for fluid replacement. one volume of whole blood achieves the same increase in plasma as two to three volumes of plasma. if the patient's blood type is unknown and type a-negative whole blood is not available, any dog blood can be administered to a dog in acute need if the dog has never had a transfusion before. if mismatched blood is given, the patient will become sensitized, and after days, destruction of the donor rbcs will begin. in addition, any subsequent mismatched transfusions may cause an immediate reaction (usually mild) and rapid destruction of the transfused rbcs. the clinical signs of a transfusion reaction typically only are seen when type a blood is administered to a type a-negative recipient that has been sensitized previously. incompatible blood transfusions to breeding females can result in isoimmunization and in hemolytic disease in the puppies. the a-negative bitch that receives a transfusion with a-positive and that produces a litter from an a-positive stud can have puppies with neonatal isoerythrolysis. cats with severe anemia in need of a blood transfusion are typically extremely depressed, lethargic, and anorexic. the stress of restraint and handling can push these critically ill patients over the edge and cause them to die. extreme gentleness and care are mandatory in restraint and handling. the critically ill cat should be cradled in a towel or blanket. supplemental flow-by or mask oxygen should be administered, whenever possible, although it may not be clinically helpful until oxygen-carrying capacity is replenished with infusion of rbcs or hemoglobin. blood can be administered by way of cephalic, medial saphenous, or the jugular vein. intramedullary infusion is also possible, if vascular access cannot be accomplished. the average -to -kg cat can accept to ml of whole blood injected intravenously over a period of to minutes. administer filtered blood at a rate of to ml/kg/hour. the following formula can be used to estimate the volume of blood required for transfusion in a cat: anticoagulated blood volume (ml) = body mass (kg) × × pcv desired − pcv of recipient pcv of donor in anticoagulant the exact overall incidence and clinical significance of transfusion reactions in veterinary medicine are unknown. several studies have been performed that document the incidence of transfusion reactions in dogs and cats. overall, the incidence of transfusion reactions in dogs and cats is . % and %, respectively. transfusion reactions can be immune-mediated and non-immune-mediated and can happen immediately or can be delayed until after a transfusion. acute reactions usually occur within minutes to hours of the onset of transfusion but may occur up to hours after the transfusion has been stopped. acute immunologic reactions include hemolysis and acute hypersensitivity including rbcs, platelets, and leukocytes. signs of a delayed immunologic reaction include hemolysis, purpura, immunosuppression, and neonatal isoerythrolysis. acute nonimmunologic reactions include donor cell hemolysis before onset of transfusion, circulatory volume overload, bacterial contamination, citrate toxicity with clinical signs of hypocalcemia, coagulopathies, hyperammonemia, hypothermia, air embolism, acidosis, and pulmonary microembolism. delayed nonimmunologic reactions include the transmission and development of infectious diseases and hemosiderosis. clinical signs of a transfusion reaction typically depend on the amount of blood transfused, the type and amount of antibody involved in the reaction, and whether the recipient has had previous sensitization. monitoring the patient carefully during the transfusion period is essential in recognizing early signs of a transfusion reaction, including those that may become life threatening. a general guideline for patient monitoring is first to start the transfusion slowly during the first minutes. monitor temperature, pulse, and respiration every minutes for the first hour, hour after the end of the transfusion, and every hours minimally thereafter. also obtain a pcv immediately before the transfusion, hour after the transfusion has been stopped, and every hours thereafter. monitor coagulation parameters such as an act and platelet count at least daily in patients requiring transfusion therapy. the most common documented clinical signs of a transfusion reaction include pyrexia, urticaria, salivation/ptyalism, nausea, chills, and vomiting. other clinical signs of a transfusion reaction may include tachycardia, tremors, collapse, dyspnea, weakness, hypotension, collapse, and seizures. severe intravascular hemolytic reactions may occur within minutes of the start of the transfusion, causing hemoglobinemia, hemoglobinuria, disseminated intravascular coagulation, and clinical signs of shock. extravascular hemolytic reactions typically occur later and will result in hyperbilirubinemia and bilirubinuria. pretreatment of patients to help decrease the risk of a transfusion reaction remains controversial, and in most cases, pretreatment with glucocorticoids and antihistamines is ineffective at preventing intravascular hemolysis and other reactions should they occur. the most important component of preventing a transfusion reaction is to screen each recipient carefully and process the donor component therapy carefully before the administration of any blood products. treatment of a transfusion reaction depends on its severity. in all cases, stop the transfusion immediately when clinical signs of a reaction occur. in most cases, discontinuation of the transfusion and administration of drugs to stop the hypersensitivity reaction will be sufficient. once the medications have taken effect, restart the transfusion slowly and monitor the patient carefully for further signs of reaction. in more severe cases in which a patient's cardiovascular or respiratory system become compromised and hypotension, tachycardia, or tachypnea occurs, immediately discontinue the transfusion and administer diphenhydramine ( mg/kg im), dexamethasone-sodium phosphate ( . to . mg/kg iv), and epinephrine to the patient. the patient should have a urinary catheter and central venous catheter placed for measurement of urine output and central venous pressures. aggressive fluid therapy may be necessary to avoid renal insufficiency or renal damage associated with severe intravascular hemolysis. overhydration with subsequent pulmonary edema generally can be managed with supplemental oxygen administration and intravenous or intramuscular administration of furosemide ( to mg/kg). plasma products with or without heparin can be administered for disseminated intravascular coagulation. the hbocs can be stored at room temperature and have a relatively long shelf life compared with red blood component products. the hbocs function to carry oxygen through the blood and can diffuse oxygen past areas of poor tissue perfusion. an additional characteristic of hbocs is as a potent colloid, serving to maintain fluid within the vascular space. for this reason, hbocs must be used with caution in euvolemic patients and patients with cardiovascular insufficiency. central venous pressure (cvp) measures the hydrostatic pressure in the anterior vena cava and is influenced by vascular fluid volume, vascular tone, function of the right side of the heart, and changes in intrathoracic pressure during the respiratory cycle. the cvp is not a true measure of blood volume but is used to gauge fluid therapy as a method of determining how effectively the heart can pump the fluid that is being delivered to it. thus the cvp reflects the interaction of the vascular fluid volume, vascular tone, and cardiac function. measure cvp in any patient with acute circulatory failure, large volume fluid diuresis (i.e., toxin or oliguric or anuric renal failure), fluid in-and-out monitoring, and cardiac dysfunction. the placement of central venous catheters and thus cvp measurements is contraindicated in patients with known coagulopathies including hypercoagulable states. to perform cvp monitoring, place a central venous catheter in the right or left jugular vein. in cats and small dogs, however, a long catheter placed in the lateral or medial saphenous vein can be used for trends in cvp monitoring. first, assemble the equipment necessary for jugular catheter (see vascular access techniques for how to place a jugular or saphenous long catheter) and cvp monitoring (box - ). after placing the jugular catheter, take a lateral thoracic radiograph to ensure that the tip of the catheter sits just outside of the right atrium for proper cvp measurements (see to establish an intravenous catheter for cvp, follow this procedure: . assemble the cvp setup such that the male end of a length of sterile intravenous catheter extension tubing is inserted into the t port of the jugular or medial/lateral saphenous catheter. make sure to flush the length of tubing with sterile saline before connecting it to the patient to avoid iatrogenic air embolism. . next, insert the male end of a three-way stopcock into the female end of the extension tubing. . attach a -ml syringe filled with heparinized sterile . % saline to one of the female ports of the three-way stopcock and either a manometer or a second length of intravenous extension tubing attached to a metric ruler. . lay the patient in lateral or sternal recumbancy. . turn the stopcock off to the manometer/ruler and on to the patient. infuse a small amount of heparinized saline through the catheter to flush the catheter. . next, turn the stopcock off to the patient and on to the manometer. gently flush the manometer or length of extension tubing with heparinized saline from the syringe. use care not to agitate the fluid and create air bubbles within the line or manometer that will artifactually change the cvp measured. . next, lower the cm point on the manometer or ruler to the level of the patient's manubrium (if the patient is in lateral recumbancy) or the point of the elbow (if the patient is in sternal recumbancy). . turn the stopcock off to the syringe, and allow the fluid column to equilibrate with the patient's intravascular volume. once the fluid column stops falling and the level rises and falls with the patient's heartbeat, measure the number adjacent to the bottom of the meniscus of the fluid column. this is the cvp in centimeters of water (see figure - ). . repeat the measurement several times with the patient in the same position to make sure that none of the values has been increased or decreased artifactually in error. alternately, attach the central catheter to a pressure transducer and perform electronic monitoring of cvp. there is no absolute value for normal cvp. the normal cvp for small animal patients is to cm h o. values less than zero are associated with absolute or relative hypovolemia. values of to cm h o are borderline hypervolemia, and values greater than cm h o suggest intravascular volume overload. values greater than cm h o may be correlated with congestive heart failure and the development of pulmonary edema. in individual patients, the trend in change in cvp is more important than absolute values. as a rule of thumb, when using cvp measurements to gauge fluid therapy and avoid vascular and pulmonary overload, the cvp should not increase by more than cm h o in any -hour period. if an abrupt increase in cvp is found, repeat the measurement to make sure that the elevated value was not obtained in error. if the value truly has increased dramatically, temporarily discontinue fluid therapy and consider administration of a diuretic. delaforcade am, rozanski ea: central venous pressure and arterial blood pressure measurements, vet clin north am small anim pract ( ) the diagnosis of intracellular fluid deficit is difficult and is based more on the presence of hypernatremia or hyperosmolality than on clinical signs. an intracellular fluid deficit is expected when free water loss by insensible losses and vomiting, diarrhea, or urine is not matched by free water intake. consideration of the location of the patient's fluid deficit, history of vomiting and diarrhea, no visible clinical signs of deficit % dry mucous membranes, mild skin tenting % increased skin tenting, dry mucous membranes, mild tachycardia, normal pulse* % increased skin tenting, dry mucous membranes, tachycardia, weak pulse pressure % increased skin tenting, dry corneas, dry mucous membranes, % elevated or decreased heart rate, poor pulse quality, altered level of consciousness* the respiratory system further contributes to acid-base status by changes in the elimination of carbon dioxide. hyperventilation decreases the blood pco and causes a respiratory alkalosis. hypoventilation increases the blood pco and causes a respiratory acidosis. depending on the altitude, the pco in dogs can range from to mm hg. in cats, normal is to mm hg. venous pco values are to mm hg in dogs and to mm hg in cats. use a systematic approach whenever attempting to interpret a patient's acid-base status. ideally, obtain an arterial blood sample so that you can monitor the patient's oxygenation and ventilation. once an arterial blood sample has been obtained, follow these steps: . determine whether the blood sample is arterial or venous by looking at the oxygen saturation (sao ). the sao should be greater than % if the sample is truly arterial, although it can be as low as % if a patient has severe hypoxemia. . consider the patient's ph. if the ph is outside of the normal range, an acid-base disturbance is present. if the ph is within the normal range, an acid-base disturbance may or may not be present. if the ph is low, the patient is acidotic. if the ph is high, the patient is alkalotic. . next, look at the base excess or deficit. if the base excess is increased, the patient has higher than normal bicarbonate. if there is a base deficit, the patient may have a low bicarbonate or increase in unmeasured anions (e.g., lactic acid or ketoacids). . next, look at the bicarbonate. if the ph is low and the bicarbonate is low, the patient has a metabolic acidosis. if the ph is high and the bicarbonate is elevated, the patient has a metabolic alkalosis. . next, look at the paco . if the patient's ph is low and the paco is elevated, the patient has a respiratory acidosis. if the patient's ph is high and the paco is low, the patient has a respiratory alkalosis. . finally, if you are interested in the patient's oxygenation, look at the pao . normal pao is greater than mm hg. the metabolic acidosis early in renal failure may be hyperchloremic and later may convert to typical increased anion gap acidosis. . next, you must determine whether the disorders present are primary disorders or an expected compensation for disorders in the opposing system. for example, is the patient retaining bicarbonate (metabolic alkalosis) because of carbon dioxide retention (respiratory acidosis)? use the chart in table - to evaluate whether the appropriate degree of compensation is occurring. if the adaptive response falls within the expected range, a simple acid-base disorder is present. if the response falls outside of the expected range, a mixed acid-base disorder is likely present. . finally, you must determine whether the patient's acid-base disturbance is compatible with the history and physical examination findings. if the acid-base disturbance does not fit with the patient's history and physical examination abnormalities, question the results of the blood gas analyses and possibly repeat them. the most desirable method of assessing the acid-base status of an animal is with a blood gas analyzer. arterial samples are preferred over venous samples, with heparin used as an anticoagulant (table - ) . potassium primarily is located in the intracellular fluid compartment. serum potassium is regulated by the actions of the sodium-potassium-adenosinetriphosphatase pump on cellular membranes, including those of the renal tubular epithelium. inorganic metabolic acidosis artifactually can raise serum potassium levels because of redistribution of extracellular potassium in exchange for intracellular hydrogen ion movement in an attempt to correct serum ph. metabolic acidosis potassium is one of the major players in the maintenance of resting membrane potentials of excitable tissue, including neurons and cardiac myocytes. changes in serum potassium can affect cardiac conduction adversely. hyperkalemia lowers the resting membrane potential and makes cardiac cells, particularly those of the atria, more susceptible to depolarization. characteristic signs of severe hyperkalemia that can be observed on an ecg rhythm strip include an absence of p waves, widened qrs complexes, and tall tented or spiked t waves. further increases in serum potassium can be associated with bradycardia, ventricular fibrillation, and cardiac asystole (death). treatment of hyperkalemia consists of administration of insulin ( . to . units/kg, iv regular insulin) and dextrose ( g dextrose per unit of insulin administered, followed by . % dextrose iv cri to prevent hypoglycemia), calcium ( to ml of % calcium gluconate administered iv slowly to effect), or sodium bicarbonate ( meq/kg, iv slowly). insulin plus dextrose and bicarbonate therapy help drive the potassium intracellularly, whereas calcium antagonizes the effect of hyperkalemia on the myocardial cells. all of the treatments work within minutes, although the effects are relatively short-lived ( minutes to hour) unless the cause of the hyperkalemia is identified and treated appropriately (box - ). dilution of serum potassium also results from restoring intravascular fluid volume and correcting metabolic acidosis, in most cases. treatment with a fluid that does not contain potassium (preferably . % sodium chloride) is recommended. hypokalemia elevates the resting membrane potential and results in cellular hyperpolarization. hypokalemia may be associated with ventricular dysrhythmias, but the ecg changes are not as characteristic as those observed with hyperkalemia. causes of hypokalemia include renal losses, anorexia, gastrointestinal loss (vomiting, diarrhea), intravenous fluid diuresis, loop diuretics, and postobstructive diuresis (box - ). if the serum potassium concentration is known, potassium supplementation in the form of potassium chloride or potassium phosphate can be added to the patient's intravenous fluids. correct serum potassium levels less than . meq/l or greater than . meq/l. potassium rates should not exceed . meq/kg/hour (table - ) . metabolic acidosis from bicarbonate depletion often corrects itself with volume restoration in most small animal patients. patients with moderate to severe metabolic acidosis may benefit from bicarbonate supplementation therapy. the metabolic contribution to acid-base balance is identified by measuring the total carbon dioxide concentration or calculating the bicarbonate concentration. if these measurements are not available, the degree of expected metabolic acidosis can be estimated subjectively by the severity of underlying disease that often contributes to metabolic acidosis: hypovolemic or traumatic shock, septic shock, diabetic ketoacidosis, or oliguric/anuric renal failure. if the metabolic acidosis is estimated to be mild, moderate, or severe, add sodium bicarbonate at , , and meq/kg body mass, respectively. patients with diabetic ketoacidosis may not require bicarbonate administration once volume replacement and perfusion is restored, and the ketoacids are metabolized to bicarbonate. if the bicarbonate measurement of base deficit is known, the following formula can be used as a gauge for bicarbonate supplementation: base deficit × . = body mass (kg) = meq bicarbonate to administer osmolality osmolality is measured by freezing point depression or a vapor pressure osmometer, or it may be calculated by the following formula: mosm/kg = [(na + ) + (k + )] + bun/ . + glucose/ where sodium and potassium are measured in milliequivalents, and bun and glucose are measured in milligrams per deciliter. osmolalities less than mosm/kg or greater emergency care than mosm/kg are serious enough to warrant therapy. the difference between the measured osmolality and the calculated osmolality (the osmolal gap) should be less than mosm/kg. if the osmolal gap is greater than mosm/kg, consider the presence of unmeasured anions such as ethylene glycol metabolites. the volume of extracellular fluid is determined by the total body sodium content, whereas the osmolality and sodium concentration are determined by water balance. serum sodium concentration is an indication of the amount of sodium relative to water in the extracellular fluid and provides no direct information about the total body sodium content. unlikely to cause hyperkalemia in presence of normal renal function unless iatrogenic (e.g., continuous infusion of potassium-containing fluids at an excessively rapid rate) acute mineral acidosis (e.g., hydrochloric acid or ammonium chloride) insulin deficiency (e.g., diabetic ketoacidosis) acute tumor lysis syndrome reperfusion of extremities after aortic thromboembolism in cats with cardiomyopathy hyperkalemic periodic paralysis (one case report in a pit bull) mild hyperkalemia after exercise in dogs with induced hypothyroidism infusion of lysine or arginine in total parenteral nutrition solutions nonspecific β-blockers (e.g., propranolol)* cardiac glycosides (e.g., digoxin)* urethral obstruction ruptured bladder anuric or oliguric renal failure hypoadrenocorticism selected gastrointestinal disease (e.g., trichuriasis, salmonellosis, or perforated duodenal ulcer) late pregnancy in greyhound dogs (mechanism unknown but affected dogs had gastrointestinal fluid loss) chylothorax with repeated pleural fluid drainage hyporeninemic hypoaldosteronism † angiotensin-converting enzyme inhibitors (e.g., enalapril)* angiotensin receptor blockers (e.g., losartan)* cyclosporine and tacrolimus* potassium-sparing diuretics (e.g., spironolactone, amiloride, and triamterene)* nonsteroidal antiinflammatory drugs* heparin* trimethoprim* from dibartola sp: fluid, electrolyte and acid-base disorders in small animal practice, st louis, , saunders. *likely to cause hyperkalemia only in conjunction with other contributing factors (e.g., other drugs, decreased renal function, or concurrent administration of potassium supplements). † not well documented in veterinary medicine. if refractory hypokalemia is present, supplement magnesium at . meq/kg/day for hours. alone unlikely to cause hypokalemia unless diet is aberrant administration of potassium-free (e.g., . % sodium chloride or % dextrose in water) or potassium-deficient fluids (e.g., lactated ringer's solution over several days) bentonite clay ingestion (e.g., cat litter) alkalemia insulin/glucose-containing fluids catecholamines hypothermia hypokalemic periodic paralysis (burmese cats) albuterol overdosage patients with hyponatremia or hypernatremia may have decreased, normal, or increased total body sodium content (boxes - and [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] ). an increased serum sodium concentration implies hyperosmolality, whereas a decrease in serum sodium concentration usually, but not always, implies hypoosmolality. the severity of clinical signs of hypernatremia and hyponatremia is related primarily to the rapidity of the onset of the change rather than to the magnitude of the associated plasma hyperosmolality or hypoosmolality. clinical signs of neurologic disturbances include disorientation, ataxia, and seizures, and coma may occur at serum sodium concentrations less than meq/l or greater than meq/l in dogs. therapy of hypernatremia or hyponatremia with fluid containing low or higher concentrations of sodium should proceed with caution, for rapid changes (decreases or increases) of serum sodium and osmolality can cause rapid changes in the intracellular and extracellular fluid flux, leading to intracellular dehydration or edema, even though the serum sodium has not been returned to normal. a rule of thumb is to not raise or lower the serum sodium by more than meq/l during any one -hour period. restoration of the serum sodium concentration over a period of to hours is better. in almost all circumstances, an animal will correct its sodium balance with simple fluid restoration. if severe hypernatremia exists that suggests a free water deficit, however, the free water deficit should be calculated from the following formula: hypernatremia can be corrected slowly with . % sodium chloride plus . % dextrose, % dextrose in water, or lactated ringer's solution (sodium content: meq/l). correct hyponatremia initially with . % sodium chloride. sodium is balanced predominantly by chloride and bicarbonate. the difference between these concentrations, (na , has been called the anion gap. the normal anion gap is between and meq/l. when the anion gap exceeds , consider the possibility of an accumulation of unmeasured anions (e.g., lactate, ketoacids, phosphate, sulfate, ethylene glycol metabolites, and salicylate). abnormalities in the anion gap may be helpful in determining the cause of metabolic acidosis (boxes - and - ). the colloid oncotic pressure of blood is associated primarily with large-molecular-weight colloidal substances in circulation. the major player in maintaining intravascular and interstitial oncotic pressure, the water-retaining property of each fluid compartment, is albumin. albumin contributes roughly % to the colloidal oncotic pressure of blood. the majority of albumin is located within the interstitial space. hypoalbuminemia can result from increased loss in the form of protein-losing enteropathy or nephropathy and wound exudates, or it may be due to lack of hepatic albumin synthesis. serum albumin pools are in a constant flux with interstitial albumin. once interstitial albumin pools become depleted from replenishing serum albumin, serum albumin levels can continue to decrease, which can lead to a decrease in colloidal oncotic pressure. serum albumin less than . g/dl has been associated with inadequate intravascular fluid retention and the development of peripheral edema and third spacing of fluid. oncotic pressure can be restored with the use of artificial or synthetic colloids or natural colloids (see colloids). maintenance fluid requirements have been extrapolated from the formulas used to calculate a patient's daily metabolic energy requirements because it takes ml of water to metabolize kcal of energy (table - ) . the patient's daily metabolic water (fluid) requirements can be calculated by the following formula: administration of an isotonic crystalloid fluid for maintenance requirements often can produce iatrogenic hypokalemia. in most cases, supplemental potassium must be added to prevent hypokalemia resulting from inappetance, kalliuresis, and supplementation with isotonic crystalloid fluids. the most reliable method of determining the degree of fluid deficit is by weighing the animal and calculating acute weight loss. acute weight loss in a patient with volume loss in the form of vomiting, feces, wound exudates, and urine is due to fluid loss and not loss of muscle or fat. lean body mass normally is not gained or lost rapidly enough to cause major changes in body weight. one milliliter of water weighs approximately g. this fact allows calculation of the patient's fluid deficit, if ongoing losses can be measured. when a patient first presents, however, the body weight before a fluid deficit has occurred rarely is known. instead, one must rely on subjective measures of dehydration to estimate the patient's percent dehydration and to calculate the volume of fluid required to rehydrate the patient over the next hours. to calculate the volume deficit, use the following formula: body mass (kg) × (% dehydration) × = fluid deficit (ml) the patient's fluid deficit must be added to the daily maintenance fluid requirements and administered over a -hour period. ongoing losses can be determined by measuring urine output, weighing the patient at least to times a day, and measuring the volume or weight of vomitus or diarrhea. a crystalloid fluid contains crystals of salts with a composition similar to that of the extracellular fluid space and can be used to maintain daily fluid requirements and replace fluid deficits or ongoing fluid losses (table - ) . metabolic, acid-base, and electrolyte imbalances also can be treated with isotonic fluids with or without supplemental electrolytes and buffers. depending on the patient's clinical condition, choose the specific isotonic crystalloid fluid to replace and maintain the patient's acid-base and electrolyte status ( table - ) . crystalloid fluids are readily available, are relatively inexpensive, and can be administered safely in large volumes to patients with no preexisting cardiac or renal disease or cerebral edema. following infusion, approximately % of the volume of a crystalloid fluid infused will redistribute to the interstitial fluid compartment. as such, crystalloid fluids alone are ineffective for ongoing intravascular volume depletion when given as a bolus. the crystalloid fluid bolus must be followed by a constant rate infusion, taking into consideration the patient's daily maintenance fluid requirements and ongoing fluid losses. administration of a large volume of crystalloid fluids can cause dilutional anemia and coagulopathies. * × bw kg + = kcal/day = ml/day. note: this formula will slightly underestimate the requirements for patients that are less than kg and will slightly overestimate the requirements for patients greater than kg. retain fluid in the vascular space, the volume of crystalloid fluid infused (maintenance + deficit + ongoing losses) should be decreased by % to % to avoid vascular volume overload. two major classes of colloids exist: natural and synthetic. natural colloids (whole blood, packed rbcs, plasma) are discussed elsewhere in this text. concentrated human albumin is a natural purified colloid that recently has become more popular in the treatment of advanced hypoalbuminemia and hypoproteinemia and will be discussed here. synthetic colloids are starch polymers and include dextrans and hetastarch. concentrated human albumin is available as a % or % solution. the % solution has an osmolality similar to that of serum ( mosm/l), whereas the % solution is hyperoncotic ( mosm/l). a % albumin solution draws fluid from the interstitial space into the intravascular space. concentrated albumin solutions often are used to restore circulating volume when synthetic colloids are not available. albumin not only is important at maintaining the colloidal oncotic pressure of blood but also serves as a valuable free-radical scavenger and carrier of drugs and hormones necessary for normal tissue function and healing. albumin levels less than . g/dl have been associated with increased morbidity and mortality. concentrated human albumin solutions can be administered as an effective method of restoring interstitial and serum albumin concentrations in situations of acute and chronic hypoalbuminemia. albumin ( %) is available in -and -ml vials and is more cost-efficient as an albumin replacement than procurement and administration of fresh frozen plasma. recommended albumin infusion rates are to ml/kg over hours, after pretreatment with diphenhydramine. although concentrated human albumin is structurally similar to canine albumin, closely monitor the patient for signs of allergic reaction during and after the infusion. dextran- is a synthetic high-molecular-weight polysaccharide (sucrose polymer) with a molecular weight of , d. particles less than , d, are cleared rapidly by the kidneys, whereas larger particles are cleared more slowly by the hepatic reticuloendothelial system. dextran- can coat platelets and inhibit platelet function and so must be used with caution in patients with known coagulopathies. the total daily dosage should not exceed ml/kg/day. hetastarch (hydroxyethyl starch) is a large-molecular-weight amylopectin polymer, has molecules with a molecular weight that exceeds , d, and has an average half-life of to hours in circulation. hetastarch can bind with vwf and cause prolongation of the act and aptt; however, it does not cause a coagulopathy. recommended rates of hetastarch infusion are -to -ml incremental boluses for the treatment of hypotension and to ml/kg/day as a constant rate infusion for maintenance of colloidal oncotic pressure. many are the acceptable ways to administer the fluids prescribed for each patient based on the degree of dehydration, estimation of ongoing losses, ability to tolerate oral fluid, and metabolic, acid-base, and electrolyte derangements. administer the fluids in a manner that is best for the patient and most appropriate for the practice. to determine the rate of intravenous fluid infusion, take the total volume of fluids that have been prescribed and divide the total volume by the total number of hours in a day that intravenous fluids can be delivered safely and monitored. the safest and most accurate way to deliver intravenous fluids, particularly in extremely small animals or those with congestive heart failure, is through an intravenous fluid pump. fluid should not be administered intravenously if the patient cannot be monitored to make sure that the fluids are being delivered at a safe rate and that the fluid line has not become disconnected. supplement fluids over as many hours as possible to allow the patient as much time as possible to redistribute and fully utilize the fluids administered. fluids administered too quickly can cause a diuresis to occur, such that the majority of the fluids administered will be excreted in the urine. if time is limited or if extra time is needed for safe administration of fluids, consider using a combination of intravenously and subcutaneously emergency care administered fluids. intravenous is the preferred route of administration of fluids in any patient with dehydration and hypovolemia. as intravascular volume depletion occurs, reflex peripheral vasoconstriction occurs to restore core perfusion. the subcutaneous tissue are not perfused well and therefore fluids administered subcutaneously will not be absorbed well into the interstitial and intravascular spaces. subcutaneously administered fluids can be absorbed slowly and delivered effectively in the management of mild interstitial dehydration and in the treatment of renal insufficiency. subcutaneously administered fluids should never take the place of intravenously administered fluids in a hypovolemic patient or one with severe interstitial dehydration. intramedullary (intraosseous) infusion works well in small patients in which vascular access cannot be established. shock doses of fluids and other substances, including blood products, can be administered under pressure through an intraosseous cannula. because of the inherent discomfort and risk of osteomyelitis with intraosseous infusion, establish vascular access as soon as possible. the safest and most efficient method of intravenous fluid infusion is through a fluid pump. in cases in which a fluid pump is unavailable, infusion by gravity feed is the next option. infusion sets from various manufacturers have calibrated drip chambers such that a specific number of drops will equal ml of fluid. fluid rates can be calculated based on the number of drops that fall into the drip chamber per minute: fluid volume to be infused (ml) = ml/hour number of hours available many pediatric drip sets deliver drops/ml, such that milliliters/hour equals drops/ minute. carefully record fluid orders so that the volume to be administered is recorded as milliliters/hour, milliliters/day, and drops/minute. this will allow personnel to detect major discrepancies and calculation errors more readily. the volume actually delivered should be recorded in the record by nursing personnel. all additives should be listed clearly on the bottle on a piece of adhesive tape or a special label manufactured for this purpose. a strip of adhesive tape also can be attached to the bottle and marked appropriately to provide a quick visualization of the estimate of volume delivered. includes a large-bore flexible orogastric lavage tube, permanent marker or white tape, lubricating jelly, warm water, two large buckets, a roll of -inch white tape, and a manual lavage pump. to perform the orogastric lavage, follow this procedure: . place all animals under general anesthesia with a cuffed endotracheal tube in place to protect the airway and prevent aspiration of gastric contents into the lungs. . place a roll of -inch white tape into the animal's mouth, and secure the tape around the muzzle. you will insert the tube through the hole in the center of the roll of tape. . next, place the distal end of the tube at the level of the last rib, directly adjacent to the animal's thorax and abdomen. measure the length of the tube from the most distal end to the point where it comes out of the mouth, and label this location on the tube with a permanent marker or piece of white tape. . lubricate the distal portion of the tube, and gently insert it through the roll of tape in the animal's mouth. . gently push the tube down the esophagus. palpate the tube within the esophagus. two tubes should be palpable, the orogastric tube, and the patient's trachea. push the tube down into the stomach. you can verify location by blowing into the proximal end of the tube and simultaneously auscultating the stomach for borborygmi. . insert the manual pump to the proximal end of the tube, and instill the warm water. alternate instilling water with removal of fluid and gastric debris by gravity. repeat the process until the efflux fluid is clear of any debris. . save fluid from the gastric efflux fluid for toxicologic analyses. hackett tb: emergency approach to intoxications, clin tech small anim pract ( ): - , . hypoxia, or inadequate tissue oxygenation, is the primary reason for supplemental oxygen therapy. major causes of hypoxia include hypoventilation, ventilation-perfusion mismatch, physiologic or right-to-left cardiac shunt, diffusion impairment, and decreased fraction of inspired oxygen (table - ) . inadequate tissue perfusion caused by low cardiac output or vascular obstruction also can result in circulatory hypoxia. finally, histiocytic hypoxia results from inability of cells to use oxygen that is delivered to them. this form of hypoxia can be observed with various toxin ingestions (bromethalin, cyanide) and in septic shock. a patient's oxygenation status can be monitored invasively by drawing of arterial blood gas samples or noninvasively through pulse oximetry, in most cases (see acid-base physiology and pulse oximetry). inspired air at sea level has a po of mm hg. as the air travels through the upper respiratory system to the level of the alveolus, the po drops to mm hg. tissue oxygen saturation in a normal healthy animal is mm hg. after oxygen has been delivered to the tissues, the oxygen left in the venous system (pvo ) is approximately mm hg. normally, oxygen diffuses across the alveolar capillary membrane and binds reversibly with hemoglobin in rbcs. a small amount of oxygen is carried in an unbound diffusible form in the plasma. when an animal has an adequate amount of hemoglobin and hemoglobin becomes fully saturated while breathing room air, supplemental oxygen administration will only increase the sao a small amount. the unbound form of oxygen dissolved in plasma will increase. if, however, inadequate hemoglobin saturation is obtained by breathing room air, as in a case of pneumonia or pulmonary edema, for example, breathing a higher fraction of inspired oxygen (fio ) will improve bound and unbound hemoglobin levels. the formula for calculating oxygen content of arterial blood is as follows: where cao is the arterial oxygen content, . is the amount of oxygen that can be carried by hemoglobin (hb), sao is the hemoglobin saturation, and . × pao is the amount of oxygen dissolved (unbound) in plasma. dissolved oxygen actually contributes little to the total amount of oxygen carried in the arterial blood, and the majority depends on the amount or availability of hemoglobin and the ability of the body (ph and respiratory status) to saturate the hemoglobin at the level of the alveoli. oxygen therapy is indicated whenever hypoxia is present. the underlying cause of the hypoxia also must be identified and treated, for chronic, lifelong oxygen therapy is rarely feasible in veterinary patients. if hemoglobin levels are low due to anemia, oxygen supplementation must occur along with rbc transfusions to increase hemoglobin mass. whenever possible, use arterial blood gas analyses or pulse oximetry to gauge a patient's response to oxygen therapy and to determine when an animal can be weaned from supplemental oxygen. the goal of oxygen therapy is to increase the amount of oxygen bound to hemoglobin in arterial blood. oxygen supplementation can be by hood, oxygen cage or tent, nasal or nasopharyngeal catheter, or tracheal tube. in rare cases, administration of oxygen with mechanical ventilation may be indicated. administration of supplemental oxygen to patients with chronic hypoxia is sometimes necessary but also dangerous. with chronic hypoxia the patient develops a chronic respiratory acidosis (elevated paco ) and depends almost entirely on the hypoxic ventilatory drive to breathe. administration of supplemental oxygen increases pao and may inhibit the central respiratory drive, leading to hypoventilation and possibly respiratory arrest. therefore, closely monitor animals with chronic hypoxia that are treated with supplemental oxygen. oxygen hoods can be purchased from commercial sources or can be manufactured in the hospital using a rigid elizabethan collar, tape, and plastic wrap. to make an oxygen hood, place several lengths of plastic wrap over the front of the elizabethan collar and tape them in place. leave the ventral third of the collar open to allow moisture and heat to dissipate and carbon dioxide to be eliminated. place a length of flexible oxygen tubing under the patient's collar into the front of the hood, and run humidified oxygen at a rate of to ml/kg/minute. animals may become overheated with an oxygen hood in place. carefully monitor the patient's temperature so that iatrogenic hyperthermia does not occur. commercially available plexiglass oxygen cages can be purchased from a variety of manufacturers. the best units include a mechanical thermostatically controlled compressor cooling unit, a circulatory fan, nebulizers or humidifiers to moisten the air, and a carbon dioxide absorber. alternately, a pediatric (infant) incubator can be purchased from hospital supply sources, and humidified oxygen can be run into the cage at to l/minute (depending on the size of the cage). high flow rates may be required to eliminate nitrogen and carbon dioxide from the cage. in most cases, the fio inside the cage reaches % to % using this technique. disadvantages of using an oxygen cage are high consumption/ use of oxygen, rapid decrease in the fio within the cage whenever the cage must be opened for patient treatments, lack of immediate access to the patient, and potential for iatrogenic hyperthermia. one of the most common methods for oxygen supplementation in dogs is nasal or nasopharyngeal oxygen catheters: . to place a nasal or nasopharyngeal catheter, obtain a red rubber catheter ( f to f, depending on the size of the patient). a. for nasal oxygen supplementation, measure the distal tip of the catheter from the medial canthus of the eye to the tip of the nose. b. for nasopharyngeal oxygen supplementation, measure the catheter from the ramus of the mandible to the tip of the nose. . mark the tube length at the tip of the nose with a permanent marker. . instill topical anesthetic such as proparacaine ( . %) or lidocaine ( %) into the nostril before placement. . place a stay suture adjacent to (lateral aspect) the nostril while the topical anesthetic is taking effect. . lubricate the tip of the tube with sterile lubricant. . gently insert the tube into the ventral medial aspect of the nostril to the level made with the permanent marker. if you are inserting the tube into the nasopharynx, push the nasal meatus dorsally while simultaneously pushing the lateral aspect of the nostril medially to direct the tube into the ventral nasal meatus and avoid the cribriform plate. . once the tube has been inserted to the appropriate length, hold the tube in place with your fingers adjacent to the nostril, and suture the tube to the stay suture. if the tube is removed, you can cut the suture around the tube and leave the stay suture in place for later use, if necessary. . suture or staple the rest of the tube dorsally over the nose and in between the eyes to the top of the head, or laterally along the zygomatic arch. . attach the tube to a length of flexible oxygen tubing, and provide humidified oxygen at to ml/kg/minute. . secure an elizabethan collar around the patient's head to prevent the patient from scratching at the tube and removing it. the rule of s states that if a patient's pao is less than mm hg, or if the paco is mm hg, mechanical ventilation should be considered. for mechanical ventilation, anesthetize the patient and intubate the patient with an endotracheal tube. alternately, a temporary tracheostomy can be performed and the patient can be maintained on a plane of light to heavy sedation and ventilated through the tracheostomy site. this method, a noninvasive means of determining oxygenation is through the use of pulse oximetry. a pulse oximeter uses different wavelengths of light to distinguish characteristic differences in the properties of the different molecules in a fluid or gas mixture, in this case, oxygenated (oxyhemoglobin) and deoxygenated hemoglobin (deoxyhemoglobin) in pulsatile blood. the process is termed pulse oximetry. oxyhemoglobin and deoxyhemoglobin are different molecules that absorb and reflect different wavelengths of light. oxyhemoglobin absorbs light in the infrared spectrum, allowing wavelengths of light in the red spectrum to transmit through it. conversely, deoxyhemoglobin absorbs wavelengths of the red spectrum and allows wavelengths in the infrared spectrum to transmit through the molecule. the spectrophotometer in the pulse oximeter transmits light in the red ( nanometers) and infrared ( nanometers) spectra. the different wavelengths of light are transmitted across a pulsatile vascular bed and are detected by a photodetector on the other side. the photodetector processes the amount of light of varying wavelengths that reaches it, then transmits an electrical current to a processor that calculates the difference in the amount of light originally transmitted and the amount of light of similar wavelength that actually reaches the photodetector. the difference in each reflects the amount of light absorbed in the pulsatile blood and can be used to calculate the amount or ratio of oxyhemoglobin to deoxyhemoglobin in circulation, or the functional hemoglobin saturation by the formula: where hbo is oxygenated hemoglobin, and hb is deoxygenated hemoglobin. four molecules of oxygen reversibly bind to hemoglobin for transport to the tissues. carbon monoxide similarly binds to hemoglobin and forms carboxyhemoglobin, a molecule that is detected similarly as oxygenated hemoglobin. thus sao as detected by a pulse oximeter is not reliable if carboxyhemoglobin is present. in most cases, pulse oximetry or sao corresponds reliably to the oxyhemoglobin dissociation curve. oxygen saturation greater than % corresponds to a pao greater than mm hg. above this value, large changes in pao are reflected in relatively small changes in sao , making pulse oximetry a relatively insensitive method of determining oxygenation status when pao is normal. because pulse oximetry measures oxygenated versus nonoxygenated hemoglobin in pulsatile blood flow, it is fairly unreliable when severe vasoconstriction, hypothermia, shivering or trembling, or excessive patient movement are present. additionally, increased ambient lighting and the presence of methemoglobin or carboxyhemoglobin also can cause artifactual changes in the sao , and thus the measurement is not reliable or accurate. most pulse oximeters also display a waveform and the patient's heart rate. if the photodetector does not detect a good quality signal, the waveform will not be normal, and the heart rate displayed on the monitor will not correlate with the patient's actual heart rate. the efficiency of ventilation is evaluated using the paco value on an arterial blood gas sample. alternatively, a noninvasive method to determine end-tidal carbon dioxide is through use of a capnograph. the science of capnometry uses a spectrophotometer to measure carbon dioxide levels in exhaled gas. the capnometer is placed in the expiratory limb of an anesthetic circuit. a sample of exhaled gas is aliquoted from the breath, and an infrared light source is passed across the sample. a photodetector on the other side of the sample flow measures the amount or concentration of carbon dioxide in the sample of expired gas. the calculated value is displayed as end-tidal carbon dioxide. this value also can be displayed as a waveform. when placed in graphic form, a waveform known as a capnograph is displayed throughout the ventilatory cycle. normally, at the onset of exhalation, the gas exhaled into the expiratory limb of the tubing comes from the upper airway or physiologic dead space and contains relatively little carbon dioxide. as exhalation continues, a steep uphill slope occurs as more carbon dioxide is exhaled from the bronchial tree. near the end of exhalation, the capnogram reaches a plateau, which most accurately reflects the carbon dioxide level at the level of the alveolus. because carbon dioxide diffuses across the alveolar basement membrane so rapidly, this reflects arterial carbon dioxide levels. if a plateau is not reached and notching of the waveform occurs, check the system for leaks. if the baseline waveform does not reach zero, the patient may be rebreathing carbon dioxide or may be tachypneic, causing physiologic positive end-expiratory pressure. the soda-sorb in the system should be replaced if it has expired. conversely, low end-tidal carbon dioxide may be associated with a decrease in perfusion or blood flow. decreased perfusion can be associated with low end-tidal carbon dioxide values, particularly during cardiopulmonary cerebral resuscitation. end-tidal carbon dioxide levels are one of the most accurate predictors of the efficacy of cardiopulmonary cerebral resuscitation and patient outcome. additionally, the difference between arterial carbon dioxide levels (paco ) and end-tidal carbon dioxide can be used to calculate dead-space ventilation. increases in the difference also occur with poor lung perfusion and pulmonary diffusion impairment. thoracocentesis refers to the aspiration of fluid or air from within the pleural space. thoracocentesis may be diagnostic to determine whether air or fluid is present and to characterize the nature of the fluid obtained. thoracocentesis also can be therapeutic when removing large volumes of air or fluid to allow pulmonary reexpansion and correction of hypoxemia and orthopnea. to perform thoracocentesis, follow this procedure: . first, assemble the equipment necessary (box - ). . next, clip a -cm square in the center of the patient's thorax on both sides. . aseptically scrub the clipped area. . ideally, thoracocentesis should be performed within the seventh to ninth intercostal space. rather than count rib spaces in an emergent situation, visualize the thoracic cage as a box, and the clipped area as a box within the box. you will insert your needle or catheter in the center of the box and then direct the bevel of the needle dorsally or ventrally to penetrate pockets of fluid or air present. . attach the needle or catheter hub to the length of intravenous extension tubing. attach the female port of the intravenous extension tubing to the male port of the three-way stopcock. attach the male port of the -ml syringe to one of the female ports of the three-way stopcock. the apparatus is now assembled for use. . insert the needle through the intercostal space such that the bevel of the needle initially is directed downward. . next, push down on the hub of the needle such that the needle becomes parallel with the thoracic wall. by moving the hub of the needle in a clockwise or counterclockwise manner, the bevel of the needle will move within the thoracic cavity to penetrate pockets of air or fluid. in general, air is located dorsally and fluid is located more ventrally, although this does not always occur. . aspirate air or fluid. save any fluid obtained for cytologic and biochemical analyses and bacterial culture and susceptibility testing. in cases of pneumothorax, if the thoracocentesis needs to be repeated more than times, consider using a thoracostomy tube. place a thoracostomy tube in cases of pneumothorax whenever negative suction cannot be obtained or repeated accumulation of air requires multiple thoracocentesis procedures. thoracostomy tubes also can be placed to drain rapidly accumulating pleural effusion and for the medical management of pyothorax. before attempting thoracostomy tube placement, make sure that all necessary supplies are assembled (box - ; table - ) . to place a thoracostomy tube, follow this procedure: . lay the patient in lateral recumbency. . clip the patient's entire lateral thorax. . aseptically scrub the lateral thorax. . palpate the tenth intercostal space. . have an assistant pull the patient's skin cranially and ventrally toward the point of the elbow. this will facilitate creating a subcutaneous tunnel around the thoracostomy tube. . draw up mg/kg % lidocaine ( mg/kg for cats) along with a small amount of sodium bicarbonate to take away some of the sting. . insert the needle at the dorsal aspect of the tenth intercostal space and to the seventh intercostal space. inject the lidocaine into the seventh intercostal space at the point where the trocarized thoracic drainage catheter will penetrate into the thoracic cavity. slowly infuse the lidocaine as you withdraw the needle to create an anesthetized tunnel through which to insert the catheter. . while the local anesthetic is taking effect, remove the trocar from the catheter and cut the proximal end of the catheter with a mayo scissors to facilitate adaptation with the christmas tree adapter. . attach the christmas tree adapter to the three-way stopcock and the three-way stopcock to a length of intravenous extension tubing and the -ml syringe so that the apparatus can be attached immediately to the thoracostomy tube after placement. . aseptically scrub the lateral thorax a second time and then drape it with sterile huck towels secured with towel clamps. . wearing sterile gloves, make a small stab incision at the dorsal aspect of the tenth intercostal space. . insert the trocar back into the thoracostomy drainage tube. insert the trocar and tube into the incision. tunnel the tube cranially for approximately intercostal spaces while an assistant simultaneously pulls the skin cranially and ventrally toward the point of the elbow. . at the seventh intercostal space, direct the trocar and catheter perpendicular to the thorax. grasp the catheter apparatus at the base adjacent to the thorax to prevent the trocar from going too far into the thorax. . place the palm of your dominant hand over the end of the trocar, and push the trocar and catheter into the thoracic cavity, throwing your weight into the placement in a swift motion, not by banging the butt of your hand on the end of the stylette. for small individuals, standing on a stool, or kneeling over the patient on the triage table can create leverage and make this process easier. the tube will enter the thorax with a pop. . gently push the catheter off of the stylette, and remove the stylette. . immediately attach the christmas tree adapter and have an assistant start to withdraw air or fluid while you secure the tube in place. . first, place a horizontal mattress suture around the tube to cinch the skin securely to the tube. use care to not penetrate the tube with your needle and suture. . next, place a purse-string suture around the tube at the tube entrance site. leave the ends of the suture long, so that you can create a finger-trap suture to the tube, holding the tube in place. . place a large square of antimicrobial-impregnated adhesive tape over the tube for further security and sterility. . if antimicrobial adhesive is not available, place a gauze pad × inches square over the tube, and then wrap the tube to the thorax with cotton roll gauze and elastikon adhesive tape. . draw the location of the tube on the bandage to prevent cutting it with subsequent bandage changes. an alternate technique to use if a trocar thoracic drainage catheter is not available is the following: . prepare the lateral thorax and infuse local lidocaine anesthetic as listed before. . make a small stab incision with a no. scalpel blade, as listed before. . obtain the appropriately sized red rubber catheter and cut multiple side ports in the distal end of the catheter, taking care to not cut more than % of the circumference of the diameter of the tube. . insert a rigid, long urinary catheter into the red rubber catheter to make the catheter more rigid during insertion into the pleural space. . grasp the distal end of the catheter(s) in the teeth of a large carmalt. tunnel a metzenbaum scissors under the skin to the seventh intercostal space and make a puncture through the intercostal space. . remove the metzenbaum scissors, and then tunnel the carmalt and red rubber tube under the skin to the hole created in the seventh intercostal space with the metzenbaum scissors. . insert the tips of the carmalt and the red rubber catheter through the hole, and then open the teeth of the carmalt. . push the red rubber catheter cranially into the pleural cavity. . remove the carmalt and the rigid urinary catheter, and immediately attach the suction apparatus. secure the red rubber catheter in place as listed before. placement of a temporary tracheostomy can be lifesaving to relieve upper respiratory tract obstruction, to facilitate removal of airway secretions, to decrease dead space ventilation, to provide a route of inhalant anesthesia during maxillofacial surgery, and to facilitate mechanical ventilation. in an emergent situation in which asphyxiation is imminent and endotracheal intubation is not possible, any cutting instrument placed into the trachea distal to the point of obstruction can be used. to perform a slash tracheostomy, quickly clip the fur and scrub the skin over the third tracheal ring. make a small cut in the trachea with a no. scalpel blade, and insert a firm tube, such as a syringe casing. alternately, insertion of a -gauge needle attached to intravenous extension tubing and adapted with a -ml syringe case to attach to a humidified oxygen source also temporarily can relieve obstruction until a temporary tracheostomy can be performed. in less emergent situations, place the patient under general anesthesia and intubate the patient. assemble all the equipment necessary before starting the temporary tracheostomy procedure (box - ). to perform a tracheostomy, follow this procedure: . place the patient in dorsal recumbency. . clip the ventral cervical region from the level of the ramus of the mandible caudally to the thoracic inlet and dorsally to midline. . aseptically scrub the clipped area, and then drape with sterile huck towels secured with towel clamps. . make a -cm ventral midline skin incision over the third to sixth tracheal rings, perpendicular to the trachea. . bluntly dissect through the sternohyoid muscles to the level of the trachea. . carefully pick up the fascia overlying the trachea and cut it away with a metzenbaum scissors. . place two stay sutures through/around adjacent tracheal rings. . incise in between trachea rings with a no. scalpel blade. take care to not cut more than % of the circumference of the trachea. . using the stay sutures, pull the edges of the tracheal incision apart, and insert the tracheostomy tube. the shiley tube contains an internal obturator to facilitate placement into the tracheal lumen. remove the obturator, and then insert the inner cannula, which can be removed for cleaning as needed. . once the tube is in place, secure the tube around the neck with a length of sterile umbilical tape. postoperative care of the tracheostomy tube is as important as the procedure itself. because the tracheostomy tube essentially bypasses the protective effects of the upper respiratory system, one of the most important aspects of tracheostomy tube care and maintenance is to maintain sterility at all times. any oxygen source should be humidified with sterile water or saline to prevent drying of the respiratory mucosa. if supplemental oxygen is not required, instill to ml of sterile saline every to hours to moisten the mucosa. wearing sterile gloves, remove the internal tube and place it in a sterile bowl filled with sterile hydrogen peroxide and to be cleaned every hours (or more frequently as necessary). if a shiley tube is not available, apply suction to the internal lumen of the tracheostomy tube every to hours (or more frequently as needed) with a sterile f red rubber catheter attached to a vacuum pump to remove any mucus or other debris that potentially could plug the tube. unless the patient demonstrates clinical signs of fever or infection, the prophylactic use of antibiotics is discouraged because of the risk of causing a resistant infection. after the temporary tracheostomy is no longer necessary, remove the tube and sutures, and leave the wound to heal by second intention. primary closure of the wounds could predispose the patient to subcutaneous emphysema and infection. baker gd: trans-tracheal oxygen therapy in dogs with severe respiratory compromise due to tick (i. holocyclus) toxicity, aust vet pract ( ) urohydropulsion is a therapeutic procedure for removal of uroliths from the urethra of the male dog. the technique works best if the animal is heavily sedated or is placed under general anesthesia (figure - ) . to perform urohydropulsion, follow this procedure: . place the animal in lateral recumbency. . clip the fur from the distal portion of the prepuce. . aseptically scrub the prepuce and flush the prepuce with to ml of antimicrobial flush solution. . have an assistant who is wearing gloves retract the penis from the prepuce. . while wearing sterile gloves, lubricate the tip of a rigid urinary catheter as for urethral catheterization. . gently insert the tip of the catheter into the urethra until you meet the resistance of the obstruction. . pinch the tip of the penis around the catheter. . have an assistant insert a gloved lubricated finger into the patient's rectum and press ventrally on the floor of the rectum to obstruct the pelvic urethra. . attach a -ml syringe filled with sterile saline into proximal tip of the catheter. . quickly inject fluid into the catheter and alternate compression and relaxation on the pelvic urethra such that the urethra dilates and suddenly releases the pressure, causing dislodgement of the stone. small stones may be ejected from the tip of the urethra, whereas larger stones may be retropulsed back into the urinary bladder to be removed surgically at a later time. the type of catheter that you choose for vascular access depends largely on the size and species of the patient, the fragility of the vessels to be catheterized, the proposed length of time that the catheter will be in place, the type and viscosity of the fluid or drug to be administered, the rate of fluid flow desired, and whether multiple repeated blood samples will be required (table - ) . a variety of over-the-needle, through-the-needle, and over-the-wire catheters are available for placement in a variety of vessels, including the jugular, cephalic, accessory cephalic, medial saphenous, lateral saphenous, dorsal pedal artery, and femoral artery. one of the most important aspects of proper catheter placement and maintenance is to maintain cleanliness at all times. the patient's urine, feces, saliva, and vomit are common sources of contamination of the catheter site. before placing a peripheral or central catheter in any patient, consider the patient's physical status including whether vomiting, diarrhea, excessive urination, or seizures. in a patient with an oral mass that is drooling excessively or a patient that is vomiting, peripheral cephalic catheterization may not be the most appropriate, to prevent contamination. conversely, in a patient with excessive urination or diarrhea, a lateral or medial saphenous catheter is likely to become contaminated quickly. whenever one places or handles a catheter or intravenous infusion line, the person should wash the hands carefully and wear gloves to prevent contamination of the intravenous catheter and fluid lines. one of the most common sources of catheter contamination in veterinary hospitals is through caretakers' hands. in emergent situations, placement of a catheter may be necessary under less than ideal circumstances. remove those catheters as soon as the patient is more stable, and place a second catheter using aseptic techniques. in general, once the location of the catheter has been decided, set up all equipment necessary for catheter placement before starting to handle and restrain the patient. lists the equipment needed for most types of catheter placement. after setting up all of the supplies needed, clip the fur over the site of catheter placement. make sure to clip all excess fur and long feathers away from the catheter site, to prevent contamination. for catheter placement in limbs, clip the fur circumferentially around the site of catheter placement to facilitate adherence of the tape to the limb and to facilitate catheter removal with minimal discomfort at a later date. next, aseptically scrub the catheter site with an antimicrobial scrub solution such as hibiclens. the site is now ready for catheter insertion. consider using a central venous catheter whenever multiple repeated blood samples will need to be collected from a patient during the hospital stay. central venous catheters also can be used for cvp measurement, administration of hyperoncotic solutions such as parenteral nutrition, and administration of crystalloid and colloid fluids, anesthesia, and other injectable drugs (figures - and - ) . to place a jugular central venous catheter, place the patient in lateral recumbancy and extend the head and neck such that the jugular furrow is straight. clip the fur from the ramus of the mandible caudally to the thoracic inlet and dorsally and ventrally to midline. wipe the clipped area with gauze × -inch squares to remove any loose fur and other debris. aseptically scrub the clipped area with an antimicrobial cleanser. venocaths (abbott laboratories) are a through-the-needle catheter that is contained within a sterile sleeve for placement. alternately, other over-the-wire central venous catheters can be placed by the seldinger technique. sterility must be maintained at all times, regardless of the type of catheter placed. wearing sterile gloves, drape the site of catheter placement with sterile drapes, and occlude the jugular vein at the level of the thoracic inlet. pull the clear ring and wings of emergency diagnostic and therapeutic procedures figure - : lateral thoracic radiograph of a central venous catheter. note that the tip of the catheter is inserted in its proper location, just outside of the right atrium. the catheter cover down toward the catheter itself to expose the needle. remove the guard off of the needle. lift the skin over the proposed site of catheter insertion and insert the needle under the skin, with the bevel of the needle facing up. next, reocclude the vessel and pull the skin tight over the vessel to prevent movement of the vessel as you attempt to insert the needle. in some cases, it may be difficult actually to see the vessel in obese patients. if you cannot visualize or palpate the needle, gently bounce the needle over the vessel with the bevel up. the vessel will bounce in place slightly, allowing a brief moment of visualization to facilitate catheter placement. once the vessel has been isolated and visualized, insert the needle into the vessel at a -to -degree angle. watch closely for a flash of blood in the catheter. when blood is observed, insert the needle a small distance farther, and then push the catheter and stylette into the vessel for the entire length, until the catheter and stylette can be secured in the catheter hub. if the catheter cannot be inserted fully into the vessel for its entire length, the tip of the needle may not be within the entire lumen, the catheter may be directed perivascularly, and the catheter may be caught at the thoracic flexure and may be moving into one of the tributaries that feeds the forelimb. extend the patient's head and neck, and lift the forelimb up to help facilitate placement. do not force the catheter in because the catheter potentially can form a knot and will need to be removed surgically. remove the needle from the vessel, and have an assistant place several × -inch gauze squares over the site of catheter placement with some pressure to control hemorrhage. secure the catheter hub into the needle guard, and remove the stylette from the catheter. immediately insert a -to -ml syringe of heparinized saline and flush the catheter and draw back. if you are in the correct place, you will be able to draw blood from the catheter. to secure the catheter in place, tear a length of -inch white tape that will wrap around the patient's neck. pull a small length of the catheter out of the jugular vein to make a semicircle. the semicircle should be approximately / inch in diameter. let the length of catheter lie on the skin, and then place × -inch gauze squares impregnated with antimicrobial ointment over the site of catheter insertion. secure the proximal end of white tape around the white and blue pieces of the catheter, and wrap the tape around the patient's neck so that the tape adheres to the skin and fur. repeat the process by securing the gauze to the skin with two additional lengths of white tape, starting to secure the gauze in place by first wrapping the tape dorsally over the patient's neck, rather than under the patient's neck. in between each piece of tape and bandage layer, make sure that the catheter flushes and draws back freely, or else occlusion can occur. gently wrap layers of cotton roll gauze, kling, and elastikon or vetrap over the catheter. secure a male adapter or t port that has been flushed with heparinized saline, and then label the catheter with the size and length of catheter, date of catheter placement, and initials of the person who placed the catheter. the catheter is ready for use. monitor the catheter site daily for erythema, drainage, vessel thickening, or pain upon infusion. if any of these signs occur, or if the patient develops a fever of unknown origin, remove the catheter, culture the catheter tip aseptically, and replace the catheter in a different location. as long as the catheter is functional without complications, the catheter can remain in place. central catheters also can be placed via the seldinger or over-the-wire technique. a number of companies manufacture kits that contain the supplies necessary for over-the-wire catheter placement. each kit minimally should contain an over-the-needle catheter to place into the vessel, a long wire to insert through the original catheter placed, a vascular dilator to dilate the hole in the vessel created by the first catheter, and a long catheter to place into the vessel over the wire. additional accessories can include a paper drape, sterile gauze, a scalpel blade, local anesthetic, -gauge needles, and -or -ml syringes. restrain the patient and prepare the jugular furrow aseptically as for the percutaneous through-the-needle catheter placement. the person placing the catheter should wear sterile gloves throughout the process to maintain sterility. pick up the skin over the site of catheter placement, and insert a small bleb of local anesthetic through the skin. the local anesthetic should not be injected into the underlying vessel (figure - ) . make a small nick into the skin through the local anesthetic with a no. or no. scalpel blade. use care to avoid lacerating the underlying vessel. next, occlude the jugular vein as previously described, and insert the over-the-needle catheter into the vessel. watch for a flash of blood in the catheter hub. remove the stylette from the catheter. next, insert the long wire into the catheter and into the vessel (figures - and - ) . never let go of the wire. remove the catheter, and place the vascular dilator over the wire and into the vessel (figure - ) . gently twist to place the dilator into the vessel a short distance, creating a larger hole in the vessel. the vessel will bleed more after creating a larger hole. remove the vascular dilator, and leave the wire in place within the vessel. insert the long catheter over the wire into the vessel (figure - ) . push the catheter into the vessel to the catheter hub (figure - ) . slowly thread the wire through a proximal port in the catheter. once the catheter is in place, remove the wire, and suture the catheter in place to the skin with nonabsorbable suture. cover the catheter site with sterile gauze and antimicrobial ointment, cotton roll bandaging material, gauze, and kling or vetrap. flush the catheter with heparinized saline solution, and then use the catheter for infusion of parenteral nutrition, blood products, crystalloid and colloid fluids, medications, and frequent blood sample collection. examine the catheter site daily for evidence of infection or thrombophlebitis. the catheter can remain in place as long as it functions and no complications occur. place the patient in sternal recumbency as for cephalic venipuncture. clip the antebrachium circumferentially, and wipe the area clean of any loose fur and debris (figure - ) . aseptically scrub the clipped area, and have an assistant occlude the cephalic vein at the crook of the elbow. the person placing the catheter should grasp the distal carpus with the nondominant hand and insert the over-the-needle catheter into the vessel at a -to -degree angle ( figure - ) . watch for a flash of blood in the catheter hub, and then gently push the catheter off of the stylette (figure - ) . have the assistant occlude the vessel over the catheter to prevent backflow. flush the catheter with heparinized saline solution. make sure that the skin and catheter hub are clean and dry to ensure that the tape adheres to the catheter hub and skin. secure a length of / -inch white tape tightly around the catheter and then around the limb. make sure that the catheter hub does not "spin" in the tape, or else the catheter will fall out. next, secure a second length of -inch adhesive tape under the catheter and around the limb and catheter hub (figure - ). this piece of tape helps to stabilize the catheter in place. finally, place a flushed t port or male adapter in the catheter hub and secure to the limb with white tape. make sure that the tape is adhered to the skin securely, but not so tightly as to impede venous outflow (figure - ) . the catheter site can be covered with a cotton ball impregnated with antimicrobial ointment and layers of bandage material. label all catheters with the date of placement, the type and gauge of catheter inserted, and the initials of the person who placed the catheter. the femoral artery can be catheterized for placement of an indwelling arterial catheter. indwelling arterial catheters can be used for continuous invasive arterial blood pressure monitoring and for procurement of arterial blood samples. place the patient in lateral recumbancy, and tape the down leg in an extended position. clip the fur over the femoral artery and aseptically scrub the clipped area. palpate the femoral artery as it courses distally on the medial surface of the femur and anterior to the pectineus muscle. make a small nick incision over the proposed site of catheter placement using the bevel of an -gauge needle. place a long over-the-needle catheter through the nick in the skin and direct it toward the palpable pulse. place the tip of the catheter so that the needle tip rests in the subcutaneous tissue between the artery and the palpating index finger. advance the needle steeply at a -degree angle to secure the superficial wall of the vessel and then the deep wall of the vessel. the spontaneous flow of blood in the catheter hub ensures that the catheter is figure - : catheter is taped in place with a t-port. situated in the lumen of the artery. feed the catheter off of the stylette, and cover the hub with a catheter cap. flush the catheter with sterile heparinized saline solution, and then secure it in place. some persons simply tape the catheter in place with pieces of / -and -inch adhesive tape. others use a "butterfly" piece of tape around the catheter hub and suture or glue the tape to the adjacent skin for added security. the dorsal pedal artery commonly is used for catheter placement. to place a dorsal pedal arterial catheter, place the patient in lateral recumbency. clip the fur over the dorsal pedal artery, and then aseptically scrub the clipped area. tape the distal limb so that the leg is twisted slightly medially for better exposure of the vessel, or the person placing the arterial catheter can manipulate the limb into the appropriate position. palpate the dorsal pedal pulse as it courses dorsally over the tarsus. place an over-the-needle catheter percutaneously at a -to -degree angle, threading the tip of the needle carefully toward the pulse. advance the needle in short, blunt movements, and watch the catheter hub closely for a flash of pulsating blood that signifies penetration into the lumen of the artery. then thread the catheter off of the stylette, and cover the catheter hub with a catheter cap. secure the catheter in place with lengths of / -and -inch adhesive tape as with any other intravenous catheter, and then flush it with heparinized saline solution every to hours. any vessel that can be catheterized percutaneously also can be catheterized with surgical cutdown. restrain the patient and clip and aseptically scrub the limb or jugular vein as for a percutaneous catheterization procedure. block the area for catheter placement with a local anesthetic before cutting the skin over the vessel with a no. scalpel blade. while wearing sterile gloves, pick up the skin and incise the skin over the vessel. direct the sharp edge of the blade upward to avoid lacerating the underlying vessel. using blunt dissection, push the underlying subcutaneous fat and perivascular fascia away from the vessel with a mosquito hemostat. make sure that all tissue is removed from the vessel. using the mosquito hemostat, place two stay sutures of absorbable suture under the vessel. elevate the vessel until it is parallel with the incision, and gently insert the catheter and stylette into the vessel. secure the stay sutures loosely around the catheter. suture the skin over the catheter site with nonabsorbable suture, and then tape and bandage the catheter in place as for percutaneous placement. remove catheters placed surgically as soon as possible and exchange them for a percutaneously placed catheter to avoid infection and thrombophlebitis. the most important aspect of catheter maintenance is to maintain cleanliness and sterility at all times. an indwelling catheter can remain in place for as long as it is functional and no complications occur. change the bandage whenever it becomes wet or soiled to prevent wicking of bacteria and debris from the environment into the vessel. check the bandages and catheter sites at least once a day for signs of thrombophlebitis: erythema, vessel hardening or ropiness, pain upon injection or infusion, and discharge. also closely examine the tissue around and proximal and distal to the catheter. swelling of the paw can signify that the catheter tape and bandage are too tight and are occluding venous outflow. swelling above the catheter site is characteristic of perivascular leakage of fluid and may signify that the catheter is no longer within the lumen of the vessel. remove the catheter if it is no longer functional, if there is pain or resistance upon infusion, if there is unexplained fever or leukocytosis, or if there is evidence of cellulitis, thrombophlebitis, or catheter-related bacteremia or septicemia. aseptically culture the tip of the indwelling catheter for bacteria. animals should wear elizabethan collars or other forms of restraint if they lick or chew at the catheter or bandage. catheter patency may be maintained with constant fluid infusion or by intermittent flushing with heparinized saline ( units of unfractionated heparin per to ml of saline) every hours. flush arterial catheters more frequently (every hours). disconnect intravenous connections only when absolutely necessary. wear gloves whenever handling the catheter or connections. label all fluid lines and elevate them off of the floor to prevent contamination. date each fluid line and replace it once every to hours. if an intravenous catheter cannot be placed because of small patient size, hypovolemia, hypothermia, or severe hypotension, needles can be placed into the marrow cavity of the femur, humerus, and tibia for intraosseous infusion of fluids, drugs, and blood products. this technique is particularly useful in small kittens and puppies and in exotic species. contraindications to intraosseous infusion is in avian species (which have air in their bones), fractures, and sepsis, because osteomyelitis can develop. an intraosseous catheter is relatively easy to place and maintain but can cause patient discomfort and so should be changed to an intravenous catheter as soon as vascular access becomes possible. to place an intraosseous catheter, clip and aseptically scrub the fur over the proposed site of catheter placement. the easiest place for intraosseous placement is in the intertrochanteric fossa of the femur. inject a small amount of a local anesthetic through the skin and into the periosteum where the trocar or needle will be inserted. place the patient in lateral recumbency, and grasp the leg in between your fingers, with the stifle braced against the palm of your hand. push the stifle toward the abdomen (medially) to abduct the proximal femur away from the body. this will shift the sciatic nerve out of the way of catheter placement. insert the tip of the needle through the skin and into the intertrochanteric fossa. gently push with a simultaneous twisting motion, pushing the needle parallel with the shaft of the femur, toward your palm. you may feel a pop or decreased resistance as the needle enters the marrow cavity. gently flush the needle with heparinized saline. if the needle is plugged with bone debris, remove the needle and replace it with a fresh needle of the same type and size in the hole that you have created. a spinal needle with an internal stylette also can be placed. the stylette will prevent the needle from becoming clogged with bone debris during insertion. secure the hub of the needle with a butterfly length of white adhesive tape and then suture it to the skin to keep the catheter in place. the catheter is now ready for use. the patient should wear an elizabethan collar to prevent disruption or removal of the catheter. the intraosseous catheter can be maintained as any peripheral catheter, with frequent flushing and daily evaluation of the catheter site. the definition of pain has been debated philosophically over the ages and has changed as knowledge has increased. pain is defined as an unpleasant sensory or emotional experience associated with actual or perceived tissue damage. until recognition of a noxious stimulus occurs in the cerebral cortex, no response or adaptation results. rational management of pain requires an understanding of the underlying mechanisms involved in pain and an appreciation of how analgesic agents interact to disrupt pain mechanisms. multiple factors and causes produce pain in human beings and domestic animal species. the causes of pain, psychological and physical, may derive from many different mechanisms within emergency medicine, among them trauma, infectious disease, neglect, environmental stress, surgery, and acute decompensation of chronic medical conditions. the two major classes of pain are acute and chronic pain. box - gives specific categories and causes of pain. the pain sensing and response system can be divided into the following categories: nociceptors, which detect and filter the intensity of the noxious stimuli; primary afferent nerves, which transmit impulses to the central nervous system (cns); ascending tracts, which are part of the dorsal horn and the spinal cord that conveys stimuli to higher centers in the brain; higher centers, which are involved in pain discrimination, some memory, and motor control; and modulating or descending systems, which are a means of processing, memorizing, and modifying incoming impulses. current analgesic therapies may inhibit afferent nociceptive transmission within the brain and spinal cord; directly interrupt neural impulse conduction through the dorsal horn, primary afferent nerves, or dorsal root ganglion; or prevent the nociceptor sensitization that accompanies initial pain and inflammation. the physiologic aspects of pain are believed to be produced by the transmission, transduction, and integration of initial nerve endings, peripheral neuronal input, and ascending afferent nerves via the thalamus to the cerebral cortex. ascending afferent nerves to the limbic system are believed to be responsible for the emotional aspects of pain. there are several classification schemes for different types of pain. acute pain, such as that which results from trauma, surgery, or infectious agents, is abrupt in onset, relatively short in duration, and may be alleviated easily by analgesics. in contrast, chronic pain is a long-standing physical disorder or emotional distress that is slow in onset and difficult to treat. both types of pain can be classified further based on site of origin. somatic pain arises from superficial skin, subcutaneous tissue, body wall, or appendages. visceral pain arises from abdominal or thoracic viscera and primarily is associated with serosal irritation. analgesia, then, is the loss of pain without the loss of consciousness. this is in contrast to anesthesia, which is the loss of sensation in the whole body or a part of the body with the loss of consciousness or at least depression of the cns. untreated pain causes immediate changes in the neurohormonal axis, which in turn causes restlessness, agitation, increased heart and respiratory rates, fever, and blood pressure fluctuations, all of which are detrimental to the healing of the animal. a catabolic state is created as a result of increased secretion of catabolic hormones and decreased secretion of anabolic hormones. the net effect the majority of neurohormonal changes produce is an increase in the secretion of catabolic hormones. hyperglycemia is produced and may persist because of production of glucagon and relative lack of insulin. lipolytic activity is stimulated by cortisol, catecholamines, and growth hormone. cardiorespiratory effects of pain include increased cardiac output, vasoconstriction, hypoxemia, and hyperventilation. protein catabolism is a common occurrence and major concern regarding healing. pain associated with inflammation causes increase in tissue and blood levels of prostaglandins and cytokines, both of which promote protein catabolism indirectly by increasing the energy expenditure of the body. powerful evidence indicates that local anesthetic, sympathetic agonist, and opioid neural blockade may produce a modification of the responses to these physiologic changes. variable reduction in plasma cortisol, growth hormone, antidiuretic hormone, β-endorphin, aldosterone, epinephrine, norepinephrine, and renin is based on the anesthetic technique and the drugs selected. prophylactic administration of analgesics blunts the response before it occurs; analgesics administered following perception or pain are not as effective, and higher doses are generally necessary to achieve an equivalent level of analgesia. effective pain control can be achieved only when the signs of pain can be assessed effectively, reliably, and regularly. the experience of pain is unique to each individual, which makes pain assessment difficult, especially in traumatized and critical patients. most attempts to assess clinical pain use behavioral observations and interactive variables in addition to assessment of physiologic responses such as heart rate and respiratory rate, blood pressure, and temperature. but many factors can influence the processing and outward projection of pain, including altered environments, species differences, withinspecies variations (age, breed, sex), and the type, severity, and chronicity of pain. within-species differences (age, breed, and sex) further complicate the pain assessment. most notable is that different breeds of dogs act differently when confronted with pain or fear. labrador retrievers tend to be stoic, whereas greyhounds and teacup breeds tend to react with a heightened state of arousal around even the simplest of procedures (e.g., subcutaneous injections and nail trims). the individual character and temperament of the animal further influences its response. pediatric and neonatal animals seem to have a lower threshold for pain and anxiety than older animals. in any species, the duration and type of pain make it more (acute) or less (chronic) likely to be expressed or exhibited outwardly. unfamiliarity with normal behaviors typical of a particular species or breed makes recognition of their painful behaviors and responses impossible. the definition and recognition of pain in an individual animal is challenging. because of all the differences discussed, there is no straight line from insult, albeit actual or perceived, to degree of pain experienced. nor is there a formula for treating "x" type of pain with "y" type of analgesic. a goal of analgesia is to treat all animals with analgesic drugs and modalities as preemptively as possible and using a multimodal approach. use analgesic treatment as a tool for diagnosis of pain in the event that recognition of these phenomena is difficult for the patient. in other words, with countless drugs and treatment modalities available, analgesic administration should never be withheld in an animal, even if pain is questionable. it is important to remember that no behavior or physiologic variable in and of itself is pathognomonic for pain. interactive and unprovoked (noninteractive) behavior assessments and trending of physiologic data are useful to determine the pain in an individual animal. this is known as pain scoring. baseline observations, especially those observations from someone who has known the animal well, can be helpful to serial behavior and pain assessments. pain scoring systems have been developed and are reviewed elsewhere; the purposes of these systems are to evaluate and to help guide diagnostic and analgesic treatments (table - ) . regardless of the scale or method used to assess pain, the caregiver must recognize the limitations of the scale. if in doubt of whether pain is present or not, analgesic therapy should be used as a diagnostic tool. classic behaviors associated with pain in dogs and cats include abnormal postures, gaits, movements, and behaviors (boxes - and . stoicism is the apparent apathy and pain: assessment, prevention, and management indifference in the presence of pain and is perhaps the no. sign of ineffective pain relief or persistent pain in many animals, because so many display apathy and classically normal physiologic parameters even in the face of severe distress, overt suffering, or blatant trauma and illness. the absence of normal behaviors is also a clinical sign of pain, even when abnormal behaviors are not observed. acute pain results in many of the aforementioned behavioral and physiologic signs, but chronic pain in small animals is an entirely different and distinct entity. chronic pain is often present in the absence of obvious tissue pathology and changes in physical demeanor. again, the severity of the pain may not correlate with the severity of any pathologic condition that may or may not be present. chronic pain, especially if insidious in onset (cancer, dental, or degenerative pain), may well go unnoticed in dogs and cats, even by family members or intermittent caregivers. inappetance, lack of activity, panting in a species classically designed to be nose breathers, decreased interest in surroundings, different activity patterns, and abnormal postures are just a few signs of chronic pain in cats and dogs. cats are a species that in particular are exemplary in their abilities to hide chronic pain. they will exhibit marked familial withdrawal, finding secluded areas where they may remain for days to weeks when they experience acute and chronic pain. when deciding on a pain management protocol for a patient, always perform a thorough physical examination and include a pain score assessment before injury and pain has occurred, whenever possible. form a problem list to guide your choice of anesthesia and analgesia. for example, using a nonsteroidal antiinflammatory drug (nsaid) in an animal with renal failure would not be wise. remember to account for current medications that the patient may be taking that may augment or interfere with the analgesic or anesthetic drugs. use multimodal techniques and regional therapy and drugs to target pain at different sites before it occurs. once a strategy is decided upon, frequently reassess the patient and tailor the protocol to meet each patient's response and needs. drug therapy (in particular, opioids with or without α -agonists) is a cornerstone for acute pain treatment and surgical preemptive pain prevention. however, local anesthetics delivered epidurally, via perineural or plexus injection, intraarticular or trigger point injection, are also effective analgesics for acute and chronic forms of pain and inflammation. the nsaids that classically have been reserved for treatment of more chronic or persistent pain states now are being used regularly for treatment of acute and perioperative pain once blood pressure, coagulation, and gastrointestinal parameters have been normalized. an opioid is any natural or synthetic drug that is derived from the poppy, which interacts with opiate receptors identified on cell membranes. the drugs from this class constitute the most effective means of controlling acute, perioperative, and chronic pain in human and veterinary medicine (table - ) . their physiologic effects result from the interaction with one or more of at least five endogenous opioid receptors (µ, σ, δ, ε, and κ). µ-receptor agonists are noted for their ability to produce profound analgesia with mild sedation. these drugs diminish "wind-up," the hyperexcitable state resulting from an afferent volley of nociceptive impulses. they elevate the pain threshold and are used preemptively to prevent acute pain. as a class, opioids cause cns depression with their intense analgesia. dose-related respiratory depression reflects diminished response to carbon dioxide levels. cardiac depression is secondary only to bradycardia and is more likely with certain opioids such as morphine and oxymorphone. narcotics produce few if any clinically significant cardiovascular effects in dogs and cats; they are considered cardiac soothing or sparing. because opioids increase intracranial and intraocular pressure, use them more cautiously in patients with severe cranial trauma and or ocular lesions. opioids directly stimulate the chemoreceptor trigger zone and may cause nausea and vomiting. most opioids depress the cough reflex via a central mechanism; this may be helpful in patients recovering from endotracheal intubation irritation. a key characteristic of opioids that makes them desirable for use in emergency and critical care situations is their reversibility. antagonists block or reverse the effect of agonists by combining with receptors and producing minimal or no effects. administer all reversal agents, such as naloxone and naltrexone, slowly if given intravenously and to effect. α α -agonists as a class of drugs, α -agonists warrant special attention because most members of the group possess potent analgesic power at doses that are capable of causing sedation, cns depression, cardiovascular depression, and even general anesthetic states. originally developed for antihypertensive use, α -agonists quickly have attained sedative analgesic status in veterinary medicine (table - ) . like the opioids, α -agonists produce their effects by aggravating α-adrenergic receptors in the cns and periphery. emergency care among them cyclooxygenase- (cox- ), the major constitutive enzyme primarily involved in normal physiologic functions, and cox- , the enzyme responsible for most of the hyperalgesia and pain responses experienced after tissue injury or trauma. some nsaids inhibit cyclooxygenase and lipoxygenase activity. most of the currently available oral and parenteral nsaids for small animal medicine and surgery target the cyclooxygenase pathways predominantly, although one (tepoxalin) is thought to inhibit both pathways. inhibition of cox- and cox- can inhibit the protective effects and impair platelet aggregation and lead to gastrointestinal ulceration. there are definite contraindications and relative contraindications for the use of nsaids. nonsteroidal antiinflammatory drugs should not be administered to patients with renal or hepatic insufficiency, dehydration, hypotension or conditions that are associated with low circulating volume (congestive heart failure, unregulated anesthesia, shock), or evidence of ulcerative gastrointestinal disease. trauma patients should be stabilized completely regarding vascular volume, tone, and pressure before the use of nsaids. patients receiving concurrent administration of other nsaids or corticosteroids, or those considered to be cushingoid, should be evaluated carefully for an adequate "washout" period (time of clearance of drug from the system) before use of an nsaid or before switching nsaids. patients with coagulopathies, particularly those that are caused by platelet number or function defects or those caused by factor deficiencies, and patients with severe, uncontrolled asthma or other bronchial disease are probably not the patients in which to use nsaids. other advice is that nsaids not be administered to pregnant patients or to females attempting to become pregnant because cox- induction is necessary for ovulation and subsequent implantation of the embryo. the administration of nsaids should be considered only in the well-hydrated, normotensive dog or cat with normal renal or hepatic function, with no hemostatic abnormalities, and no concurrent steroid administration. nonsteroidal antiinflammatory drugs can be used in many settings of acute and chronic pain and inflammation. among these are the use in well-stabilized musculoskeletal trauma and surgical pain, osteoarthritis management, meningitis, mastitis, animal bite and other wound healing, mammary or transitional cell carcinoma, epithelial (dental, oral, urethral) inflammation, ophthalmologic procedures, and dermatologic or otic disease. whereas opioids seem to have an immediate analgesic effect when administered, most nsaids will take up to minutes for their effect to be recognized. as such, most perioperative or acute nsaids use is part of a balanced pain management scheme, one that uses narcotics and local anesthetic techniques. nonsteroidal antiinflammatory drugs are devoid of many of the side effects of narcotic administration; namely, decreased gastrointestinal motility, altered sensorium, nausea/vomition, and sedation. nonsteroidal antiinflammatory drugs are also devoid of many of the side effects of steroid administration; namely, suppression of the pituitary adrenal axis. the toxic effects of salicylates in cats are well documented. cats are susceptible because of slow clearance and dose-dependent elimination because of deficient glucuronidation in this species. because of this, the dose and the dosing interval of most commonly used nsaids need to be altered in order for these drugs to be used. cats that have been given canine doses of nsaids (twice daily or even once daily repetitively) may show hyperthermia, hemorrhagic or ulcerative gastritis, kidney and liver injury, hyperthermia, respiratory alkalosis, and metabolic acidosis. acute and chronic toxicities of nsaids have been reported in cats, especially after repeat once daily dosing. ketoprofen, flunixin, aspirin, carprofen, and meloxicam have been administered safely to cats, although like most antibiotics and other medications, they are not approved and licensed for use in cats. an important note, though, is that dosing intervals ranging from to hours have been used, and antithrombotic effects often can be achieved at much lower doses than those required to treat fevers and inflammation. i recommend the use of no loading doses, minimum -hour dosing intervals, and assurance of adequate circulating blood volume, blood pressure, and renal function. because many of the nsaids are used off-label in cats, it is imperative that the clinician carefully calculate the dose, modify the dosing interval, and communicate this information to the client before dispensing the drug. even drugs that come in liquid form (meloxicam), if administered to cats via box-labeled directions used for dogs, will be given in near toxic doses. to worsen the misunderstanding about dosages for cats, drops from manufacturer's bottles often are calibrated drops; when these same liquids are transferred into pharmacy syringes for drop administration, the calibration of course is lost, and the animal potentially is overdosed. a more accurate method of dispensing and administering oral nsaids in cats is to calculate the dose in milligrams and determine the exact number of milliliters to administer, rather than use the drop method. ketamine classically was considered a dissociative anesthetic, but it also has potent activity as an n-methyl-d-aspartate (nmda) receptor antagonist. this receptor located in the cns mediates windup and central sensitization (a pathway from acute to chronic pain). blockade of this receptor with microdoses of ketamine results in the ability to provide body surface, somatic, and skin analgesia with potentially lower doses of opioids and α-agonists. loading doses of . to mg/kg are used intravenously with continuous rate infusions of to µg/kg/minute. in and of itself, this drug possesses little to no analgesic ability and indeed in high doses alone often can aggravate, sensitize, or excite the animal in subacute or acute pain. amantadine is another nmda blocker that has been used for its antiviral and parkinson's stabilizing effects. amantadine has been used for neuropathic pain in human beings but is only available in an oral form. suggested starting doses for cats and dogs range from to mg/kg po daily. when the drug is given orally and intravenously, patients are unlikely to develop behavioral or cardiorespiratory effects with ketamine or amantadine. tramadol is an analgesic that possesses weak opioid µ-agonist activity and norepinephrine and serotonin reuptake inhibition. tramadol is useful for mild to moderate pain in small animals. although the parent compound has very weak opioid activity, the metabolites have excellent binding affinity for the µ-receptor. tramadol has been used for perisurgical pain control when given orally in cats and dogs at a dose of to mg/kg po sid to bid. cats appear to require only once daily dosing. regardless of its affinity for the opioid receptors, the true mechanism of action of tramadol in companion animals remains largely unknown. gabapentin is a synthetic analog of γ-aminobutyric acid (gaba). originally introduced as an antiepileptic drug, the mechanism of action of gabapentin remains somewhat unclear in veterinary medicine. the drug is among a number of commonly used antiepileptic medications used to treat central pain in human beings. the rationale for use is the ability of the drugs to suppress discharge in pathologically altered neurons. gabapentin does this through calcium channel modulation without binding to glutamate receptors. chronic, burning, neuropathic, and lancinating pain in small animals responds well to to mg/kg po daily. local anesthetic agents are the major class used as a peripheral-acting analgesic ( table - ) . local anesthetics block the transmission of pain impulses at the peripheral nerve nociceptor regions. local anesthetics may be used to block peripheral nerves or inhibit nerve "zones" using regional techniques. although all local anesthetics are capable of providing pain relief, agents with a longer duration of action are preferred for pain management purposes. bupivacaine is an example of a long-acting local anesthetic drug that is used along with lidocaine for long-acting pain relief. a single dose of bupivacaine injected at a local site will provide local anesthesia and analgesia for to hours. when lidocaine is administered as an intravenous constant rate infusion ( to µg/kg/minute in dogs, to µg/kg/minute in cats) is effective in the treatment of chronic neuropathic pain and periosteal and peritoneal pain (e.g., pancreatitis). mexiletine, an oral sodium channel blocker, can be used as an alternative to injectable lidocaine for provision of background analgesia. many drugs (table - ) are used in combination with opioids, α -agonists, and ketamine to provide anxiolysis and sedation. injection of local anesthetic solution into the connective tissue surrounding a particular nerve produces loss of sensation (sensory blockade) and/or paralysis (motor nerve blockade) in the region supplied by the nerve. local anesthetics also may be administered epidurally, intrathoracically, intraperitoneally, and intraarticularly. lidocaine and bupivacaine are the most commonly administered local anesthetics. lidocaine provides for quick, short-acting sensory and motor impairment. bupivacaine provides for later-onset, longerlasting desensitization without motor impairment. combinations of the two agents diluted with saline are used frequently to provide for quick-onset analgesia that lasts between and hours in most patients. adding narcotic and/or α agent often maximizes the analgesia and increases the pain-free interval to to hours. epinephrine and preservative-free solutions are recommended. precision placement of anesthetic close to nerves, roots, or plexuses is improved with the use of a stimulating nerve locator. cats seem to be more sensitive to the effects of local anesthetics; as such the lower ends of most dosing ranges are used for blockades in this species. unlike most instances of general anesthesia, during which the animal is rendered unconscious and nerve transmission is decreased by virtue of cns depression, local and regional techniques block the initiation of noxious signals, thereby effectively preventing pain from entering the cns. this is an effective means of not only preventing initial pain but also reducing the changes that take place in the dorsal horn of the spinal cord, spinothalamic tracts, limbic and reticular activating centers, and cortex. frequently, the neurohormonal response that is stimulated in pain and stress is blunted as well. overall, the patient has fewer local and systemic adverse effects of pain, disease processes are minimized, chronic pain states are unlikely, and outcome is improved. regional techniques are best used as part of an analgesic regimen that consists of their continuous administration, narcotics, α-agonists, anxiolytics, and good nursing. lidocaine can be added to sterile lubricant in a one-to-one concentration to provide decreased sensation for urinary catheterization, nasal catheter insertion, minor road burn analgesia, and pyotraumatic dermatitis analgesia. proparacaine is a topical anesthetic useful for corneal or scleral injuries. local anesthetics can be used to infiltrate areas of damage or surgery by using long-term continuous drainage catheters and small, portable infusion pumps. this is an effective means of providing days of analgesia for massive surgical or traumatic soft tissue injury. even without the catheter, incisional or regional soft tissue blocking using a combination of to mg/kg lidocaine and . to mg/kg bupivacaine diluted with equal volume of saline and : with sodium bicarbonate is effective for infiltrating large areas of injury. administration of local anesthetic drugs around the infraorbital, maxillary, ophthalmic mental, and alveolar nerves can provide excellent analgesia for dental, orofacial, and ophthalmic trauma and surgical procedures. each nerve may be desensitized by injecting . to . ml of a % lidocaine hydrochloride solution and . to . ml of . % bupivacaine solution using a . -to . -cm, -to -gauge needle. precise placement perineurally versus intraneurally (neuroma formation common) is enhanced by using catheters in the foramen versus needle administration. always perform aspiration before administration to rule out intravascular injection of agents. this block is used to provide analgesia for thoracic, lower cervical, cranial abdominal, and diaphragmatic pain. following aseptic preparation, place a small through-the-needle ( -to -gauge) catheter in the thoracic cavity between the seventh and ninth intercostal space on the midlateral aspect of the thorax. aseptically mix a . to mg/kg lidocaine and a . to . mg/kg bupivacaine dose with volume of saline equal to the volume of bupivacaine, and slowly inject it over a period of to minutes following aspiration to ensure that no intravascular injection occurs. depending on where the lesion is, position the patient to allow the intrapleural infusion to "coat" the area. most effective is positioning the patient in dorsal recumbency for several minutes following the block to make sure local anesthetic occupies the paravertebral gutters and hence the spinal nerve roots. the block should be repeated every hours in dogs and every to hours in cats. secure the catheter to the skin surface for repetitive administration. administration of local anesthetic around the brachial plexus provides excellent analgesia for forelimb surgery, particularly that distal to the shoulder, and amputations. nerve locator-guided techniques are much more accurate and successful than blind placement of local anesthetic; however, even the latter is useful. to administer a brachial plexus blockade, follow this procedure: . aseptically prepare a small area of skin over the point of the shoulder. . insert a -gauge, / -to -inch spinal needle medial to the shoulder joint, axial to the lesser tubercle, and advance it caudally, medial to the body of the scapula, and toward the costochondral junction of the first rib. aspirate first before injection to make sure that intravenous injection does not occur. . inject one third of the volume of local anesthetic mix, and then slowly withdraw the needle and fan dorsally and ventrally while infusing the remaining fluid. . local anesthetic doses are similar to those for intrapleural blockade. epidural analgesia refers to the injection of an opioid, a phencyclidine, an α-agonist, or an nsaid into the epidural space. epidural anesthesia refers to the injection of a local anesthetic. in most patients a combination of the two is used. epidural analgesia and anesthesia are used for a variety of acute and chronic surgical pain or traumatically induced pain in the pelvis, tail, perineum, hind limbs, abdomen, and thorax (table - ) . procedures in which epidural analgesia and anesthesia are useful include forelimb and hind limb amputation, tail or perineal procedures, cesarean sections, diaphragmatic hernia repair, pancreatitis, peritonitis, and intervertebral disk disease. epidural blocks performed using opioids or bupivacaine will not result in hind limb paresis or decreased urinary or anal tone (incontinence), unlike lidocaine or mepivicaine epidural blocks. morphine is one of the most useful opioids for administration in the epidural space because of its slow systemic absorption. epidural catheters used for the instillation of drugs through constant rate infusion or intermittent injection can be placed in dogs and cats. routinely placed at the lumbosacral junction, these catheters are used with cocktails including preservative-free morphine, bupivacaine, medetomidine, and ketamine. extremely effective for preventing windup pain in the peritoneal cavity or caudal half of the body, the catheters may be maintained if placed aseptically for to days. to provide epidural analgesia or anesthesia, follow this procedure: . position the animal in lateral or sternal recumbency. . clip and aseptically scrub over the lumbosacral site. . palpate the craniodorsal-most extent of the wings of the ileum bilaterally and draw an imaginary line through them to envision the spine of l located immediately behind the imaginary line. . advance a -to -gauge, / -to -inch spinal or epidural needle through the skin just caudal to the spine of l . . the needle will lose resistance as it is introduced into the epidural space. drop saline into the hub of the needle, and the saline will be pulled into the epidural space as the needle enters. discrete intercostal nerve blocks can provide effective analgesia for traumatic or postsurgical pain. identify the area of the injury, and infiltrate three segments on either side of the injury with analgesic. to perform an intercostal nerve block, follow this procedure: . clip and aseptically scrub the dorsal and ventral third of the chest wall. . palpate the intercostal space as far dorsally as possible. . use a -gauge, . -inch needle at the caudolateral aspect of the affected rib segments and those cranial and caudal. . direct the tip of the needle caudally such that the tip of the needle "drops" off of the caudal rib. (this places the needle tip in proximity to the neuromuscular bundle that contains the intercostal nerve that runs in a groove on the caudomedial surface of the rib.) . aspirate to confirm that the drug will not go intravenously. . inject while slowly withdrawing the needle. inject . to . ml at each site, depending on the size of the animal. gaynor js, an acute condition in the abdomen is defined as the sudden onset of abdominal discomfort or pain caused by a variety of conditions involving intraabdominal organs. many animals have the primary complaint of lethargy, anorexia, ptyalism, vomiting, retching, diarrhea, hematochezia, crying out, moaning, or abnormal postures. abnormal postures can include generalized rigidity, walking tenderly or as if "on eggshells," or a prayer position in which the front limbs are lowered to the ground while the hind end remains standing. in some cases, it may be difficult initially to distinguish between true abdominal pain or referred pain from intervertebral disk disease. rapid progression and decompensation of the patient's cardiovascular status can lead to stupor, coma, and death in the most extreme cases, making rapid assessment, treatment, and definitive care extremely challenging. often the patient's signalment and history can increase the index of suspicion for a particular disease process. a thorough history often is overlooked or postponed in the initial stages of resuscitation of the patient with acute abdominal pain. often, asking the same question in a variety of methods can elicit an answer from the client that may lead to the source of the problem and the reason for acute abdominal pain. important questions to ask the client include the following: • what is your chief complaint or reason that you brought your animal in on emergency? • when did the signs first start, or when was your animal last normal? • do you think that the signs have been the same, better, or getting worse? • does your animal have any ongoing or past medical problems? • have similar signs occurred in the past? • does your animal have access to any known toxins, or does he or she run loose unattended? as with any other emergency, the clinician must follow the abcs of therapy, treating the most life-threatening problems first. first, perform a perfunctory physical examination. examination of the abdomen ideally should be performed last, in case inciting a painful stimulus precludes you from evaluating other organ systems more thoroughly. briefly observe the patient from a distance. are there any abnormal postures? is there respiratory distress? is the animal ambulatory, and if so, do you observe any gait abnormalities? do you observe any ptyalism or attempts to vomit? auscultate the patient's thorax for crackles that may signify aspiration pneumonia resulting from vomiting. examine the patient's mucous membrane color and capillary refill time, heart rate, heart rhythm, and pulse quality. many patients in pain have tachycardia that may or may not be accompanied by dysrhythmias. if a patient's heart rate is inappropriately bradycardic, consider hypoadrenocorticism, whipworm infestation, or urinary obstruction or trauma as a cause of hyperkalemia. assess the patient's hydration status by evaluating skin turgor, mucous membrane dryness, and whether the eyes appear sunken in their orbits. a brief neurologic examination should consist of whether the patient is actively having a seizure, or whether mental dullness, stupor, coma, or nystagmus are present. posture and spinal reflexes can assist in making a diagnosis of intervertebral disk disease versus abdominal pain. perform a rectal examination to evaluate for the presence of hematochezia or melena. finally, examination of the abdomen should proceed first with superficial and then deeper palpation. visually inspect the abdomen for the presence of external masses, bruising, or penetrating injuries. reddish discoloration of the periumbilical area often is associated with the presence of intraabdominal hemorrhage. it may be necessary to shave the fur to inspect the skin and underlying structures visually for bruising and ecchymoses. auscultate the abdomen for the presence or absence of borborygmi to characterize gut sounds. next, perform percussion and ballottement to evaluate for the presence of a gas-distended viscus or peritoneal effusion. finally, perform first superficial and then deep palpation of all quadrants of the abdomen, noting abnormal enlargement, masses, or whether focal pain is elicited in any one area. once the physical examination has been performed, implement initial therapy in the form of analgesia, fluid resuscitation, and antibiotics. treatment for any patient with an acute condition in the abdomen and shock is to treat the underlying cause, maintain tissue oxygen delivery, and prevent end-organ damage and failure. a more complete description of shock and oxygen delivery is given in the section on shock. emergency care the administration of analgesic agents to any patient with acute abdominal pain is one of the most important therapies in the initial stages of case management. many patients with acute abdominal pain are clinically dehydrated or are in hypovolemic shock because of hemorrhage. careful titration of intravenous crystalloid and colloid fluids including blood products is necessary based on the patient's perfusion parameters including heart rate, capillary refill time, blood pressure, urine output, and pcv. fluid therapy also should be based on the most likely differential diagnoses, with specific fluid types administered according to the primary disease process. in dogs, a shock volume of fluids is calculated based on the total blood volume of ml/kg/hour. in cats, shock fluid rate is based on plasma volume of ml/kg/hour. in most cases, any crystalloid fluid can be administered at an initial volume of one fourth of a calculated shock dose and then titrated according to whether the patient's cardiovascular status responds favorably or not. in cases of an acute condition in the abdomen from known or suspected hypoadrenocorticism, severe whipworm infestation, or urinary tract obstruction or rupture, . % sodium chloride fluid without added potassium is the fluid of choice. when hemorrhage is present, the administration of whole blood or packed rbcs may be indicated if the patient has clinical signs of anemia and shows clinical signs of lethargy, tachypnea, and weakness. fresh frozen plasma is indicated in cases of hemorrhage resulting from vitamin k antagonist rodenticide intoxication or hepatic failure or in cases of suspected disseminated intravascular coagulation (dic). a more thorough description of fluid therapy is given under the sections on shock and fluid therapy. the empiric use of broad-spectrum antibiotics is warranted in cases of suspected sepsis or peritonitis as a cause of acute abdominal pain. ampicillin sulbactam ( mg/kg iv q - h) and enrofloxacin ( mg/kg once daily) are the combination treatment of choice to cover gram-negative, gram-positive, aerobic, and anaerobic infections. alternative therapies include a second-generation cephalosporin such as cefotetan ( mg/kg iv tid) or cefoxitin ( mg/kg iv tid) or added anaerobic coverage with metronidazole ( to mg/kg iv tid). tissue oxygen delivery depends on a number of factors, including arterial oxygen content and cardiac output. if an animal has had vomiting and subsequent aspiration pneumonitis, treatment of hypoxemia with supplemental oxygen in the form of nasal, nasopharyngeal, hood, or transtracheal oxygen administration is important (see oxygen supplementation under emergency diagnostic and therapeutic procedures). perform a complete blood count in all cases of acute abdominal pain to determine if lifethreatening infection or coagulopathy including dic is present. in cases of sepsis, infection, or severe nonseptic inflammation, the white blood cell count may be normal, elevated, or low. examine a peripheral blood smear for the presence of toxic neutrophils, eosinophils, atypical lymphocytes, nucleated rbcs, platelet estimate, anisocytosis, and blood parasites. a falling pcv in the face of rbc transfusion suggests ongoing hemorrhage. perform a biochemistry panel to evaluate organ system function. azotemia with elevated bun and creatinine may be associated with prerenal dehydration, impaired renal function, or postrenal obstruction or leakage. the bun also can be elevated when gastrointestinal hemorrhage is present. serum amylase may be elevated with decreased renal function or in cases of pancreatitis. a normal serum amylase, however, does not rule out pancreatitis as a source of abdominal pain. serum lipase may be elevated with gastrointestinal inflammation or pancreatitis. like amylase, a normal serum lipase does not rule out pancreatitis. total bilirubin, alkaline phosphatase, and alanine transaminase may be elevated with primary cholestatic or hepatocellular diseases or may be due to extrahepatic causes including sepsis. obtain a urinalysis via cystocentesis whenever possible, except in cases of suspected pyometra or transitional cell carcinoma. azotemia in the presence of a nonconcentrated (isosthenuric or hyposthenuric) urine suggests primary renal disease. secondary causes of apparent renal azotemia and lack of concentrating ability also occur in cases of hypoadrenocorticism and gram-negative sepsis. renal tubular casts may be present in cases of acute renal ischemia or toxic insult to the kidneys. bacteriuria and pyuria may be present with infection and inflammation. when a urinalysis is obtained via free catch or urethral catheterization, the presence of bacteriuria or pyuria also may be associated with pyometra, vaginitis, or prostatitis/prostatic abscess. serum lactate is a biochemical indicator of decreased organ perfusion, decreased oxygen delivery or extraction, and end-organ anaerobic glycolysis. elevated serum lactate greater than mmol/l has been associated with increased morbidity and need for gastric resection in cases of gdv and increased patient morbidity and mortality in other disease processes. rising serum lactate in the face of adequate fluid resuscitation is a negative prognostic sign. obtain abdominal radiographs as one of the first diagnostic tests when deciding whether to pursue medical or surgical management. the presence of gdv, linear foreign body, pneumoperitoneum, pyometra, or splenic torsion warrants immediate surgical intervention. if a loss of abdominal detail occurs because of peritoneal effusion, perform additional diagnostic tests including abdominal paracentesis (abdominocentesis) and abdominal ultrasound to determine the cause of the peritoneal effusion. abdominal ultrasonography is often useful in place of or in addition to abdominal radiographs. the sensitivity of abdominal ultrasonography is largely operator dependent. indications for immediate surgical intervention include loss of blood flow to an organ, linear bunching or placation of the intestinal tract, intussusception, pancreatic phlegmon or abscess, a fluid-filled uterus suggestive of pyometra, gastrointestinal obstruction, intraluminal gastrointestinal foreign body, dilated bile duct, or gallbladder mucocele, or gas within the wall of the stomach or gallbladder (emphysematous cholecystitis). the presence of peritoneal fluid alone does not warrant immediate surgical intervention without cytologic and biochemical evaluation of the fluid present. see also abdominal paracentesis and diagnostic peritoneal lavage. abdominal paracentesis (abdominocentesis) often is the deciding factor in whether to perform immediate surgery. abdominocentesis is a sensitive technique for detecting peritoneal effusion when more than ml/kg of fluid is present within the abdominal cavity. abdominal effusion collected should be saved for bacterial culture and evaluated biochemically and cytologically based on your index of suspicion of the primary disease process. if creatinine, urea nitrogen (bun) or potassium is elevated compared with that of serum, uroabdomen is present. elevated abdominal fluid lipase or amylase compared with serum supports a diagnosis of pancreatitis. elevated lactate compared with serum lactate or an abdominal fluid glucose less than mg/dl is highly sensitive and specific for bacterial/ septic peritonitis. the presence of bile pigment or bacteria is supportive of bile and septic peritonitis, respectively. free fibers in abdominal fluid along with clinical signs of abdominal pain strongly support gastrointestinal perforation, and immediate surgical exploration is required. text continued on p. the following are clinical conditions, patient signalment, common history, physical examination, and characteristic findings of various diagnostic tests. a blank column next to a condition indicates no specific signalment, history, physical examination, or diagnostic test characteristic for a particular disease process. lack of contiguity of body wall surgical ( medical unless perforation present present c-shaped abnormal gas pattern with plication on radiographs surgical (immediate) dilation of bowel cranial to foreign object, radiopaque object in surgical (immediate) stomach or intestines, hypochloremic metabolic acidosis on bloodwork if pyloric outflow obstruction is present elevated or decreased wbc; foreign material, wbcs and medical unless perforation bacteria on abdominal fluid, elevated lactate and decreased present glucose on abdominal fluid target shaped soft tissue density on abdominal u/s, soft tissue surgical (immediate): density with gas dilation cranially on abdominal radiographs medical management of primary cause colonic distension with hard feces on radiographs medical increased or decreased wbc, septic abdominal effusion surgical (immediate) elevated t bili, alt, alk phos, and wbc hypoechoic hepatic medical after biopsy parenchyma on ultasound hepatomegaly elevated t bili, alt, alk phos, and wbc hyperechoic foci in surgical (immediate) gallbladder or sludge on u/s, free gas in wall of gall bladder abdominal effusion, bile pigment in effusion surgical (immediate) elevated t bili, alk phos, alt surgical (immediate) elevated or decreased wbc, elevated t bili, alk phos and surgical (immediate) alt, free gas in hepatic parenchyma on rads, hypoechoic mass with hyperechoic material in hepatic parenchyma on u/s heteroechoic liver with hyperechoic center on ultrasound surgical (immediate) mixed echogenic mass on ultrasound, soft tissue mass surgical (immediate or density on radiographs, elevated alk phos, alt, delayed) t bili, hypoglycemia pain-cont'd elevated t bili, alk phos, alt, amylase and/or lipase, elevated medical in most cases or decreased wbc, hypocalcemia, focal loss of detail in right unless abscess or cranial quadrant on radiographs hypo-to hyperechoic phlegmon is present pancreas with hyperechoic peri-pancreatic fat on ultrasound, abdominal and/or pleural effusion on radiographs and ultrasound pancreatic soft tissue mass effect on radiographs and surgical if mass identified, ultrasound, elevated amylase and lipase, hypoglycemia, otherwise medical elevated serum insulin management of hypoglycemia splenomegaly on radiographs, hyperechoic spleen with no surgical (immediate) blood flow on ultrasound soft tissue mass effect and loss of abdominal detail on surgical (immediate) radiographs, cavitated mass with abdominal effusion on u/s hyperechoic spleen with no blood flow on abdominal u/s, surgical (immediate) abdominal effusion, thrombocytopenia loss of abdominal detail on radiographs, peritoneal effusion medical unless refractory on u/s, hemoabdomen on abdominocentesis hypotension diagnosis based primarily on clinical signs medical fracture of the os penis on radiographs largely medical unless urethral tear diagnosis based primarily on clinical signs medical, although prepuce may need to be incised to allow replacement of penis into sheath prostatomegaly on radiographs and ultrasound hypoechoic medical prostate on u/s, pyuria and bacteriuria and u/a prostatomegaly on radiographs and ultrasound hypo-to surgical (delayed) hyperechoic prostate on u/s, bacteriuria and pyuria on u/a prostatomegaly on radiographs and ultrasound, prostatic medical/surgical mineralization on radiographs and ultrasound hypoechoic kidneys on u/s, pyuria on u/a, elevated wbc, medical azotemia pyuria, bacteriuria on u/a medical pyelectasia in abdominal u/s, azotemia surgical (immediate) renomegaly on radiographs, azotemia renal mass on u/s, renomegaly on radiographs surgical (immediate) renal mass on u/s, azotemia, lack of renal blood flow surgical (delayed) on u/s calculi in renal pelvis on radiographs and ultrasound, azotemia medical unless both kidneys affected ureteral calculi on radiographs and ultrasound, hydronephrosis, medical unless both azotemia kidneys affected ureteral calculi on radiographs and ultrasound, hydronephrosis, surgical (delayed until fluid or soft tissue density on u/s, azotemia electrolyte stabilization) diagnosis largely based on physical examination medical unless cannot pass findings urethral catheter azotemia, no peritoneal effusion, lack of urine output or surgical (delayed until outflow with ureteral catheterization, double contrast electrolyte stabilization) cystourethrogram indicated transitional cellular casts on u/a, hematuria, mass effect or surgical and medical thickened irregular urethra on ultrasound or management cystourethrogram hypoechoic swollen testicle on testicular ultrasound surgical (immediate) fluid or gas-filled tubular structure on abdominal ultrasound or surgical (immediate) abdominal radiographs soft tissue tubular structure on radiographs, fluid-filled uterus surgical ( in the event of a negative abdominocentesis, but peritoneal effusion or bile or gastrointestinal perforation are suspected, perform a diagnostic peritoneal lavage. peritoneal dialysis kits are commercially available but are often expensive and impractical (see p. ). animals that have acute abdominal pain can be divided into three broad categories, depending on the primary cause of pain and the initial definitive treatment (table - ) . some diseases warrant a nonsurgical, medical approach to case management. other conditions require immediate surgery following rapid stabilization. other conditions initially can be managed medically until the patient is hemodynamically more stable and then may or may not require surgical intervention at a later time. specific management of each disease entity is listed under its own subheading. box - lists specific indications for exploratory laparotomy. the best means to explore the abdominal cavity accurately and thoroughly is to open the abdomen on midline from the level of the xyphoid process caudally to the pubis for full exposure and then to evaluate all organs in every quadrant in a systematic manner. address specific problems such as gastric or splenic torsion, enteroplication, and foreign body removal, and then copiously lavage the abdomen with warmed sterile saline solution. suction the saline solution thoroughly from the peritoneal cavity so as to not impair macrophage function. in cases of septic peritonitis, the abdomen may be left open, or a drain may be placed for further suction and lavage. the routine use of antibiotics in irrigation solutions is contraindicated because the antibiotics can irritate the peritoneum and delay healing. when the abdominal cavity is left open, secure sterile laparotomy towels and water-impermeable dressings over the abdominal wound with umbilical tape, and then change these daily or as strike-through occurs. open abdomen cases are often effusive and require meticulous evaluation and management of electrolyte imbalances and hypoalbuminemia. the abdomen can be closed and/or the abdominal drain removed when the volume of the effusion decreases, when bacteria are no longer present, and when the neutrophils become more healthy in appearance. bischoff mg: radiographic techniques and interpretation of the acute abdomen, clin tech small anim pract ( ) anaphylactic shock occurs as an immediate hypersensitivity reaction to a variety of inciting stimuli (box - ). in animals, the most naturally occurring anaphylactic reaction results from wasp or bee stings. most other reactions occur as a result of an abnormal sensitivity to items used in making medical diagnoses or treatment. during an anaphylactic reaction, activation of c a and the complement system results in vascular smooth muscle dilation and the release of a cascade of inflammatory mediators, including histamine, slow-reacting substance of anaphylaxis, serotonin, heparin, acetylcholine, and bradykinin. clinical signs associated with anaphylaxis differ between dogs and cats. in dogs, clinical signs may include restlessness, vomiting, diarrhea, hematochezia, circulatory collapse, coma, and death. in cats, clinical signs often are associated with respiratory system abnormalities. clinical signs may include ptyalism, pruritus, vomiting, incoordination, bronchoconstriction, pulmonary edema and hemorrhage, laryngeal edema, collapse, and death. the most important steps to remember in any emergency is to follow the abcs of airway, breathing, and circulation. first, establish an airway through endotracheal intubation or emergency tracheostomy, if necessary. concurrently, an assistant should establish vascular or intraosseous access to administer drugs and fluids (box - ). the patient should be hospitalized until complete resolution of clinical signs. after initial stabilization and treatment, it is important to maintain vascular access and continue intravenous fluid therapy until the patient is no longer hypotensive, and vomiting and diarrhea have resolved. in cases of fulminant pulmonary hemorrhage and edema, administer supplemental oxygen until the patient is no longer hypoxemic or orthopneic on room air. normalize and maintain blood pressure using positive inotropes (dobutamine, - µg/kg/ minute cri) or pressors (dopamine, to µg/kg/minute iv cri; see shock). if bloodtinged vomitus or diarrhea has been observed, administer antibiotics to decrease the risk of bacterial translocation and sepsis (cefoxitin, mg/kg iv tid; metronidazole, mg/kg iv tid). also consider using gastroprotectant drugs (famotidine, . to . mg/kg iv; ranitidine, . to . mg/kg po, iv, im bid; sucralfate, . to . g po tid; omeprazole, . to . mg/kg po sid). a second and less serious form of allergic reaction is manifested as angioneurotic edema and urticaria. in most cases, clinical signs develop within minutes of an inciting allergen. although this type of reaction causes patient discomfort, it rarely poses a life-threatening problem. most animals have mild to severe swelling of the maxilla and periorbital regions. the facial edema also may be accompanied by mild to severe generalized urticaria. some animals may paw at their face, rub at their eyes, or have vomiting or diarrhea. the treatment for angioneurotic edema involves suppressing the immune response by administration of short-acting glucocorticoid drugs and blocking the actions of histamine by the synergistic use of histamine and histamine receptor blockers (box - ). in some cases, the inciting cause is a known recent vaccination or insect sting. many times, however, the inciting cause is not known and is likely an exposure to a stinging insect or arachnid. differential diagnoses for acute facial swelling and/or urticaria include acetaminophen toxicity (cats), anterior caval syndrome, lymphadenitis, vasculitis, hypoalbuminemia, and contact dermatitis. observe animals that have presented for angioneurotic edema for a minimum of to minutes after injection of the short-acting glucocorticoids and antihistamines. monitor blood pressure to make sure that the patient does not have concurrent anaphylaxis and hypotension. after partial or complete resolution of clinical signs, the animal can be discharged to its owner for observation. in dogs, mild vomiting or diarrhea may occur within to days after this type of reaction. wherever possible, exposure to the inciting allergen should be avoided. • administer short-acting glucocorticoid: complications observed while a patient is under anesthesia can be divided into two broad categories: ( ) those related to equipment malfunction or human error and ( ) the patient's physiologic response to the cardiorespiratory effects of the anesthetic drugs. careful observation of the patient and familiarity with anesthetic equipment, drug protocols, and monitoring equipment is necessary for the safest anesthesia to occur. despite this, however, anesthetic-related complications are frequent and need to be recognized and treated appropriately. many anesthetic drugs have a dose-dependent depressive effect on the respiratory system and cause a decrease in respiratory rate and tidal volume, leading to hypoventilation. respiratory rate alone is not a reliable indicator of the patient's oxygenation and ventilatory status. the respiratory tidal volume can be measured with a wright's respirometer. perform pulse oximetry and capnography as noninvasive measures of the patient's oxygenation and ventilation. ventilation can be impaired as a result of anesthetic drugs, patient position, pneumothorax, pleural effusion (chylothorax, hemothorax, pyothorax), equipment malfunction, rebreathing of carbon dioxide, thoracic wall injury, or alveolar fluid (pulmonary edema, hemorrhage, or pneumonia). problems such as a diaphragmatic hernia, gdv, or gravid uterus can impede diaphragmatic excursions once the patient is placed on its back and can lead to impaired ventilation. the work of breathing also may be increased because of increased resistance of the anesthesia circuit and increased dead space ventilation. this is particularly important in small toy breeds. clinical signs of inadequate ventilation and respiratory complications include abnormal respiratory pattern, sudden changes in heart rate, cardiac dysrhythmias, cyanosis, and cardiopulmonary arrest. end-tidal carbon dioxide, or capnography, gives a graphic display of adequacy of ventilation. rapid decreases in end-tidal carbon dioxide can be caused by disconnection or obstruction of the patient's endotracheal tube or poor perfusion, namely, cardiopulmonary arrest (see capnometry [end-tidal carbon dioxide monitoring]). postoperatively, hypoventilation can occur because of the residual effects of the anesthetic drugs, hypothermia, overventilation during intraoperative support, surgical techniques that compromise ventilation (thoracotomy, cervical disk surgery, atlantooccipital stabilization), postoperative bandaging of the abdomen or thorax, ventilatory muscle fatigue, or injury to the cns. cardiac output is a function of heart rate and stroke volume. factors that influence stroke volume include vascular and cardiac preload, cardiac afterload, and cardiac contractility. the patient's cardiac output can be affected adversely by the negative inotropic and chronotropic and vasodilatory effects of anesthetic drugs, all leading to hypotension. emergency care bradycardia, tachycardia, cardiac dysrhythmias, and vascular dilation can lead to hypotension and inadequate organ perfusion. table - lists the normal heart rate and blood pressure in dogs and cats. bradycardia is defined as a heart rate below normal values. many anesthetic drugs can cause bradycardia. causes of bradycardia include the use of narcotics or α -agonist drugs, deep plane of anesthesia, increased vagal tone, hypothermia, and hypoxia. table - lists the causes of bradycardia and the necessary immediate action or treatment. tachycardia is defined as a heart rate above normal values. common causes of tachycardia include vasodilation, drugs, inadequate anesthetic depth and perceived pain, hypercapnia, hypoxemia, hypotension, shock, or hyperthermia. table - lists the causes and immediate action or treatment for tachycardia. hypotension is defined as physiologically low blood pressure (mean arterial pressure less than mm hg). a mean arterial blood pressure less than mm hg can result in inadequate tissue perfusion and oxygen delivery. the coronary arteries are perfused during diastole. inadequate diastolic blood pressure, less than mm hg, can cause decreased coronary artery perfusion and myocardial hypoxemia that can predispose the heart to dysrhythmias. causes of perianesthetic hypotension include peripheral vasodilation by anesthetic drugs, bradycardia or tachyarrhythmias, hypothermia, inadequate cardiac preload from vasodilation or hemorrhage, decreased venous return from patient position or surgical manipulation of viscera, and decreased cardiac contractility. electrocardiogram monitoring is useful for the early detection of cardiac dysrhythmias during the perianesthetic period. clinical signs of cardiac dysrhythmias include irregular pulse rate or pressure, abnormal or irregular heart sounds, pallor, cyanosis, hypotension, and an abnormal ecg tracing. remember that the single best method of detecting cardiac emergency care vagolytic drugs atropine allow time for the drug to wear off. glycopyrrolate allow time for the drug to wear off. sympathomimetic drugs epinephrine allow time for the drug to wear off; administer a β-blocker; turn off infusion. isoproterenol administer a β-blocker. turn off infusion; administer a β-blocker. allow time for drug to wear off. inadequate anesthetic depth increase anesthetic depth. hypercapnia increase ventilation (assisted ventilation). hypoxemia increase gas flow and oxygenation. hypotension decrease anesthetic depth; administer an intravenous crystalloid or colloid bolus, positive inotrope drug, positive chronotrope drug, or pressor. hyperthermia apply ambient or active cooling measures; administer dantrolene sodium if malignant hyperthermia is suspected. hypothermia provide ambient rewarming. hypocalcemia * administer calcium chloride ( mg/kg iv) or calcium gluconate ( mg/kg). decrease vaporizer setting/anesthetic depth. reverse with opioids or a -agonists. vasodilation administer an intravenous crystalloid bolus ( ml/kg). administer an intravenous colloid bolus ( ml/kg). administer a pressor (epinephrine, phenylephrine dysrhythmias is with your fingertips (palpate a pulse or apex heartbeat) and ears (auscultate the heart). confirm the dysrhythmia by auscultating the heart rate and rhythm, identify the p waves and the qrs complexes, and evaluate the relationship between the p waves and qrs complexes. is there a p wave for every qrs, and a qrs for every p wave? during anesthesia, fluid, acid-base, and electrolyte imbalances can predispose the patient to dysrhythmias. sympathetic and parasympathetic stimulation, including the time of intubation, can predispose the patient to dysrhythmias. if the patient's plane of anesthesia is too light, perception of pain can cause catecholamine release, sensitizing the myocardium to ectopic beats. atrioventricular blockade can be induced with the administration of α -agonist medications, including xylazine and medetomidine. thiobarbiturates (thiopental) can induce ventricular ectopy and bigeminy. although these dysrhythmias may not be harmful in the awake patient, anesthetized patients are at a particular risk of dysrhythmia-induced hypotension. carefully monitor and treat all dysrhythmias (see cardiac dysrhythmias). box - lists steps to take to prevent perianesthetic dysrhythmias. awakening during anesthesia can occur and can be caused by equipment failure and simply, although no one likes to admit it, human error. table - lists causes of arousal during anesthesia and appropriate immediate actions. awaken patient, and administer dantrolene arousal (e.g., malignant hyperthermia) sodium. • stabilize acid-base and electrolyte balance before anesthetic induction, whenever possible. • rehydrate patient before anesthetic induction. • select anesthetic agents appropriate for the particular patient. • be aware of the effects of the drugs on the myocardium. • ensure adequate anesthetic depth and oxygenation before anesthetic induction. • ensure ventilatory support during anesthesia. • monitor heart rate, rhythm, blood pressure, pulse oximetry, and capnometry during anesthesia. • ensure adequate anesthetic depth before surgical stimulation. • avoid surgical manipulation to the heart or great vessels, whenever possible. • avoid changes in perianesthetic depth. • avoid hypothermia. delayed recovery can be caused by a number of factors, including excessive anesthetic depth, hypothermia, residual action of narcotics or tranquilizers, delayed metabolism of anesthetic drugs, hypoglycemia, hypocalcemia, hemorrhage, and breed or animal predisposition. careful monitoring of the patient's blood pressure, acid-base and electrolyte status, anesthetic depth, pcv, and vascular volume intraoperatively and taking care with supportive measures to prevent abnormalities can hasten anesthetic recovery and avoid postoperative complications. gaynor the presentation of a patient with a bleeding disorder often is a diagnostic challenge for the veterinary practitioner (boxes - and - ). in general, abnormal bleeding can be caused by five major categories: ( ) vascular trauma, ( ) circulating inhibitors of coagulation heparin fibrin degradation products development of spontaneous deep hematomas, unusually prolonged bleeding after traumatic injury, bleeding at multiple sites throughout the body involving multiple organ systems, delayed onset of severe hemorrhage after bleeding, and an inability on the practitioner's part to find an organic cause of bleeding. the signalment, history, clinical signs, and results of coagulation often can aid in making a rapid diagnosis of the primary cause of the disorder and in the selection of appropriate case management. when taking a history, ask the following important questions: • what is the nature of the bleeding? • what sites are affected? • how long has the bleeding been going on? • has your animal had any previous or similar episodes? • is there any possibility of any toxin exposure? • if so, when and how much did your animal consume? • is there any possibility of trauma? • does your animal run loose outdoors unattended? • have you ever traveled, and if so, where? • has your animal been on any medications recently or currently? • has your animal been vaccinated recently? • have any known relatives of your animal had any bleeding disorders? • are there any other abnormal signs that you have seen? abnormalities found on physical examination may aid in determining whether the hemorrhage is localized or generalized (i.e., bleeding from a venipuncture site versus bleeding diathesis). note whether the clinical signs are associated with a platelet problem and superficial hemorrhage or whether deep bleeding can be associated with abnormalities of the coagulation cascade. also, make an attempt to identify any concurrent illness that can predispose the patient to a bleeding disorder (i.e., pancreatitis, snakebite, sepsis, immunemediated hemolytic anemia, or severe trauma and crush or burn injury). abnormalities associated with coagulopathies include petechiae and ecchymoses, epistaxis, gingival bleeding, hematuria, hemarthrosis, melena, and hemorrhagic cavity (pleural and peritoneal or retroperitoneal) effusions. disseminated intravascular coagulation is a complex syndrome that results from the inappropriate activation of the clotting cascade, leading to disruption of the normal balance between thrombosis and fibrinolysis. the formation of diffuse microthrombi with concurrent consumption of platelets and activated clotting factors leads to end-organ thrombosis with various degrees of clinical hemorrhage. in animals, dic always results from some other pathologic process, including various forms of neoplasia, crush and heat-induced injury, sepsis, inflammation, and immune-mediated disorders (box - ). the pathophysiologic mechanisms involved in dic include vascular endothelial damage, activation and consumption of platelets, release of tissue procoagulants, and consumption of endogenous anticoagulants. because dic always results from some other disease process, diagnosis of dic is based on a number of criteria when evaluating various coagulation tests, peripheral blood smears, platelet count, and end products of thrombosis and fibrinolysis. there is no one definitive criterion for the diagnosis of dic (box - ). thrombocytopenia occurs as platelets are consumed during thrombosis. it is important to remember that trends in decline in platelet numbers are just as important as thrombocytopenia when making the diagnosis. in some cases the platelet count still may be within the normal reference range but has significantly decreased in the last hours. early in dic the procoagulant cascade dominates, with hypercoagulability. activated clotting time, aptt, and pt may be rapid and shorter than normal. in most cases, we do not recognize the hypercoagulable state in our critically ill patients. later in dic, as platelets and activated clotting factors become consumed, the act, aptt, and pt become prolonged. antithrombin, a natural anticoagulant, also becomes consumed, and antithrombin levels decline. antithrombin levels can be measured at commercial laboratories and in some large veterinary institutions. the end products of thrombosis and subsequent fibrinolysis also can be measured. fibrinogen levels may decline, although this test is not sensitive or specific for dic. fibrin degradation (split) products also become elevated. fibrin degradation products are normally cleared by the liver, and these also become elevated in cases of hepatic failure because of lack of clearance. more recently, cageside d-dimer tests have become available to measure the breakdown product of cross-linked fibrin as a more sensitive and specific monitor of dic. management of dic first involves treating the primary underlying cause. by the time dic becomes evident, rapid and aggressive treatment is necessary. if you are suspicious of dic in any patient with a disease known to incite dic, then ideally, you should begin treatment before the hemostatic abnormalities start to occur for the best possible prognosis. treatment involves replacement of clotting factors and antithrombin and prevention of further clot formation. to replenish clotting factors and antithrombin, administer fresh whole blood or fresh frozen plasma. heparin requires antithrombin as a cofactor to inactivate thrombin and other activated coagulation factors. administer heparin ( to units/kg sq q - h of unfractionated heparin; or fractionated enoxaparin [lovenox], mg/kg sq bid). aspirin ( mg/kg po bid in dogs; every third day in cats) also can be administered to prevent platelet adhesion. management of dic also involves the rule of twenty monitoring and case management to maintain end-organ perfusion and oxygen delivery (see the rule of ). hemophilia a is a sex-liked recessive trait that is carried by females and manifested in males. female hemophiliacs can occur when a hemophiliac male is bred with a carrier female. hemophilia a has been reported in cats and a number of dog breeds, including miniature schnauzer, saint bernard, miniature poodle, shetland sheepdog, english and irish setters, labrador retriever, german shepherd, collie, weimaraner, greyhound, chihuahua, english bulldog, samoyed, and vizsla. mild to moderate internal or external bleeding can occur. clinical signs of umbilical cord bleeding can become apparent in some animals shortly after weaning. gingival hemorrhage, hemarthrosis, gastrointestinal hemorrhage, and hematomas may occur. clotting profiles in animals with factor viii deficiency include prolonged aptt and act. the pt and buccal mucosa bleeding time are normal. affected animals have low factor viii activity but normal to high levels of factor viii-related antigen. carrier females can be detected by low ( % to % of normal) factor viii activity and normal to elevated levels of factor vii-related antigen. von willebrand's disease is a deficiency or defect in von willebrand's protein. a number of variants of the disease have been described: von willebrand's disease type i is associated with a defect in factor viir/protein concentration, and von willebrand's disease type ii is associated with a defect in viiir:vwf. type i von willebrand's disease is most common in veterinary medicine. von willebrand's disease has been identified in more than breeds of dogs, with an incidence that varies from % to % depending on the breed of origin. affected breeds include doberman pinchers, german shepherd dogs, scottish terriers and standard manchester terriers, golden retrievers, chesapeake bay retrievers, miniature schnauzers, and pembroke welsh corgis. two forms of genetic expression occur: ( ) autosomal recessive disease in which homozygous von willebrand's disease individuals have a bleeding disorder, whereas heterozygous individuals carry the trait but are clinically normal. the second variant of genetic expression involves an autosomal dominant disease with incomplete expression such that heterozygous individuals are affected carriers and homozygous individuals are severely affected. von willebrand's disease has high morbidity, but fortunately a low mortality. dogs with % or less than normal vwf tend to hemorrhage. platelet counts are normal, but bleeding times can be prolonged. the aptt can be slightly prolonged when factor viii is less than % of normal. routine screening tests are nondiagnostic for this disease, although in a predisposed breed with a normal platelet count, a prolonged buccal mucosa bleeding time strongly supports a diagnosis of von willebrand's disease. documentation of clinical bleeding with low or undetectable levels of factor viii antigen or platelet-related activities of vwf support a diagnosis of von willebrand's disease. recessive animals have zero vwf:antigen (a subunit of factor iii); heterozygotes have % to % of normal. in the incompletely dominant form, levels of vwf antigen are reduced (less than % to %). clinical signs in affected animals include epistaxis, hematuria, diarrhea with melena, penile bleeding, lameness, hemarthrosis, hematoma formation, and excessive bleeding with routine procedures such as nail trimming, ear cropping, tail docking, surgical procedures (spay, neuter), and lacerations. estrous and postpartum bleeding may be prolonged. a dna test to detect carriers of the vwf gene is available through vetgen (ann arbor, michigan) and michigan state university. patients with von willebrand's disease should avoid drugs known to affect platelet function adversely (sulfonamide, ampicillin, chloramphenicol, antihistamines, theophylline, phenothiazine tranquilizers, heparin, and estrogen). hemophilia b is an x-linked recessive trait that occurs with less frequency that hemophilia a. the disease has been reported in scottish terriers, shetland and old english sheepdogs, saint bernards, cocker spaniels, alaskan malamutes, labrador retrievers, bichon frises, airdale terriers, and british shorthair cats. carrier females have low ( % to % of normal) factor ix activity. clinical signs are more severe than for hemophilia a. congenital deficiencies of factor vii have been reported as an autosomal, incompletely dominant characteristic in beagles. heterozygotes have % factor vii deficiency. bleeding tends to be mild. the pt is prolonged in affected individuals. factor x deficiency has been documented in cocker spaniels and resembles fading-puppy syndrome in newborn dogs. internal or umbilical bleeding can occur, and affected dogs typically die. bleeding may be mild in adult dogs. in severe cases, factor x levels are reduced to % of normal; in mild cases, factor x levels are % to % of normal. factor xii deficiency has been documented as an inherited autosomal recessive trait in domestic cats. heterozygotes can be detected because they have a partial deficiency ( % of normal) of factor xii. homozygote cats have less than % factor xii activity. deficiency of hageman factor usually does not result in bleeding or other disorders. factor xi deficiency is an autosomal disease that has been documented in kerry blue terriers, great pyrenees, and english springer spaniels. in affected individuals, protracted bleeding may be observed. homozygotes have low factor xi activity (< % of normal), and heterozygotes have % to % of normal. the management of congenital defects of hemostasis typically involves replenishing the clotting factor that is present. usually, this can be accomplished in the form of fresh frozen plasma transfusion ( ml/kg). if anemia is present because of severe hemorrhage, fresh whole blood or packed rbcs also can be administered. recent research has investigated the use of recombinant gene therapy in the treatment of specific factor deficiencies in dogs; however, the therapy is not yet available for use in clinical practice. in cases of von willebrand's disease, administration of fresh frozen plasma ( to ml/kg) or cryoprecipitate ( unit/ kg body mass) provides vwf, factor viii, and fibrinogen. doses can be repeated until hemorrhage ceases. -desamino- -d-arginine vasopressin (ddavp) also can be administered ( µg/kg sc or iv diluted in . % saline given over to minutes) to the donor and patient to increase the release of stored vwf from endothelial cells. a fresh whole blood transfusion can be obtained from the donor and immediately administered to the patient, or spun down and the fresh plasma administered if rbcs are not needed. administer a dose of ddavp to any affected dog before initiating any elective surgical procedures. a supply of fresh frozen plasma and rbcs should be on hand, should uncontrolled hemorrhage occur. platelets are essential to normal blood coagulation. after a vessel is damaged, release of vasoactive amines causes vasoconstriction and sluggish flow of blood in an attempt to squelch hemorrhage. platelets become activated by platelet activating factor, and attach to the damaged vascular endothelium. normal platelet adhesion depends on mediators such as calcium, fibrinogen, vwf:antigen, and a portion of factor viii. after adhesion, the platelets undergo primary aggregation and release a variety of chemical mediators including adenosine diphosphate, prostaglandins, serotonin, epinephrine, thromboplastin, and thromboxane a that promote secondary aggregation and contraction. platelet abnormalities can include decreased platelet production (thrombocytopenia), decreased platelet function (thrombocytopathia), increased platelet destruction, increased platelet consumption, and platelet sequestration. thrombocytopathia refers to platelet function abnormalities. alterations in platelet function can affect platelet adhesion, aggregation, or release of vasoactive substances that help form a stable clot (box - ). in von willebrand's disease there is a deficiency in vwf:antigen that results in altered platelet adhesion. vascular purpuras are reported and have been seen in collagen abnormalities such as ehlers-danlos syndrome, which can be inherited as an autosomal dominant trait with complete penetrance and has been recognized in german shepherd dogs, dachshunds, saint bernards, and labrador retrievers. thrombasthenic thrombopathia is a hereditary autosomal dominant abnormality that has been described in otterhounds, foxhounds and scottish terriers. in this condition, platelets do not aggregate normally in response to adenosine diphosphate and thrombin stimulation. evaluation of platelet function is based on a total platelet count, buccal mucosa bleeding time, and thromboelastography. platelet function defects (thrombocytopenia and thrombocytopathia) can affect both sexes. clinical signs can resemble von willebrand's disease. in most cases, buccal mucosa bleeding time will be prolonged, but platelet count and clotting tests will be normal. platelet count can be decreased because of problems with production, increased consumption, sequestration, or destruction. causes of accelerated platelet destruction are typically immune-mediated autoantibodies, drug antibodies, infection, and isoimmune destruction. consumption and sequestration usually are caused by dic, vasculitis, microangiopathic hemolytic anemia, severe vascular injury, hemolytic uremic syndrome, and gram-negative septicemia. primary thrombocytopenia with no known cause has been called idiopathic thrombocytic purpura. in approximately % of the cases, thrombocytopenia is associated with immune-mediated destruction caused by immune-mediated hemolytic anemia, systemic lupus erythematosus, rheumatoid arthritis, dic, and diseases that affect the bone marrow. in systemic lupus erythematosus, % to % of the affected dogs have concurrent idiopathic thrombocytic purpura. when immune-mediated hemolytic anemia and idiopathic thrombocytic purpura are present in the same patient, the disease is called evans syndrome. pf- is a non-complement-fixing antibody that is produced in the spleen and affects peripheral and bone marrow platelets and megakaryocytes. antibodies directed against platelets are usually of the igg subtype in animals. antiplatelet antibodies can be measured by a pf- release test. platelet counts with immune-mediated destruction typically are less than , platelets/µl. infectious causes of thrombocytopenia include ehrlichia canis, anaplasma phagocytophilum (formerly, ehrlichia equi), and rickettsia rickettsii (rocky mountain spotted fever). primary immune-mediated thrombocytopenia has an unknown cause and most frequently is seen in middle-to older-aged female dogs. breed predispositions include cocker spaniels, german shepherd dogs, poodles (toy, miniature, standard), and old english sheepdogs. thrombocytopenia usually is manifested as petechiae, ecchymoses of skin and mucous membranes, hyphema, gingival and conjunctival bleeding, hematuria, melena, and epistaxis. to make a diagnosis of idiopathic thrombocytic purpura, measure the severity of thrombocytopenia (< , platelets/µl), analyze the peripheral blood smear for evidence of platelet fragmentation or microthrombocytosis, normal to increased numbers of megakaryocytes in the bone marrow, detection of antiplatelet antibody, increased platelet counts after starting glucocorticoid therapy, and elimination of other causes of thrombocytopenia. if tick-borne illnesses are suspected, antibody titers for e. canis, a. phagocytophilum (formerly e. equi), and r. rickettsii should be performed. treatment of immune-mediated thrombocytopenia involves suppression of the immune system to stop the immune-mediated destruction and to stimulate platelet release from the bone marrow. traditionally, the gold standard to suppress the immune system is to use glucocorticoids (prednisone or prednisolone, to mg/kg po bid divided, or dexamethasone, . to . mg/kg iv or po q h). more recently human serum immunoglobulin (igg) also has been used ( . to . g/kg iv in saline over hours; pretreat with mg/kg diphenhydramine minutes before starting infusion). vincristine ( . mg/m iv once) can stimulate the release of platelets from the bone marrow if megakaryocytic precursors are present; however, the platelets released may be immature and potentially nonfunctional. treatment with fresh whole blood or packed rbcs is appropriate if anemia is present; however, unless specific platelet-rich plasma has been purchased from a blood bank, fresh whole blood contains relatively few platelets, which are shortlived ( hours) and will not effectively raise the platelet count at all. finally, long-term therapy is usually in the form of azathioprine ( mg/kg po once daily, tapered to mg/kg daily to every other day after week) and cyclosporine ( to mg/kg po divided). if a tickborne illness is suspected, administer doxycycline ( to mg/kg po bid) for weeks or if titers come back negative. thrombocytopenia also can occur in the cat. causes for thrombocytopenia in cats include infections ( %), neoplasia ( %), cardiac disease ( %), primary immune-mediated disease ( %), and unknown causes ( %). in one study of cats with feline leukemia and myeloproliferative disease, % of cases had thrombocytopenia. warfarin and coumarin derivatives are the major class of rodenticides used in the united states. vitamin k antagonist rodenticides inhibit the epoxidase reaction and deplete active vitamin k, causing a depletion of vitamin k-dependent coagulation factors (ii, vii, ix, x) within hours to week of ingestion, depending on the ingested dose. affected animals can spontaneously hemorrhage anywhere in the body. clinical signs can include hemoptysis, respiratory difficulty, cough, gingival bleeding, epistaxis, hematuria, hyphema, conjunctival bleeding, petechiae and ecchymoses, cavity hemorrhage (pleural, peritoneal, retroperitoneal) with acute weakness, lethargy or collapse, hemarthrosis with lameness, deep muscle bleeds, and intracranial or spinal cord hemorrhage. diagnosis of vitamin k antagonism includes prolonged pt. a pivka (protein induced by vitamin k absence or antagonism) test also can be performed, if possible. treatment of vitamin k antagonist rodenticide intoxication and other causes of vitamin k deficiency involves supplementation with vitamin k (phytonadione, mg/kg sq once with -gauge needle in multiple sites, and then . mg/kg po bid to tid for days). never administer injections of vitamin k intramuscularly, because of the risk of causing deep muscle hematomas, or intravenously, because of the risk of anaphylaxis. the pt should be rechecked days after the last vitamin k capsule is administered, for some of the secondgeneration warfarin derivates are fat-soluble, and treatment may be required for an additional weeks. act, activated clotting time; aptt, activated partial thromboplastin time; bmbt, buccal mucosa bleeding time; fdp, fibrin degradation products; n, normal; pt, prothrombin time. thermal burns are fortunately a relatively infrequent occurrence in veterinary patients. box - lists various causes of malicious and accidental burns. the location of the burn is also important in assessing its severity and potential to lose function. burns on the perineum, feet, face, and ears are considered to be the most severe because of loss of function and severe pain. often the severity of thermal injury is difficult to assess in animals because hair coat potentially can mask clinical signs and because the thermal injury can continue after the animal has been removed from the heat source. the skin cools slowly and warms slowly, considerations that become important when initiating therapy for burns. the severity of thermal injury is associated with the temperature to which the animal is exposed, the duration of contact, and the ability of the tissue to dissipate heat. the tissue closest to the heat source undergoes necrosis and has decreased blood flow. the severity of thermal burn injury is associated directly with the temperature to which the animal is exposed, the percentage of total body surface area affected, the thickness of injured tissue, and whether underlying complications with other body systems occur. prognosis largely depends on the total body surface area affected (table - ) . superficial partial thickness, or first-degree, burns offer the most favorable prognosis. the affected epidermis initially appears erythematous and then quickly desquamates within to days. in most cases, fur grows back without leaving a scar. deep partial thickness, or second-degree, burns involve the epidermis and dermis and are associated with subcutaneous edema, inflammation, and pain. deep partial thickness burns heal from deeper adnexal tissues and from the wound edges and are associated with an increased chance of scarring and depigmentation. the most severe type is known as full thickness, or third-degree, burns, in which thermal injury destroys the entire thickness of the skin and forms an eschar. thrombosis of superficial and deeper skin vasculature and gangrene occurs. treatment involves sequential wound debridement. healing occurs by second intention and reepithelialization or by wound reconstruction. in most cases, scarring is extensive in affected areas. burns greater than % of total body surface area will have systemic effects, including impaired cardiovascular function, pulmonary dysfunction, and impaired immune function. burned tissue, with capillary damage, has increased permeability. the release of inflammatory cytokines, oxygen-derived free radical species, prostaglandins, leukotrienes, emergency care histamine, serotonin, and kinins results in increased vascular permeability and leakage of plasma proteins into the interstitium and extravascular space. at the time of presentation, first examine the patient and ascertain whether airway obstruction, impaired ventilatory function, circulatory shock, or pain are present. if necessary, establish an airway with endotracheal intubation or emergency tracheostomy. next, cool the burned area(s) with topical cool water. use care to avoid overcooling and iatrogenic hypothermia. the best approach is to cool only one portion of the patient's body at a time, then dry, and repeat the process for all affected areas to avoid overcooling and iatrogenic hypothermia. establish vascular access and administer appropriate and judicious analgesic drugs and intravenous fluid therapy. whenever possible, avoid placing a catheter through an area of burned or damaged skin. in the early stages of burn injury, shock doses of intravenous crystalloid fluids usually are not required. later, however, as severe tissue exudation occurs, protein and fluid losses can become extensive, requiring aggressive crystalloid and colloid support to treat hypovolemia and hypoproteinemia. flush the eyes with sterile saline and examine behind the third eyelids for any particulate matter. stain the corneas to make sure that superficial corneal burns are not present. treat superficial corneal burns with triple antibiotic ophthalmic ointment. next, assess the total body surface area affected, as this will gauge prognosis. depending on the extent of the damage, decide whether the burn is superficial and local therapy is indicated or whether more severe injuries exist that may involve systemic therapy or possibly euthanasia. in most cases the diagnoses of thermal burns are based on a clinical history of being in a house fire, clothes dryer, or under a heating lamp. too frequently, however, thermal burns become apparent days after an elective surgical procedure in which the patient was placed on a faulty heating pad rather than a circulating warm water or warm air blanket. superficial burns appear as singed fur with desquamating, easily epilated hair. this condition also can resemble a superficial or deeper dermatophytosis if history is unknown. other differential diagnoses include immune-mediated vasculitis or erythema multiforme. unless the superficial dermis is blistered, it may be difficult to distinguish between a thermal burn, chemical burn, or electrical burn if the trauma went unnoticed. management of burn injury largely depends on the depth of injury and the total body surface area affected. partial thickness burns and those affecting less than % of the total body surface area will require support in the form of antibiotic ointment and systemic analgesic drugs. burns affecting greater than % of total body surface area or deep thickness burns require more aggressive therapy. central venous catheters can be placed to administer crystalloid and colloid fluids, parenteral nutrition if necessary, antibiotics, and analgesic drugs. monitor perfusion parameters closely, including heart rate, blood pressure, capillary refill time, and urine output. respiratory function can be impaired because of concurrent smoke inhalation, thermal damage to the upper airways and alveoli, and carboxyhemoglobin or methemoglobin intoxication. respiratory function also can be impaired because of burn injury to the skin around the thoracic cage. thoracic radiographs may reveal patchy interstitial to alveolar infiltrates associated with pulmonary edema, pneumonia, and atelectasis. bronchoscopy often reveals edema, inflammation, particulate matter, and ulceration of the tracheobronchial tree. in some cases, upper airway inflammation is so severe that an emergency tracheostomy must be performed to treat airway obstruction. administer supplemental humidified oxygen at to ml/kg/minute via endotracheal tube, tracheostomy, nasal or intratracheal tube, or hood oxygen if respiratory function and hypoxemia are present. perform blood work including a hematocrit, albumin, bun, creatinine, and glucose at the time of presentation. monitor serum electrolytes, albumin, and colloid oncotic pressure closely because derangements can be severe as burns become exudative. the goal of fluid therapy in the burn patient is to establish and maintain intravascular and interstitial fluid volume, normalize electrolyte and acid-base status, and maintain serum albumin and oncotic pressure. in the first hours following burn injury, direct fluid therapy to maintaining the patient's metabolic fluid requirements. crystalloid fluids in the form of normosol-r, plasmalyte-m, or lactated ringer's solution can be administered according to the patient's electrolyte and acid-base status (see fluid therapy). monitor urine output, and keep it at to ml/kg/hour. avoid overhydration in the early stages of burn injury. in affected burn patients, calculate the amount of fluid that should be administered over a -hour period from the formula − ml/kg × percent total body surface area. administer half of this calculated dose over the first hours and then the remaining half over the next hours. in cats, administer only % to % of this calculated volume. to administer this volume and also avoid fluid overload is often difficult in critically ill patients with pulmonary involvement associated with smoke inhalation injury. avoid colloids in the first hours after burn injury. monitor the patient closely for serous nasal discharge, chemosis, and rales that may signify pulmonary edema. as burns become exudative, weigh the patient at least twice daily. infused fluid should equal fluid output in the form of urine and wound exudates. acute weight loss signifies acute fluid loss and that crystalloid fluid infusion should be more aggressive. ideally, keep the patient's serum albumin equal to or greater than . g/dl and total protein between . and . g/dl using a combination of fresh frozen plasma or concentrated human albumin. adjunct colloidal support can be provided with synthetic colloids including hetastarch or hbocs. keep serum potassium within . to . meq/l using potassium chloride or potassium phosphate supplementation. if potassium supplementation exceeds to meq/l and the patient continues to have severe refractory hypokalemia, administer magnesium chloride ( . meq/kg/day) to enhance potassium retention. if anemia occurs, administer packed rbcs or whole blood (see blood component therapy). lavage wounds daily with lactated ringer's solution or . % sodium chloride solution. place wet-to-dry bandages or bandages soaked in silver sulfadiazine or nitrofurazone ointment over the wounds. depending on the thickness of the burn, epilation and eschar formation and separation may take to days. at each bandage change, debride devitalized tissue to normal tissue. perform staged partial or total escharectomy, and leave the wound to heal by second intention or by reconstruction using skin advancement flaps or grafts. maintain meticulous sterility at all times, given that burn patients are at high risk for infection. administer broad-spectrum antibiotics including cefazolin and enrofloxacin. perform wound culture if a resistant bacterial infection is suspected. the most common cause of electrical injury is associated with an animal chewing on low-voltage alternating current electrical cords in the household. damage is caused by the current flowing through the path of least resistance, causing heat and thrombosis of vessels and neurons. in some cases, the owner witnesses the event. in other cases, the owner presents the patient because of vague nonspecific signs, and characteristic abnormalities on physical examination support a diagnosis of electrocution. burns on the face, paws, commissures of the mouth, tongue, and soft palate may be present. electrocution causes a massive release of catecholamines and can predispose the patient to noncardiogenic pulmonary edema within hours of the incident. clinical signs may be isolated to the pulmonary system, including orthopnea, pulmonary crackles, and cyanosis. assess the patient's lips, tongue, soft palate, gingivae, and commissures of the mouth. early after electrocution, the wound may appear small and white, black, or yellow. later, the wound may become larger as tissue sloughs because of damaged vascular supply. assess the patient's respiratory status. auscultate the lungs to determine whether pulmonary crackles emergency care are present. if the patient is stable, thoracic radiographs may demonstrate an interstitial to alveolar lung pattern in the dorsocaudal lung fields. measure the patient's heart rate, blood pressure, oxygenation as determined by pulse oximetry or arterial blood gas and urine output. immediate treatment consists of judicious use of analgesics for the burn injury, antibiotics (cefazolin, mg/kg q h; cephalexin, mg/kg q h), and humidified supplemental oxygen ( to ml/kg/minute). direct fluid therapy at providing the patient's metabolic fluid requirements. because of the risk of development of noncardiogenic pulmonary edema, avoid overzealous administration of crystalloid fluids. differential diagnoses for the patient with electrical burn injury and electrocution include chemical or thermal burn, immune-mediated glossitis, cardiogenic pulmonary edema, and pneumonia. management of the patient with electrical burn injury and electrocution primarily involves the administration of analgesic agents, supplemental humidified oxygen, and topical treatment of electrical burns. the noncardiogenic pulmonary edema is typically unresponsive to diuretics (i.e., furosemide), bronchodilators (i.e., aminophylline), and splanchnic vascular dilators (i.e., low-dose morphine). the use of glucocorticoids has no proven benefit and may impair respiratory immune function and is therefore contraindicated. oral burns may require debridement and advancement flaps if large defects or oronasal fistulas develop. if oral injury is severe, place an esophagostomy or percutaneous gastrostomy tube to ensure adequate nutrition during the healing process. if an animal survives the initial electrocution, prognosis is generally favorable with aggressive supportive care. chemical burns are associated with a number of inciting causes, including oxidizing agents, reducing agents, corrosive chemicals, protoplasmic poisons, desiccants, and vesicants. the treatment for chemical burns differs slightly from that for thermal burns, so it remains important to investigate the cause of the burn when providing initial treatment, whenever possible. at the scene, advise the owner to wrap the patient in a clean towel for transport. chilling can be avoided by then wrapping the patient in a second or third blanket. placement of ointments by well-doers should be avoided. encourage immediate transport to the nearest triage facility. the first and foremost consideration when treating a patient with chemical burn is to remove the animal from the inciting cause or offending agent. make no attempt to neutralize alkaline or acid substances because the procedure potentially could cause an exothermic reaction, leading to thermal injury in addition to the chemical injury. remove collars or leashes that may act as tourniquets or constricting devices. flush affected areas with copious amounts of cool water for several minutes, not cooling more than % to % of the body at any one time to prevent iatrogenic hypothermia. support breathing by extending the patient's head and neck. carefully clip the fur over affected areas for further evaluation of the extent of the injury. lavage exposed eyes with sterile saline, and stain the cornea to evaluate for any corneal burns. debride any wounds carefully, knowing that the full extent of the wound may not manifest itself for several days. then cover the wounds with antibiotic burn ointment such as silver sulfadiazine and an occlusive dressing. without a history of exposure, the differential diagnosis for any chemical burn includes thermal burn, necrotizing vasculitis, erythema multiforme, or superficial or deep pyoderma. contact local or national animal poison control regarding whether to attempt neutralization. perform daily bandage changes with staged debridement as the full extent of the wound manifests itself. place antimicrobial ointment and silver sulfadiazine ointment over the wound to prevent infection. the routine use of antibiotics may promote the development of a resistant bacterial infection. first-generation cephalosporin can be administered. if a more serious infection develops, perform culture and susceptibility testing to direct appropriate antibiotic therapy. the wound can heal by second intention or may require reconstructive repair for definitive closure. the primary cause of radiation injury in small animal patients is radiation therapy for neoplastic conditions. the goal of radiation therapy is to kill neoplastic cells. an unfortunate side effect is damage to adjacent normal tissue that results in necrosis, fibrosis, and impaired circulation to the affected area. radiation burns result in dermatitis, mucositis, impaired surgical wound healing, and chronic nonhealing wounds. in many cases, the degree of secondary radiation injury to normal tissue can be prevented or decreased with careful radiation planning and mapping of the radiation field, such that radiation exposure to normal tissue is limited to the smallest extent possible. with the advent of three-dimensional imaging modalities such as computed tomography (ct) and magnetic resonance imaging (mri), this has become more routine in veterinary oncology to date. radiation injury can be early and appear at the later stage of the course of radiation therapy. late effects can be delayed and occur months to years after treatment. the degree of radiation injury is categorized based on the depth of tissue affected. first-degree changes cause cutaneous erythema. second-degree changes cause superficial desquamation. thirddegree changes cause deeper moist desquamation, and fourth-degree changes are associated with complete dermal destruction and ulceration. during the early stages of radiation injury, affected tissues may appear erythematous and edematous. wound exudates may be moist, or the skin may appear dry and scaly with desquamation or ulceration. later, the area may scar and depigment or may have induration, atrophy, telangiectasia, keratosis, and decreased adnexal structures. treatment for radiation dermatitis is to irrigate the area with warmed saline and to protect the area from self-mutilation. no-bite, or elizabethan, collars or loose clothing can be used to protect the area for patient-induced injury. mucositis can be treated with topical green tea baths and the administration of an oral solution of l-glutamine powder ( g/m ). local irrigation of xylocaine or lidocaine viscous jelly can be used in dogs but should be avoided in cats because of the risk of inducing hemolytic anemia and neurotoxicity. topical and systemic antibiotics (cephalexin, mg/kg po tid) also can be administered. avoid antibiotics that can be sensitized by radiation (i.e., metronidazole). because most radiation burns are associated with a known exposure to radiation therapy, the cause of the patient's injury usually is known. if an animal presents to you with a scar, however, differential diagnoses may include nasal planum solar dermatitis, pemphigus foliaceus, discoid lupus, superficial necrolytic dermatitis, superficial or deep pyoderma, chemical burn, or thermal burn. treatment of radiation injury involves making the patient as comfortable as possible with analgesic drugs, prevention of self-mutilation, and staged debridement techniques. wounds can heal by second intention or may require reconstructive surgery. distress syndrome (ards), and anesthetic agents. the acute onset of bradycardia, change in mucous membrane color and capillary refill time, change in respiratory pattern, and change in mentation are signs of possible deterioration and impending cardiopulmonary arrest. the diagnosis of cardiopulmonary arrest is based on the absence of effective ventilation, severe cyanosis, absence of a palpable pulse or apex heartbeat, absence of heart sounds, and ecg evidence of asystole or other nonperfusing rhythm such as electricalmechanical dissociation (aka pulseless electrical activity) or ventricular fibrillation. the goals of cpcr are to obtain airway access, provide artificial ventilation and supplemental oxygen, implement cardiac compressions and cardiovascular support, recognize and treat dysrhythmias and arrhythmias, and provide stabilization and treatment for cardiovascular, pulmonary, and cerebral function in the event of a successful resuscitation. even with aggressive treatment and management, the overall success of cpcr is less than % in critically ill or traumatized patients and % to % in anesthetized patients. basic life support involves rapid intubation to gain airway access, artificial ventilation, and cardiac compressions to promote blood flow and delivery of oxygen to the brain and other important tissues (figure - ). perform the abcs or cabs of cpcr, where a is airway, b is breathing, and c is compression and circulation. recently, the paradigm has shifted to cabs. while a team member is grabbing an endotracheal tube, clearing the airway of foreign debris, and establishing airway access through endotracheal intubation, a second person starts external cardiac compressions to deliver oxygen that is in the bloodstream to the vital organs. the patient should be positioned in dorsal (> kg) or lateral (< kg) recumbency for external cardiac compressions. approximately to external compressions should be performed over the patient's sternum. a team member should palpate for a peripheral pulse to determine whether cardiac compressions are actually effective. if a peripheral pulse cannot be palpated for every chest compression, change the patient's position and have a larger individual perform compressions, or initiate open-chest cardiac resuscitation. once the patient is intubated, tie in the endotracheal tube and attach it to an oxygen source (anesthetic machine or mechanical ventilator or ambu bag) for artificial ventilation. the oxygen flow rate should be ml/kg/minute. give two long breaths, and then to breaths per minute. simultaneous ventilation with thoracic compression increases the pressure difference in the thorax and allows more forward flow of oxygenated blood through the great vessels into the periphery. if possible, a third team member can initiate interposed abdominal compressions, compressing the abdomen when the thoracic cage is relaxed, to improve forward flow. if only one person is available to perform the thoracic compressions and ventilation, give two breaths for every compressions (i.e., thoracic compressions followed by two long breaths, and then start thoracic compressions again). the jen chung maneuver can be performed by placing a -to -gauge hypodermic needle through the skin of the nasal philtrum and twisting the needle into the periosteum to stimulate respirations. this maneuver appears to work better in cats than dogs at return to spontaneous respiration. advanced life support during cpcr involves ecg, pulse oximetry and capnometry monitoring, administration of drugs, and the administration of intravenous fluids (in select cases). most of the drugs used during cpcr can be administered directly into the lungs from the endotracheal tube (intratracheal tube). therefore, only in select instances is it necessary to establish vascular or intraosseous access during cpcr (figure - ) . if an animal experiences cardiopulmonary arrest because of extreme hemorrhage or hypovolemia, inappropriate vasodilation caused by sepsis or systemic inflammation, or vasodilation resulting from anesthesia, the administration of shock volumes ( ml/kg/hour in dogs and ml/kg/hour in cats) is appropriate. if a patient is euvolemic and experiences cardiopulmonary arrest, however, an increase in circulating fluid volume actually can impair coronary artery perfusion by increasing diastolic arterial blood pressure and is asystole is one of the most common rhythm disturbances that causes cardiac arrest in small animal patients. one of the most important things to do when the ecg looks like asystole is to make sure that the ecg monitor is working properly and that all ecg leads are attached properly to the patient. if asystole is truly present, reverse any opiate, α -agonist, or benzodiazepine drugs with their appropriate reversal agents. lowdose epinephrine ( . to . mg/kg diluted with ml sterile saline) can be administered directly into the endotracheal tube via a rigid or red rubber catheter. if vascular access is available, epinephrine ( . to . mg/kg) can be administered intravenously. no drug should ever be administered directly into the heart by intracardiac injection. unless the heart is in the veterinarian's hand during open-chest cpcr, intracardiac injection is risky and potentially could lacerate a coronary artery or cause the myocardium to become more irritable and refractory to other therapies, if a drug is delivered into the myocardium and not into the ventricle. for these reasons, intracardiac injections are contraindicated. administer atropine ( . mg/kg iv, io, or . mg/kg it) immediately after the epinephrine. atropine, a vagolytic drug, serves to decrease tonic vagal inhibition of the sinoatrial and atrioventricular node and increase heart rate. administer atropine and epinephrine every to minutes during asystole while cardiac compressions, interposed abdominal compressions, and artificial ventilation are continued. although discontinuation of thoracic compressions can decrease the chance of success during cpcr, you must intermittently evaluate the ecg monitor for any rhythm change that may require different drug therapies. if the cardiac arrest was not witnessed or more than to minutes have passed without successful return to a perfusing rhythm, perform open-chest cpcr, if the client wishes. administer sodium bicarbonate ( to meq/kg iv) every to minutes during cpcr. sodium bicarbonate is the only drug used in cpcr that should not be administered intratracheally because of inactivation of pulmonary surfactant. electrical-mechanical dissociation also is known as pulseless electrical activity and is an electrical rhythm that may look wide and bizarre and irregular with no associated mechanical contraction of the ventricles. the rhythm can appear different from patient to patient. electrical-mechanical dissociation is one of the more common nonperfusing rhythms observed during cardiopulmonary arrest in small animal patients (figure - ) . when electrical-mechanical dissociation is identified, first confirm the rhythm and proceed with cpcr as previously described. electrical-mechanical dissociation is thought to be associated with high doses of endogenous endorphins and high vagal tone. the treatment of choice for electrical-mechanical dissociation is high-dose atropine ( mg/kg iv, it [ times the normal dose]) and naloxone hydrochloride ( . mg/kg iv, io, it). administer epinephrine ( . to . mg/kg diluted in ml sterile . % saline it). if the rhythm does not change within minutes, consider open-chest cardiac massage. ventricular fibrillation can be coarse (figure - ) . patients with coarse ventricular fibrillation are easier to defibrillate than those with fine defibrillation. if ventricular fibrillation is identified, initiate cpcr as described previously (figure - ) . if an electrical defibrillator is available, administer j/kg of direct current externally. when a patient in cardiopulmonary arrest is attached to ecg leads, it is important to use contact electrode paste, water-soluble gel such as ky jelly, or water, rather than any form of alcohol. electrical defibrillation of a patient who has alcohol on the ecg leads can lead to fire and thermal burns. reverse any opioid, α -agonist, and phenothiazine drugs that have been administered to the patient. if fine ventricular fibrillation is identified, administer epinephrine figure - : electrical-mechanical dissociation (emd), also known as pulseless electrical activity (pea). the complexes often appear wide and bizarre without a palpable apex beat or functional contraction of the heart. this is just one example of emd, as many shapes and complexes may be observed. organized according to whether an electrical defibrillator is available. after each intervention step, the ecg should be reevaluated and the next step initiated if v-fib is still seen. if a new arrhythmia develops, the appropriate therapy for that rhythm should be inititated. if a sinus rhythm is seen with a palpable apex beat, postresuscitation measures should be implemented. perform open-chest cpcr immediately if a pathologic condition exists that prevents enough of a change in intrathoracic pressure that closed-chest cpcr will not be effective in promoting forward blood flow (box . to perform open-chest cpcr, place the patient in right lateral recumbency. clip a wide strip of fur over the left fifth to seventh intercostal space and quickly aseptically scrub over the clipped area. using a no. scalpel blade, incise over the fifth intercostal space through the skin and subcutaneous tissue to the level of the intercostal muscles. with a mayo scissors, make a blunt stab incision through the intercostal muscles in the left sixth intercostal space. make sure that the person who is breathing for the patient deflates the lungs as you make the stab incision to avoid iatrogenic lung puncture. after the stab incision, open the tips of the mayo scissors and quickly open the muscle dorsally and ventrally to the sternum with a sliding motion. avoid the internal thoracic artery at the sternum and the intercostal arteries at the caudal aspect of each rib. cut the rib adjacent to the sternum and push it behind the rib in front of and at the caudal aspect of the incision to allow more room and better visualization if a rib spreading retractor is not available. visualize the heart in the pericardial sac. visualize the phrenic nerve, and incise the pericardium just ventral to the phrenic nerve. make sure to not cut the phrenic nerve. grasp the heart in your hand(s) and gently squeeze it from apex to base, allowing time for the ventricle to fill before the next "contraction." if the heart does not seem to be filling, administer fluids intravenously or directly into the right atrium. the descending aorta can be cross-clamped with a rummel tourniquet or red rubber catheter to improve perfusion to the brain and heart. postresuscitation care and monitoring (prolonged life support) postresuscitation care involves careful monitoring and management of the adverse effects of hypoxia and reperfusion injury on the brain and other vital organs. the first hours after an arrest are most critical, because this is the time period in which an animal is most likely to rearrest unless the underlying cause of the initial arrest has been determine and treated (table - ) . until an animal is adequately ventilating on its own, artificial ventilation by manual bagging or attaching the patient to a mechanical ventilator with supplemental oxygen must continue. the efficacy of oxygenation and ventilation can be monitored using a wright's respirometer, pulse oximetry, capnometry, and arterial blood . once an animal is extubated, administer supplemental oxygen ( to ml/ kg/minute) (see oxygen supplementation). the brain is sensitive to ischemia and reperfusion injury. the effects of cellular hypoxia and reperfusion include the development of oxygen-derived free radical species that contribute to cerebral edema. administer mannitol ( . to g/kg iv over to minutes), followed by furosemide ( mg/kg iv) minutes later, to all patients that have experienced cardiopulmonary arrest and have had successful resuscitation. mannitol and furosemide work synergistically to decrease cerebral edema formation and scavenge oxygen-derived free radical species. the combination of cardiac arrest, myocardial ischemia and acidosis, and external or internal cardiac compressions often make the myocardium irritable and predisposed to dysrhythmias following successful cpcr. start lidocaine ( to mg/kg iv, followed by to µg/kg/minute iv cri) in all patients following successful resuscitative efforts. monitor the ecg continuously for the presence of cardiac dysrhythmias and recurrence of nonperfusing rhythms. perform direct or indirect blood pressure monitoring. if a patient's systolic blood pressure is less than mm hg, diastolic pressure is less than mm hg, or mean arterial blood pressure is less than mm hg, administer positive inotropic drugs (dobutamine, to µg/kg/minute) and pressor agents (epinephrine, . to . mg/kg iv, io, it) to improve cardiac contractility, cardiac output, and core organ perfusion. the kidneys are sensitive to decreased perfusion and cellular hypoxia. place a urinary catheter and monitor urine output. in a euvolemic patient, normal urine output should be no less than to ml/kg/hour. if urine output is low, administer low-dose dopamine ( to µg/kg/minute iv cri) in an attempt to dilate afferent renal vessels and improve renal perfusion. maintain acid-base and electrolyte status within normal reference ranges. monitor serum lactate as a rough indicator of organ perfusion and cellular oxygen extraction. the presence of elevated or rising serum lactate in the face of aggressive cardiorespiratory and cerebral support makes prognosis less favorable. cole sg, otto cm, hughes d: cardiopulmonary cerebral resuscitation: a clinical practice review part i, j vet emerg crit care ( ) immediate action depends largely on recognition of the primary or secondary cause of the dysrhythmia and treating the dysrhythmia and underlying cause. diagnosis of cardiac dysrhythmias is based on physical examination findings of abnormal thoracic/cardiac auscultation, the presence of abnormal pulse rhythm and quality, and recognition of ecg abnormalities. the ecg is critical to the accurate diagnosis of dysrhythmias. ventricular dysrhythmias arise from ectopic foci in the ventricles that cause the wave of depolarization to spread from cell to cell rather than spread through fast-conducting tissue. this causes the qrs complex to appear wide and bizarre, unless the ectopic focus originates close to the atrioventricular node high in the ventricle. other ecg features of ventricular dysrhythmias include a t wave polarity that is opposite to the qrs complex and nonrelated p waves. ventricular dysrhythmias may manifest as isolated ventricular premature complexes, couplets, or triplets; bigeminy; or ventricular tachycardia. relatively slow ventricular tachycardia is known as an idioventricular rhythm and is not as hemodynamically significant as faster ventricular tachycardia. idioventricular rhythm usually is less than beats per minute and may alternate spontaneously with sinus arrhythmias (figures - to . supraventricular dysrhythmias arise from ectopic foci in the atria and are commonly associated with atrial dilatation and structural heart disease such as advanced acquired or congenital heart disease, cardiomyopathies, cardiac neoplasia, or advanced heartworm disease. occasionally, supraventricular dysrhythmias may be associated with respiratory or other systemic illness. sustained supraventricular tachycardia in the absence of underlying structural heart or systemic disease is disturbing and should alert the clinician that an accessory pathway conduction disturbance may be present, particularly in labrador retrievers. supraventricular dysrhythmias can manifest as isolated premature complexes (atrial premature complexes or contractions), sustained or paroxysmal supraventricular tachycardia (atrial tachycardia), or atrial fibrillation or flutter. in the dog, atrial fibrillation most commonly is associated with dilative cardiomyopathy. rarely and primarily in giant breed dogs, lone atrial fibrillation can occur with no underlying heart disease. atrial fibrillation and the resultant sustained elevation in ventricular rate are presumed to progress to dilative cardiomyopathy in such breeds. by comparison, atrial fibrillation is relatively uncommon in cats because of the small size of their atria but is associated most commonly with hypertrophic and restrictive cardiomyopathy. the ecg is critical to the diagnosis of a supraventricular dysrhythmia. the ecg usually demonstrates a normal appearance to the qrs complex unless aberrant conduction occurs in the ventricles, in which case the qrs can be wide but still originate from above the atrioventricular node. in most cases of a supraventricular dysrhythmia, some evidence of atrial activity including p waves, atrial flutter, or atrial fibrillation is apparent. in some cases, it may be difficult to diagnose the exact rhythm without slowing the rate down mechanically or through pharmacologic intervention. once a rhythm diagnosis is made, appropriate treatment strategies can be implemented (figures - and - ). treatment of ventricular dysrhythmias largely depends on the number of ectopic foci discharging, the rate and character of the dysrhythmia, and whether the presence of the abnormal beats is of adverse hemodynamic consequence, including risk of sudden death. many ventricular dysrhythmias, including slow idioventricular rhythms, ventricular bigeminy, or intermittent ventricular premature complexes, do not warrant antiarrhythmic therapy unless the patient is hypotensive and the dysrhythmia is thought to be contributing to the hypotension. in such cases, correction of the underlying disease process including hypoxia, pain, or anxiety often alleviates or decreases the incidence of the dysrhythmia. more serious ventricular dysrhythmias that warrant antiarrhythmic therapy (table - ) include sustained ventricular tachycardia (> beats/minute in dogs; > beats/minute in cats), multifocal ventricular premature complexes originating from more than one place in the ventricles, and the presence of r-on-t phenomena where the t wave of the preceding complex is superimposed on the qrs of the next complex with no return to isoelectric shelf in between complexes. treat these ventricular dysrhythmias immediately and aggressively. in dogs, the mainstay of emergency treatment for ventricular dysrhythmias is lidocaine therapy. administer lidocaine ( to mg/kg iv bolus) over a period of minutes to prevent the adverse side effects of seizures or vomiting. the bolus can be repeated an additional times (total dose mg/kg) over minutes, or the patient can be placed on a constant rate infusion ( to µg/kg/minute) if control of ventricular tachycardia is accomplished. also correct the patient's magnesium and potassium deficiencies to maximize the success of lidocaine therapy in the treatment of ventricular tachycardia. procainamide ( mg/kg iv slowly over to minutes) also can be used to control ventricular tachycardia. if procainamide is successful at controlling ventricular tachycardia, administer it as a constant rate infusion ( to µg/kg/minute). side effects of procainamide include vomiting, diarrhea, and hypotension. chronic oral therapy may or may not be necessary in the treatment of acute ventricular tachycardia. the decision to continue antiarrhythmic therapy depends on the underlying disease process and the expectation of persistent arrhythmogenesis of the underlying disease process. oral antiarrhythmic therapy is warranted in cases in which a serious ventricular dysrhythmia is recognized but the animal does not require hospitalization, such as the syncopal boxer with intermittent ventricular dysrhythmias and no evidence of structural heart disease. it deserves emphasis that asymptomatic, low-grade ventricular dysrhythmias probably do not require treatment. if maintenance therapy for ventricular dysrhythmias is needed, use an oral drug based on the underlying disease process, clinical familiarity, class of drug, dosing frequency, owner compliance, concurrent medications, cost, and potential adverse side effects. in the cat the mainstay of antiarrhythmic therapy is the use of a β-adrenergic antagonist. in the acute management of ventricular dysrhythmias in cases of hypertrophic, restrictive, or unclassified cardiomyopathies, consider using injectable esmolol ( . to . mg/kg iv slowly to effect) or propranolol ( . to . mg/kg iv slowly to effect), particularly if the dysrhythmia results from hyperthyroidism. for chronic oral ventricular antiarrhythmic therapy in cats, propranolol ( . to . mg po per cat q h) or atenolol ( . to . mg po per cat q - h) can be used. the decision to treat supraventricular dysrhythmias depends on the ventricular rate and the hemodynamic consequences of the dysrhythmia. for intermittent isolated atrial emergency care procainamide - mg/kg po q - h tocainide* - mg/kg po q h sotalol - mg per dog q h (start low, then titrate up to effect) mexiletine - mg/kg po q h atenolol . - . mg/kg po q - h (start low, titrate upward to effect) *do not use for longer than weeks because of idiosyncratic blindness. premature contractions, couplets, and triplets, usually no treatment is required. when the ventricular rate exceeds beats/minute, diastolic filling time is shortened, causing the heart to not fill adequately. the consequence is decreased cardiac output and decreased coronary artery perfusion. the goal of therapy is rhythm control or, in most cases, rate control. in cases of atrial fibrillation and congestive heart failure, conversion to a normal sinus rhythm rarely can be achieved, although electrocardioversion or pharmacoconversion can be attempted. in the dog a vagal maneuver can be attempted by pressing on the eyeballs or massaging the carotid body. for sustained supraventricular tachycardia, diltiazem ( . mg/kg iv), esmolol ( . to . , titrated upward to a cumulative dose of . mg/kg iv), or propranolol ( . to . mg/kg iv slowly to effect) can be administered in an attempt to slow the ventricular rate in emergent situations. administer oral diltiazem ( . mg/kg po q h), diltiazem (dilacor-xr) ( . to mg/kg po q - h), propranolol ( . to . mg/kg tid, titrated up to a maximum of . mg/kg po q h), atenolol ( . to mg/kg q - h), or digoxin ( . to . mg/kg bid or . mg/m for dogs greater than kg). in the cat a vagal maneuver can be attempted by ocular or carotid massage. (diltiazem [dilacor] to po q - h), propranolol ( . to mg/kg q - h), or atenolol ( . mg q - h) also can be administered. if structural heart disease is present, treat pulmonary edema and start angiotensin-converting enzyme inhibitor therapy. table - summarizes the drugs used in the management of supraventricular dysrhythmias. severe bradycardia often results from systemic disease, drug therapy, anesthetic agents, or hypothermia and thus rarely requires specific therapy except to treat or reverse the underlying mechanisms promoting bradycardia. hemodynamically significant bradyarrhythmias that must be treated include atrial standstill, atrioventricular block, and sick sinus syndrome. atrial standstill most commonly is associated with hyperkalemia and is seen most often in urinary obstruction, renal failure, urinary trauma with uroabdomen, and hypoadrenocorticism. characteristic ecg abnormalities observed in atrial standstill are an absence of p waves, widened qrs complexes, and tall spiked t waves (figure - ). the treatment for hyperkalemia-induced atrial standstill is to correct the underlying cause and to drive potassium intracellularly and protect the myocardium from the adverse effects of hyperkalemia. regular insulin ( . to . units/kg iv) followed by dextrose ( g/unit insulin iv, followed by . % dextrose cri to prevent hypoglycemia) or sodium bicarbonate ( meq/kg iv) can be administered to drive potassium intracellularly. calcium gluconate ( . ml/kg of % solution iv over minutes) also can be administered as a cardioprotective drug until the cause of hyperkalemia has been identified and resolved. also administer sodium chloride fluids ( . % sodium chloride iv) to promote kaliuresis. less commonly, atrial standstill is associated with atrial cardiomyopathy or silent atrium syndrome. persistent atrial standstill has been recognized without electrolyte abnormalities in the english springer spaniel and the siamese cat. short-term therapy for persistent atrial standstill includes atropine ( . mg/kg sq) until definitive treatment by implantation of a cardiac pacemaker can be performed. complete or third-degree atrioventricular block or high-grade symptomatic seconddegree atrioventricular block can be hemodynamically significant when ventricular rates are less than beats/minute in the dog. classic clinical signs include weakness, exercise intolerance, lethargy, anorexia, syncope, and occasionally seizures. advanced atrioventricular block usually is caused by advanced idiopathic degeneration of the atrioventricular node. less commonly, atrioventricular block has been associated with digoxin toxicity, magnesium oversupplementation, cardiomyopathy, endocarditis, or infectious myocarditis (lyme disease). an accurate diagnosis is made based on the ecg findings of nonconducted p waves with ventricular escape beats. first-and second-degree atrioventricular block may not be hemodynamically significant and therefore may not require therapy. initially treat third-degree (complete) or symptomatic high-grade second-degree atrioventricular block (< beats/minute) with atropine ( . mg/kg sq or im). perform a follow-up ecg in to minutes. atropine is rarely successful in treating complete atrioventricular block. also attempt treatment with isoproterenol ( . to . µg/kg/minute iv cri or . mg in ml % dextrose in water iv slowly), a pure β-agonist. definitive treatment requires permanent pacemaker implantation. consultation with a veterinary cardiologist who implants pacemakers is suggested. never attempt to convert or treat the observed ventricular escape beats with lidocaine ( figure - ) . sick sinus syndrome most commonly is recognized in the miniature schnauzer, although any dog can be affected. sick sinus syndrome usually results from idiopathic degeneration of the sinus node in the dog. in the cat, sinus node degeneration usually is associated with cardiomyopathy. dysfunction of the sinus node may manifest as marked bradycardia with periods of sinus arrest followed by junctional or ventricular escape complexes. a variant of sick sinus syndrome is the presence of severe bradycardia followed by periods of supraventricular tachycardia, often termed bradycardia-tachycardia syndrome. the most common clinical signs are syncope, exercise intolerance, and lethargy. in cats, hypertrophic cardiomyopathy is the most common form of acquired cardiac disease observed. congestive heart failure resulting from hypertrophic cardiomyopathy can occur in animals as young as to months of age. hypertrophic cardiomyopathy is characterized by stiff, noncompliant ventricles that do not relax during diastole, causing an increase in left atrial pressures and left atrial enlargement. other cardiomyopathies, including unclassified, restrictive, and dilated, are less common but also can occur in the cat. cats often develop acute exacerbation of clinical signs because of stress or arterial embolization. the rapid diagnosis of chf often is made on owner history, signalment, and physical examination findings (box - ). typical physical examination findings include a cardiac murmur or gallop dysrhythmia, abnormal breath sounds, respiratory difficulty and orthopnea, tachycardia, weak pulse quality, cool peripheral extremities, and pale or cyanotic mucous membrane. initiate immediate treatment based on physical examination findings and index of suspicion. in some cases, it is difficult to distinguish between chf and feline lower airway disease (asthma) without performing thoracic radiographs. let the animal rest and become stabilized before attempting any stressful procedures, including thoracic radiographs. immediate treatment consists of administering supplemental oxygen, decreasing circulating fluid volume with furosemide, dilating pulmonary and splanchnic capacitance vessels with topical nitroglycerine and morphine, and alleviating patient anxiety and stress (box - ). primary differential diagnoses are made based primarily on the patient's breed, age, clinical signs, history, and physical examination abnormalities. the most common differential diagnoses in a patient with chf are cardiac abnormalities and respiratory disease (chronic bronchitis [asthma], pulmonary hypertension, cor pulmonale, neoplasia). postpone diagnostic tests in any patient with suspected chf until the immediate treatments have taken effect and the patient is cardiovascularly more stable. in most cases, lateral and dorsoventral thoracic radiographs are one of the most important diagnostic tools in helping make a diagnosis of chf. increased perihilar interstitial to alveolar infiltrates are characteristic of pulmonary edema. left atrial enlargement may be observed as a "backpack" sign at the caudal cardiac waist. cardiomegaly of the right or left side also may be present in cases of valvular insufficiency. in cats, increased sternal contact and a classic valentine-shaped heart may be observed in cases of hypertrophic cardiomyopathy. perform a vertebral heart score (sum) to measure cardiac size and determine whether cardiomegaly is present (box - ). also obtain arterial blood pressure and ecg readings to determine whether hypotension and dysrhythmias are present. atrial fibrillation, ventricular premature contractions, and supraventricular tachycardia are common rhythm disturbances that can affect cardiac output adversely and influence treatment choices. the echocardiogram is a useful noninvasive and nonstressful method to determine the degree of cardiac disease present. the echocardiogram is largely user-dependent. the quality of the study is based on the experience of the operator and the quality of the ultrasound machine. echocardiography can be a useful tool in making a diagnosis of pericardial effusion, dilated or hypertrophic cardiomyopathy, cardiac neoplasia, and endocarditis. the medical management of chf is designed to improve cardiac output and relieve clinical signs. the immediate goal of therapy is to reduce abnormal fluid accumulation and provide adequate cardiac output by increasing contractility, decreasing preload and ventricular afterload, and/or normalizing cardiac dysrhythmias. strict cage rest is of utmost importance when managing a patient with chf. after initial administration of furosemide, morphine, oxygen, and nitroglycerine paste, clinical signs of respiratory distress should show improvement within minutes. if no improvement is observed, administer repeated doses of furosemide. reevaluate severe cases that are refractory to this standard treatment protocol. vasodilation should be the next step in the management of refractory cases, provided that a normal blood pressure is present. sodium nitroprusside is a potent balanced vasodilator that should be administered ( to µg/kg/minute iv cri), taking care to monitor blood pressure continuously because severe vasodilation and hypotension can occur. the goal of nitroprusside therapy is to maintain a mean arterial blood pressure of mm hg. sodium nitroprusside should not be considered in cases of refractory chf with severe hypotension. for more long-term management of chf, the use of angiotensin-converting enzyme (ace) inhibitors including enalapril ( . mg/kg po q - h), benazepril ( . mg/kg po q h), and lisinopril ( . mg/kg po q h) have become the mainstay of therapy to reduce sodium and fluid retention and decrease afterload. start angiotensin-converting enzyme inhibition as soon as a patient is able to tolerate oral medications. dobutamine ( . to µg/kg/minute cri diluted in % dextrose in water) can be administered to improve cardiac contractility, particularly in cases of dilated cardiomyopathy. at low doses, dobutamine, primarily a β-adrenergic agonist, will improve cardiac output with minimal effects on heart rate. dobutamine must be given as a constant rate infusion with careful, continuous ecg monitoring. despite minimal effects on heart rate, emergency management of specific conditions the vertebral heart sum can be calculated by performing the following steps: . measure the long axis of the heart from the apex to the carina on the lateral view and mark the distance on a sheet of paper. . measure the length of the long axis of the heart in terms of vertebral bodies, starting by counting caudally from the fourth thoracic vertebra; count the number of vertebrae that are covered by the length of the long axis of the heart. . measure the short axis of the heart at the caudal vena cava, perpendicular to the long axis of the heart. . count the number of thoracic vertebrae covered by the short axis of the heart, starting at t . . add the two numbers together to yield the vertebral heart sum; a vertebral heart sum greater than . is consistent with cardiomegaly. sinus tachycardia or ventricular dysrhythmias may develop during infusion. cats are more sensitive to the effects of dobutamine than dogs. monitor carefully for seizures and facial twitching. digoxin is a cardiac glycoside that acts as a positive inotrope and negative chronotrope in the long-term management of chf. digoxin has a long ( hours in dogs, and hours in cats) half-life and so has minimal use in the emergency management of chf. in chronic management of chf resulting from dilated cardiomyopathy or advanced mitral disease, however, digoxin is extremely useful. oral digitalization protocols have been developed but are risky in that dysrhythmias and severe gastrointestinal side effects can occur. cats with chf often have fulminant pulmonary edema, pleural effusion, arterial thromboembolism, or some combination of all three. if the pleural effusion is significant, perform therapeutic thoracocentesis to relieve pulmonary atelectasis and improve oxygenation. once the diagnosis and initial management of chf has been made, formulate a plan for continued management and monitoring. tailor the therapeutic plan to the patient based on the cause of the chf, the presence of concurrent diseases, and response to therapy. an important and often overlooked part of the successful emergency management of chf is the open communication with the owner regarding the owner's emotional and financial commitment for immediate and long-term management to ensure appropriate quality of life for each patient. pathophysiology and treatment, vet j ( ) caval syndrome resulting from severe heartworm disease is caused by the rapid maturation of a large quantity of adult worms in the right atrium and cranial and caudal venae cavae. most cases of caval syndrome occur in regions of the world where heartworm disease is highly endemic and dogs spend a large portion of time living outdoors. caval syndrome is recognized by the following clinical signs and results of biochemical analyses: acute renal and hepatic failure, enlarged right atrium and posterior vena cava, ascites, hemoglobinuria, anemia, acute collapse, respiratory distress, dic, jugular pulses, circulating microfilariae, and sometimes tricuspid insufficiency. immediate action in cases of caval syndrome in dogs involves immediate stabilization of the cardiovascular and respiratory systems with supplemental oxygen, furosemide ( mg/kg iv), and careful crystalloid fluid infusion. diagnosis of caval syndrome is based on clinical signs of cardiogenic shock with right ventricular heart failure, intravascular hemolysis, and renal and hepatic failure. thoracic radiographs reveal cardiomegaly of the right side and enlarged tortuous pulmonary arteries. a right axis deviation may be seen on ecg tracings. clinicopathologic changes observed include azotemia, inflammatory leukogram, regenerative anemia, eosinophilia, elevated hepatocellular enzyme activities, hemoglobinuria, and proteinuria. circulating microfilariae may be observed on peripheral blood smears or in the buffy coat of microhematocrit tubes. heart worm antigen tests will be strongly positive. echocardiographic changes include visualization of a large number of heartworms in the right atrium, pulmonary arteries, and vena cava, tricuspid insufficiency, and right atrial and ventricular enlargement. treatment involves surgical removal of as many of the adult heartworms as possible from the right jugular vein and right atrium. glucocorticosteroids are recommended to decrease inflammation and microangiopathic disease associated with heartworm infection. for more long-term management, administer adulticide therapy several weeks following surgery, followed by routine microfilaricide therapy and then prophylaxis. calvert pericardial effusion often develops as a consequence of neoplasia in the older dog and cat. the most common types of neoplasia that affect the heart and pericardium include hemangiosarcoma, chemodectoma, mesothelioma, and metastatic neoplasia. more rarely, other causes of pericardial effusion include benign idiopathic pericardial effusion, coagulopathy, left atrial rupture in dogs with chronic mitral valvular insufficiency, infection, or pericardial cysts. regardless of the cause of the effusion, the development of pericardial tamponade adversely affects cardiac output. cardiac output is a function of heart rate and stroke volume. stroke volume depends on cardiac preload. the presence of pericardial effusion can impede venous return to the heart and thus adversely affect preload. in addition, as preload decreases, heart rate reflexively increases in an attempt to maintain normal cardiac output. as heart rate increases more than beats/minute, diastolic filling is impaired further, and cardiac output further declines. animals with pericardial effusion often demonstrate the classic signs of hypovolemic or cardiogenic shock: anorexia, weakness, lethargy, cyanosis, cool peripheral extremities, tachycardia, weak thready pulses, hypotension, and collapse. physical examination abnormalities may include muffled heart sounds, thready femoral pulses, pulsus paradoxus, jugular venous distention, weakness, tachycardia, cyanosis, and tachypnea. electrocardiogram findings may include low amplitude qrs complexes (< . mv), sinus tachycardia, ventricular dysrhythmias, or electrical alternans (figure - ) . thoracic radiographs often demonstrate a globoid cardiac silhouette, although the cardiac silhouette rarely may appear normal with concurrent clinical signs of cardiogenic shock in cases of acute hemorrhage. in such cases the removal of even small amounts of pericardial effusion by pericardiocentesis can increase cardiac output exponentially and alleviate clinical signs (table - ) . unless an animal is dying before your eyes, ideally perform an echocardiogram to attempt to determine whether a right atrial, right auricular, or heart base mass is present before pericardiocentesis. before attempting pericardiocentesis, assemble all of the required supplies (box - ) . to perform pericardiocentesis, follow this procedure: . place the patient in sternal or lateral recumbency. . attach ecg leads to monitor the patient for dysrhythmias during the procedure. . clip a -cm square caudal to the right elbow over the fifth to seventh intercostal space. . aseptically scrub the clipped area, and infuse to mg/kg of % lidocaine mixed with a small amount of sodium bicarbonate just dorsal to the sternum at the sixth intercostal space. bury the needle to the hub, and inject the lidocaine as you withdraw the needle. . while the local anesthetic is taking effect, assemble the intravenous extension tubing, three-way stopcock, and -ml syringe. . wearing sterile gloves, make a small nick incision in the skin to decrease drag on the needle and catheter during insertion. . slowly insert the needle and catheter, watching for a flash of blood in the hub of the needle, and simultaneously watching for cardiac dysrhythmias on the ecg monitor. . once a flash of blood is observed in the hub of the needle, advance the catheter off of the stylette further into the pericardial sac, and remove the stylette. . attach the length of intravenous extension tubing to the catheter, and have an assistant withdraw the fluid slowly. . place a small amount of fluid in a red-topped tube, and watch for clots. clot formation could signify that you have penetrated the right ventricle inadvertently or that active hemorrhage is occurring. withdraw as much of the fluid as possible, and then remove the catheter. monitor the patient closely for fluid reaccumulation and recurrence of clinical signs of cardiogenic shock. less rd, bright jm, orton ec: intrapericardial cyst causing cardiac tamponade in a cat, j am anim hosp assoc ( ) foreign bodies within the ear canal (e.g., foxtails) can present as emergencies because of acute inflammation and pressure necrosis of the tissue of the external auditory meatus causing pain and discomfort. clinical signs may be limited to incessant head shaking or scratching of the ear canal. complete examination of the ear canal and removal of any foreign body often requires administration of a short-acting anesthetic agent. once the animal has been restrained sufficiently and placed under anesthesia, carefully examine the ear canal and remove any foreign material with an alligator forceps. stimulation of the ear canal can cause awakening after removal of all debris and detritus, gently wipe the internal and external ear canal with a sterile gauze. place a topical antimicrobial-antifungal-steroid ointment such as otomax in the ear every to hours. if pain and discomfort is severe, systemically effective opioids or nsaids may be required. otitis externa is a common emergency that causes excessive head shaking, scratching, and purulent malodorous aural discharge. clean the ear canal with an irrigating solution such as epiotic and wipe it clean of debris. perform a complete aural examination to determine whether a foreign body or tumor is present and whether the tympanic membrane is intact. heat-fix any discharge and examine it cytologically for bacteria and fungal organisms. following careful cleansing, instill a topical antibiotic-antifungal-steroid ointment. in severe cases in which the ear canal has scarred and closed down with chronicity, consider administering systemically effective antibiotics (cephalexin, mg/kg po tid) and antifungal agents (ketoconazole, mg/kg po q h) instead of topical therapy. systemically effective steroids (prednisone or prednisolone, . mg/kg po q h) may be indicated in cases of severe inflammation to decrease pruritus and patient discomfort. presentation of a patient with otitis interna often is characterized by torticollis, head tilt, nystagmus, circling to the affected side, or rolling. fever, pain, vomiting, and severe depression may accompany clinical signs. most cases of severe otitis interna are accompanied by severe otitis media. both conditions must be treated simultaneously. the most common causes of otitis interna are staphylococcus aureus, pseudomonas, escherichia coli, or proteus spp. otitis interna can develop by infection spreading across the tympanic membrane, through the eustachian tubes, or by hematogenous spread from the blood supply to the middle ear. in most cases of otitis media, the tympanic membrane is ruptured. perform a culture and susceptibility test of the debris behind the tympanic membrane and within the aural canal. carefully clean the external ear canal. medicate with a topical combination antibiotic, antifungal, and antibiotic ointment. administer high-dose antibiotics (cephalexin, mg/kg po q h, or enrofloxacin, to mg/kg po q h). if the tympanic membrane is not ruptured but appears swollen and erythematous, a myringotomy may need to be performed. if clinical signs of otitis media persist despite topical and systemic therapy, radiographic or ct/mri examination of the tympanic bullae may be required. chronic shaking of the head and ears or aural trauma (bite wounds) causes disruption of the blood vessels and leads to the development of unilateral or bilateral aural hematomas. aural hematomas are clinically significant because they cause patient discomfort and are often due to the presence of some other underlying problem such as otitis externa, atopy, or aural foreign bodies. acute swelling of the external ear pinna with fluid is characteristic of an aural hematoma. in some cases, swelling can be so severe that the hematoma breaks open, bathing the patient and external living environment in blood. when a patient has an aural hematoma, investigate the underlying cause. perform a complete aural examination to determine whether an aural foreign body, otitis externa, or atopy are present. carefully examine and gently clean the inner ear canal. treat underlying causes. management of an aural hematoma involves draining the hemorrhagic fluid from the aural tissue and tacking the skin down in multiple places to prevent reaccumulation of fluid until the secondary cause is resolved. many techniques have been described to surgically tack down the skin overlying the hematoma. after the animal has been placed under general anesthesia, lance the hematoma down the middle with a scalpel blade and remove the fluid and blood clot. tack down the skin with multiple through-and-through interrupted or mattress sutures through the ear. some clinicians prefer to suture through and attach a sponge or length of x-ray film to the front and back of the ear for stabilization and support. more recently, a laser can be used to drill holes in the hematoma and tack the skin down in multiple areas. compress the ear against the head with a compression bandage, whenever possible, for to days after the initial surgery, and then recheck the ear. the patient must wear an elizabethan collar until the surgical wound and hematoma heal to prevent selfmutilation. also systemically treat underlying causative factors such as otitis externa with antibiotics, antifungals, and steroids as indicated. investigate and treat other underlying causes such as hypothyroidism or allergies. bass electrocution usually is observed in young animals after they have chewed on an electric cord. other causes of electrocution include use of defective electrical equipment or being struck by lightning. electric current passing through the body can produce severe dysrhythmias, including supraventricular or ventricular tachycardia and first-and thirddegree atrioventricular block. the electric current also can produce tissue destruction from heat and electrothermal burns. electrocution also commonly results in noncardiogenic pulmonary edema caused by massive catecholamine release and increase in pulmonary vascular pressures during the event. ventricular fibrillation can occur, although that depends on the intensity and path of the electrical current and duration of contact. clinical signs of electrocution include acute onset of respiratory distress with moist rales, and localized necrosis or thermal burns of the lips and tongue. often the skin at the commissures of the mouth appears white or yellow and firm to the touch. muscle fasciculations, loss of consciousness, and ventricular fibrillation may occur. thoracic radiographs often reveal an increased interstitial to alveolar lung pattern in the dorsocaudal lung fields. noncardiogenic pulmonary edema can develop up to to hours after the initial incident. the first hours are most critical for the patient, and then prognosis improves. the most important aspect in the treatment of the patient with noncardiogenic pulmonary edema is to minimize stress and to provide supplemental oxygen, with positive pressure ventilation, when necessary. although treatment with vasodilators (low-dose morphine) and diuretics (furosemide) can be attempted, noncardiogenic pulmonary edema is typically resistant to vasodilator and diuretic therapy. positive inotropes and pressor drugs may be necessary to treat shock and hypotension. opioid drugs (morphine, hydromorphone, oxymorphone) may be useful in controlling anxiety until the pulmonary edema resolves. administer broad-spectrum antibiotics (cefazolin; amoxicillin and clavulanic acid [clavamox]) to treat thermal burns. use analgesic drugs to control patient discomfort. if thermal burns are extensive and prohibit adequate food intake, place a feeding tube as soon as the patient's cardiovascular and respiratory function are stable and the patient can tolerate anesthesia. prolapse of the uterus occurs in the immediate postparturient period in the bitch and queen. excessive straining during or after parturition causes the uterus to prolapse caudally through the vagina and vulva. immediate intervention is necessary. examine the bitch or queen for a retained fetus. treatment consists of general anesthesia to replace the prolapsed tissue. if the uterus is edematous, physical replacement may be difficult or impossible. application of a hypertonic solution such as hypertonic ( %) saline or dextrose ( %) to the exposed endometrium can help shrink the tissue. that, combined with gentle massage to stimulate uterine contraction and involution and lubrication with sterile lubricating jelly, can aid in replacement of the organ into its proper place. to ensure proper placement in the abdominal cavity and to prevent recurrence, perform an exploratory laparotomy and hysteropexy. postoperatively, administer oxytocin ( to units im) to cause uterine contraction. if the uterus contracts, it is usually not necessary to suture the vulva. administer antibiotics postoperatively. recurrence is uncommon, even with subsequent pregnancies. if the tissue is damaged or too edematous to replace or if the tissue is devitalized, traumatized or necrotic, perform an ovariohysterectomy. in some instances, replacement of the damaged tissue is not necessary before removal. pyometra occurs in dogs and cats. the disease process occurs as a result of infection overlying cystic endometrial hyperplasia under the constant influence of progesterone. during the -month luteal phase after estrus or following copulation, artificial insemination, or administration of hormones (particularly estradiol or progesterone), the myometrium becomes relaxed and favors a quiescent environment for bacterial proliferation. clinical signs of pyometra are associated with the presence of bacterial endotoxin and sepsis. early, affected animals become lethargic and anorectic. polyuria with secondary polydipsia is often present because of the influence of bacterial endotoxin on renal tubular concentration. if the cervix is open, purulent or mucoid vaginal discharge may be observed. later in the course of pyometra, vomiting, diarrhea, and progressive debilitation resulting from sepsis occur. diagnosis is based on clinical signs in an intact queen or bitch and radiographic or ultrasonographic evidence of a fluid-filled tubular density in the ventrocaudal abdomen, adjacent to the urinary bladder (figures - and - ) . treatment of open and closed pyometra is correction of fluid and electrolyte abnormalities, administration of broad-spectrum antibiotics, and ovariohysterectomy. close pyometra is a life-threatening septic condition. open pyometra also can become life-threatening and so should be treated aggressively. in closed pyometra, conservative medical therapy is not advised. administration of prostaglandins and oxytocin do not reliably cause the cervix to open and can result in ascending infection from the uterus into the abdomen or uterine rupture, both of which can result in severe peritonitis. for animals with an open pyometra, ovariohysterectomy is the most reliable treatment for chronic cystic endometrial hyperplasia. although less successful than ovariohysterectomy, medical therapy may be attempted in breeding bitches as an alternative to surgery. the most widely used medical therapy in the breeding queen and bitch is administration of prostaglandin f α . this drug has not been approved for use in the queen or bitch in the united states. to proceed with medical management of pyometra, first determine the size of the uterus. start the patient on antibiotic therapy (ampicillin, mg/kg iv q h, or enrofloxacin, mg/kg po q h). administer the prostaglandin f α ( µg/kg sq q h) for to days until the size of the uterus approaches normal. measure serum progesterone concentrations if the bitch is in diestrus. as the corpus luteum degrades under the influence of prostaglandin f α , serum progesterone levels will decline. prostaglandin f α is an abortifacient and thus should not be administered to the pregnant bitch or queen. clinical signs of a reaction to prostaglandin f α can occur within to minutes in the bitch and can last for as long as minutes. clinical signs of a reaction include restlessness, hypersalivation, panting, vomiting, defecation, abdominal pain, fever, and vocalization. in a very ill animal, death can occur. the efficacy of prostaglandin f α is limited and may require more than one treatment. the bitch should be bred on the next heat cycle and then spayed because progressive cystic endometrial hyperplasia will continue to occur. acute metritis is an acute bacterial infection of the uterus that typically occurs within to weeks after parturition. the most common organism observed in metritis is e. coli ascending from the vulva and vaginal vault. sepsis can progress rapidly. clinical signs of acute metritis include inability to nurse puppies, anorexia, lethargy, foul-smelling purulentsanguineous vaginal discharge, vomiting, or acute collapse. physical examination may reveal fever, dehydration, and a turgid distended uterus. septic inflammation will be observed on vaginal cytologic examination. an enlarged uterus can be observed with abdominal radiographs and ultrasonography. treatment of acute metritis is directed at restoring hydration status with intravenous fluids and treating the infection with antibiotics. because the primary cause of metritis is e. coli infection, start enrofloxacin ( mg/kg iv or po once daily) therapy. as soon as the patient's cardiovascular status is stable enough for anesthesia, perform an ovariohysterectomy. if the patient is not critical and is a valuable breeding bitch, medical therapy can be attempted. medical management of acute bacterial metritis includes administration of oxytocin ( to units q h for three treatments) or administration of prostaglandin f α ( µg/kg/day for to days) to evacuate the uterine exudate and increase uterine blood flow. either drug should be used concurrently with antibiotics. rupture of the gravid uterus is rare in cats and dogs but has been reported. uterine rupture may occur as a consequence of parturition or result from blunt abdominal trauma. feti expelled into the abdominal cavity may be resorbed but more commonly cause the development of peritonitis. if fetal circulation is not disrupted, the fetus actually may live to term. uterine rupture is an acute surgical emergency. an ovariohysterectomy with removal of the extrauterine puppies and membranes is recommended. if only one horn of the uterus is affected, a unilateral ovariohysterectomy can be performed to salvage the remaining unaffected puppies and preserve the breeding potential for the valuable bitch. if uterine rupture occurs because of pyometra, peritonitis is likely, and copious peritoneal lavage should be performed at the time of surgery. the patient should be placed on to days of antibiotic therapy (amoxicillin or amoxicillin and clavulanic acid [clavamox] with enrofloxacin). vaginal prolapse occurs from excessive proliferation and hyperplasia of vaginal tissue while under the influence of estrogen during proestrus (figure - ) . the hyperplastic tissue usually recedes during diestrus but reappears with subsequent heat cycles. vaginal prolapse can be confused with vaginal neoplasia. the former condition occurs primarily in younger animals, whereas the latter condition occurs primarily in older animals. treatment for vaginal hyperplasia or prolapse generally is not required if the tissue remains within the vagina. the proliferation can lead to dysuria or anuria, however. in some cases, the tissue becomes emergency care dried out and devitalized or becomes traumatized by the animal. such extreme cases warrant immediate surgical intervention. the treatment for vaginal prolapse consists of ovariohysterectomy to remove the influence of estrogen, placement of an indwelling urinary catheter if the patient is dysuric, and protection of the hyperplastic tissue until it recedes on its own. although surgical resection of the hyperplastic tissue has been recommended, excessive hemorrhage after removal can occur, and so the procedure should not be attempted. the patient should wear an elizabethan collar at all times to prevent selfmutilation. administer broad-spectrum antibiotics for a minimum of to days or until the hyperplastic tissue recedes. keep the tissue clean with saline solution. dystocia, or difficult birth, can occur in the dog and cat but is more common in the dog. a diagnosis of dystocia is made based on the time of onset of visible labor and the time in which the last puppy or no puppy has been born, the intensity and timing of contractions, the timing of when the amniotic membranes first appear, the condition of the bitch, and the timing of gestation. causes of dystocia can be maternal or fetal and include primary or secondary uterine inertia, narrowing of the pelvic canal, hypocalcemia, psychological disturbances, or uterine torsion. maternal-fetal disproportion, or large fetus size in relation to the bitch or queen, also can result in dystocia (box - ). obtain an abdominal radiograph for all cases of suspected dystocia at the time of presentation to determine the size of the fetus, presentation of the fetus (both anterior or posterior presentation can be normal in the bitch or queen, but fetal malpositioning can cause dystocia), and whether there is radiographic evidence of a uterine rupture or torsion. if maternal-fetal disproportion, uterine torsion, or uterine rupture is observed, take the patient immediately to surgery. if the puppies or kittens are in a normal position for birth, medical management can be attempted. clip the perineum and aseptically scrub it. wearing sterile gloves, insert a lubricated finger into the vagina and palpate the cervix. massage (or "feather") the dorsal wall of the vagina to stimulate contractions. place an intravenous catheter, and administer oxytocin ( to units im), repeating up to times at -minute intervals. in some cases, hypoglycemia or hypocalcemia can contribute to uterine inertia. administration of a calciumcontaining solution (lactated ringer's solution) with . % dextrose is advised. alternately, administer % calcium gluconate ( mg/ kg iv slowly). if labor has not progressed after hour, immediately perform a cesarean section. uterine torsion is an uncommon emergency seen in the gravid and nongravid uterus and has been reported in dogs and cats. the onset of clinical signs of abdominal pain and straining as if to whelp/queen or defecate is usually acute and constitutes a surgical emergency. in some cases, there may have been a history of delivery of a live or dead fetus. vaginal discharge may or may not be present. radiographs or ultrasound examination reveal a fluid-filled or air-filled tubular density in the ventral abdomen. treatment consists of placing an intravenous catheter, stabilizing the patient's cardiovascular status with intravenous fluids and sometimes blood products, and performing an immediate ovariohysterectomy. if there are viable feti, the uterus should be delivered en mass and the puppies or kittens delivered. the expulsion of one or more fetus before term is known as spontaneous abortion. in dogs and cats, it is possible to expel or abort one or more fetuses and still carry viable fetuses to term and deliver normally. clinical signs of spontaneous abortion include vaginal discharge and abdominal contractions. in some cases, the fetus is found, or there may be evidence of fetal membranes or remnants. causes of spontaneous abortion in dogs include brucella canis, herpesvirus, coronavirus, and toxoplasmosis. in cats, herpesvirus, coronavirus, and feline leukemia virus can cause spontaneous abortion. in both species, trauma, hormonal factors, environmental pathogens, drugs, and fetal factors also can result in spontaneous abortion. the safest method of pregnancy termination in the bitch or queen is by performing an ovariohysterectomy. oral diethylstilbesterol is not an effective mechanism of pregnancy termination in the bitch. a so-called mismating shot, an injection of estradiol cypionate ( . mg/lb im) is effective at causing termination of an early pregnancy but can be associated with severe side effects, including bone marrow suppression and pyometra. estradiol cypionate is not approved for use in the bitch or queen and is not recommended. prostaglandin f α is a natural abortifacient in the bitch if treatment is started within days of cytologic evidence of diestrus (noncornified epithelium on a vaginal smear). the prostaglandin f α causes lysis of the corpora lutea and a rapid decline in progesterone concentration. the prostaglandin f α is administered for a total of eight injections ( µg/kg q h for days), along with atropine ( to µg/kg sq). side effects can occur within to minutes of injection and include restlessness, panting, salivation, abdominal pain, urination, vomiting, and diarrhea. walking the patient for to minutes after each treatment sometimes decreases the intensity of the reactions. bitches in the first half of the pregnancy often resorb the embryos. if prostaglandin f α is administered in the second half of the pregnancy, the fetuses are aborted within to days of treatment. measure serum progesterone concentrations at the end of treatment to ensure complete lysis of the corpus luteum. prostaglandin f α is not approved for pregnancy termination in the bitch. in cats, prostaglandin f α can terminate pregnancy after day of gestation. prostaglandin f α should be used only in healthy queens ( to µg/kg sq q h for days). side effects in the queen are similar to those observed in the bitch but typically have a shorter duration ( to minutes). prostaglandin f α is not approved for use in cats in the united states. the use of prostaglandin f α does not preclude breeding and pregnancy at a later date. biddle d, macintire dk: obstetrical emergencies, clin tech small anim pract ( ) in the dog and cat the majority of injuries to the scrotum are associated with animal fights or shearing and abrasive injuries sustained in accidents involving automobiles. scrotal injuries should be categorized as superficial or penetrating. treatment of superficial injuries to the scrotum includes cleaning the wound with dilute antimicrobial cleanser and drying it. administer antiinflammatory doses of steroids (prednisolone, . to . mg/kg po q - h) or nsaids (carprofen, . mg/kg po q h in dogs) for the first several days after scrotal injury to prevent or treat edema. administer topical antibiotic ointment until the wound heals. in most cases, place an elizabethan collar to prevent self-mutilation. prognosis is generally favorable; however, semen quality may be affected for months after injury because of scrotal swelling and increased scrotal temperature. penetrating injuries to the scrotum are more serious and are associated with severe swelling and infection. surgically explore and debride penetrating scrotal wounds. administer systemically effective antibiotics and analgesics. in extreme cases, particularly those that involve the testicle, consider castration and scrotal ablation. scrotal dermatitis is common in intact male dogs and can be associated with direct physical injury, self-infliction from licking, chemical irritation, burns, or contact dermatitis. in affected animals, the scrotum can become extremely inflamed, swollen, and painful. if left untreated, pyogranulomatous dermatitis can develop. make an attempt to determine whether an underlying systemic illness is present that could predispose the animal to scrotal dermatitis. widespread vasculitis with scrotal edema, pain, fever, and dermatitis has been associated with rickettsia rickettsii (rocky mountain spotted fever) infection. brucella canis also has been associated with scrotal irritation and dermatitis. if scrotal dermatitis follows from an infectious cause, empiric use of glucocorticosteroids potentially can make the condition worse by suppressing immune function. empiric treatment with antibiotics also potentially can confound making an accurate diagnosis. treatment of scrotal dermatitis is to eliminate predisposing causes, if possible. place an elizabethan collar at all times to prevent self-mutilation. bathe the scrotum with a mild antimicrobial soap and dry it to remove any offending chemical irritants. topical medications including tar shampoo, tetracaine, neomycin, and petroleum can cause further irritation and are contraindicated. use oral or parenteral administration of glucocorticosteroids or nsaids to control discomfort and inflammation. scrotal hernias occur when the contents of the abdomen (intestines, fat, mesentery, omentum) protrude through the inguinal ring into the scrotal sac. like inguinal hernias, scrotal definitive therapy for a scrotal hernia involves exploratory laparotomy and surgical reduction of the contents of the hernia, surgical correction of the rent in the inguinal ring, and castration. trauma to the epididymis or testicle can cause testicular pain and swelling of one or both testes. treat penetrating trauma to the testicle by castration to prevent infection and selfmutilation. administer oral antibiotics (amoxicillin or amoxicillin-clavulanate) for to days after the injury. nonpenetrating injuries to the scrotum and testicle rarely may cause acute testicular hemorrhage or hydrocele formation. palpation of the affected area often reveals a peritesticular, soft, compliant area. treatment consists of cool compresses on the scrotum and testicle and administration of antiinflammatory doses of glucocorticosteroids or nsaids. if the swelling does not resolve spontaneously in to days, consider surgical exploration and drainage. increased scrotal temperature and testicular inflammation can affect semen quality for months after the initial incident. testicular torsion, or torsion of the spermatic cord, causes rotation of the testicle, ultimately causing obstruction to venous drainage. testicular torsion often is associated with a neoplastic mass of a retained testicle within the abdomen but also can be observed with nonneoplastic testes located within the scrotum. the predominant clinical signs are pain, stiff stilted gait, and the presence of an abnormally swollen testicle (if located within the scrotum). if an intraabdominal testicular torsion is present, pain, lethargy, anorexia, and vomiting can occur (see acute condition in the abdomen). an intraabdominal mass may be palpable. perform an abdominal or testicular ultrasound, preferably with color flow doppler to evaluate perfusion to the testicle. treatment involves surgical removal of the involved testes. bacterial infections of the testicle or epididymis most commonly are caused by ascending infections of the normal bacterial flora of the prepuce or urethra. common inhabitants include escherichia coli, staphylococcus aureus, streptococcus spp., and mycobacterium canis. brucella canis and r. rickettsii are also capable of causing orchitis and epididymitis in the dog. clinical signs of orchitis or epididymitis include testicular enlargement, stiff stilted gait, and reluctance to walk. physical examination often reveals a fever and self-induced trauma to the scrotum from licking or chewing at the inflamed area. collect a semen sample by ejaculation, and culture it to identify the causative organism. alternately, collect samples by needle aspiration of the affected organ(s) and test serologically for b. canis. treatment of infectious orchitis involves a minimum of to weeks of specific antimicrobial therapy, based on culture and susceptibility testing, whenever possible. if a bacterial culture cannot be obtained, initiate fluoroquinolone therapy (enrofloxacin, mg/kg po q h). doxycycline ( mg/kg po bid for days) has been shown to suppress but not eradicate b. canis infection. testicular inflammation and increased temperature can affect sperm quality for months after infection. the most common causes of acute prostatitis are associated with acute bacterial infection (e. coli, proteus spp., pseudomonas spp., and mycoplasma spp.). less common causes include fungal infection (blastomyces dermatitidis) or anaerobic bacterial infection. acute prostatitis is characterized by fever, caudal abdominal pain, lethargy, anorexia, blood in the ejaculate, hematuria, dyschezia, and occasionally stranguria or dysuria. the patient often appears painful and depressed and may be dehydrated on physical examination. symmetric or asymmetric prostatomegaly and prostate pain may be evident on rectal palpation. in severely affected dogs, clinical signs of tachycardia, hyperemic or injected mucous membranes, bounding pulses, lethargy, dehydration, and fever may be present because of sepsis. death can occur within days if a prostatic abscess ruptures. diagnosis of acute prostatitis is confirmed based on the presenting clinical signs, neutrophilic leukocytosis (with or without a left shift), and positive urine culture results. prostatic samples may be obtained from the prostatic portion of the ejaculate, prostatic massage, urethral discharge, urine, or (less commonly) prostatic aspirate. although semen samples can yield positive bacterial cultures, dogs with acute prostatitis are often unwilling to ejaculate. radiography may reveal an enlarged prostate, but this alone does not confirm the diagnosis of prostatitis. an abdominal ultrasound often reveals prostatic abscessation and allows for the collection of samples from the affected area(s) via prostatic aspirate. aspiration of the affected tissue potentially can wick infection into periprostatic tracks. cytologic examination of the patient's ejaculate or prostatic wash from a dog with acute prostatitis reveals numerous inflammatory cells and may contain bacterial organisms. the treatment of a patient with acute prostatitis is directed at correcting dysuria and constipation associated with prostatic enlargement. enrofloxaxin ( mg/kg po sid) can penetrate the inflamed prostatic tissue and is effective in treating gram-negative and mycoplasma spp. infections. ciprofloxacin does not appear to penetrate prostatic tissue as readily. alternatives to enrofloxacin therapy are trimethoprim-sulfamethoxazole ( mg/kg po q h) or chloramphenicol ( - mg/kg po q h) for a minimum of to weeks. castration is recommended because benign prostatic hyperplasia may be a predisposing factor in the development of acute prostatitis. do not perform castration until the patient has been on antibiotic therapy for a minimum of days, to prevent the surgical complication of schirrous cords. finasteride (proscar, mg/kg po q h), an antiandrogen α-reductase inhibitor, may help reduce the size of prostatic tissue until the effects of castration are observed. if a prostatic abscess is present, perform marsupialization, surgical drainage, or ultrasonographic drainage. surgical therapy is associated with a large incidence of complications, including incontinence, chronic drainage from fistulas and stomas, septic shock, and death. fracture of the os penis is an uncommon condition encountered in male dogs. os penis fractures can occur with minimal soft tissue damage but cause hematuria and dysuria. on physical examination, urethral obstruction and crepitus in the penis are found. a lateral abdominal radiograph is usually sufficient to document the fracture. treatment consists of conservative therapy, in most cases, and consists primarily of analgesia administration. if the urethra also is damaged, place a urethral catheter for to days to allow the urethral mucosa to heal. fractures of the os penis that are comminuted or severe enough to cause urethral obstruction require open reduction and fixation, partial penile amputation, or antescrotal (prescrotal) urethrostomy. lacerations of the penis cause significant bleeding because of the extensive vascular supply to the penis. dogs and cats tend to lick penile lacerations and prevent adequate clot formation. sedation or general anesthesia often is required to evaluate and treat the laceration. after sedation or general anesthesia, place a urinary catheter and examine the penis under a stream of cold water. small lacerations can be managed with cold compresses and one to several absorbable sutures. extensive suturing usually is not required. prevent erection by isolating the patient from females in estrus or allowing excitement or excessive activity. place an elizabethan collar to prevent self-mutilation. initiate systemic antibiotic therapy to prevent infection. the inability to withdraw the penis into the prepuce in male dogs or cats is known as paraphimosis. paraphimosis usually develops following an erection in young male dogs and in emergency care older dogs after coitus. mucosal edema, hemorrhage, self-mutilation, and necrosis requiring penile amputation can occur if left untreated. treatment consists of applying cold water to the penis and reducing edema with application of an osmotic substance such as sugar. examine the base of the penis for hair rings that can prevent retraction of the penis into the prepuce. rinse the penis carefully with cold water and lubricate it with sterile lubricant and replace it into the prepuce. if the penis cannot be reduced easily into the prepuce, anesthetize the patient and make a small incision at the lateral aspect of the preputial opening. replace the penis and close the incision with absorbable suture. place a purse-string suture and leave it in place for several days to prevent recurrence. instill topical antimicrobial ointment with steroids into the prepuce several times a day. in severe cases, a urinary catheter may need to be placed to prevent urethral obstruction, until penile swelling and edema resolve. place an elizabethan collar to prevent excessive licking during the healing process. prolapse of the distal urethra is a condition usually confined to intact male english bulldogs, although isolated incidences also have been reported in yorkshire and boston terriers. the exact cause of this condition is unknown but usually is associated with a condition that causes increased intraabdominal pressure or urethral straining, including sexual excitement, coughing, vomiting, obstructed airway or brachycephalic airway syndrome, urethral calculi, genitourinary tract infection, and masturbation. the urethral prolapse usually appears as a mushroom-tip congested, irritated mass at the end of the penis that may or may not bleed (figure - ) . in some cases, bleeding occurs or worsens with sexual excitement. clinical signs associated with the prolapsed urethra include excessive licking of the prepuce, stranguria, and preputial bleeding. once the mass is observed, other differential diagnoses include transmissible venereal tumor, urethral polyp, trauma, urethritis, and neoplasia. in most cases, however, the prolapse occurs in intact young dogs, making neoplastic conditions less likely. treatment for urethral prolapse should occur at the time of diagnosis to prevent selfinduced trauma and infection. immediate therapy includes manual reduction of the prolapsed tissue and placement of a purse-string suture around an indwelling urinary catheter. the purse-string suture can remain in place for up to days until definitive repair. until the time of surgery, place an elizabethan collar on the patient to prevent self-mutilation. several forms of surgical correction have been described. in some cases, surgical resection of the prolapsed tissue with apposition of the urethral and penile mucosa can be attempted. more recently, a technique involving placement of several mattress sutures to reduce and secure the prolapsed tissue has been described. recurrence of prolapse can occur with either technique, particularly if the inciting event recurs. because there may be a genetic predisposition in this breed and because the prolapse can recur with sexual excitement, neutering should strongly be recommended. local freezing or frostbite most commonly affects the peripheral tissues of the ears, tail, paws, and genitalia that are sparsely covered with fur, are poorly vascularized, and may have been traumatized previously by cold. clinical signs of frostbite are paleness and appearance of a blanched pink to white discoloration to the skin. the skin also may appear black and necrotic. immediate treatment consists of slowly rewarming the affected area with moist heat at . ° c ( °f) or by immersion in warm water baths. analgesics may be required to alleviate patient discomfort. carefully dry the injured areas and protect them from further trauma. the use of prophylactic antibiotics is controversial because it can promote resistant bacterial infection. use of antibiotics should be based on the presence of infection. treatments that are ineffective and may be harmful include rubbing the affected areas, pressure bandages, and ointments. corticosteroids can decrease cellular immunity and promote infection and are therefore contraindicated. many frostbitten areas that appear nonviable can regain function gradually. use care when removing areas of necrotic tissue. affected areas may take several days to a week before fully manifesting areas of demarcation between healthy viable and necrotic nonviable tissue. chilling of the entire body from exposure or immersion in extremely cold water results in a decrease in core body temperature and physiologic processes that become irreversible when the body temperature falls below °c ( °f). mild hypothermia can be °to °c, moderate hypothermia from °to °c, and severe hypothermia below °c. the duration of exposure and the general condition of the animal influences its ability to survive. clinical signs and consequences associated with hypothermia include shivering, vasoconstriction, mental depression, hypotension, sinus bradycardia, hypoventilation with decreased respiratory rate, increased blood viscosity, muscle stiffness, atrial and ventricular irritability, decreased level of consciousness, decreased oxygen consumption, metabolic (lactic) acidosis, respiratory acidosis, and coagulopathies including dic. if the animal is breathing, administer warm, humidified oxygen at to breaths per minute. if the animal is not breathing or is severely hypoventilating, endotracheal intubation with mechanical ventilation may be necessary. place an intravenous catheter and infuse warmed crystalloid fluids. if the blood glucose is less than mg/dl, add supplemental dextrose ( . %) to the crystalloid fluids. monitor the core body temperature and ecg closely. rewarming should occur in the form of external circulating warm water blankets, radiant heat, and circulating warm air blankets (bair hugger). never use a heating pad, to avoid iatrogenic thermal burn injury. severe hypothermia may require core rewarming in the form of intraperitoneal fluids ( to ml/kg of lactated ringer's solution warmed to . °c [ °f]). place a temporary peritoneal dialysis catheter, and repeat the dialysis every minutes until the patient's body temperature reaches . °to . °c ( °to °f). the body temperature should rise slowly, ideally no more than °f per hour. because the response of the body to drugs is unpredictable, avoid administering drugs whenever possible, until the body temperature returns to normal. complications observed during rewarming include dic, cardiac dysrhythmias including cardiac arrest, pneumonia, pulmonary edema, cns edema, ards, and renal failure. heat stroke and heat-induced illness in dogs can be associated with excessive exertion, exposure to high environmental temperatures, stress, and other factors that cause an inability to dissipate heat. brachycephalic breeds, obesity, laryngeal paralysis, and older animals with cardiovascular disease can be particularly affected. hyperthermia is defined as a rectal temperature of °to °c ( °to °f). clinical signs of hyperthermia include congested hyperemic mucous membranes, tachycardia, and panting. more severe clinical signs include collapse (heat prostration), ataxia, vomiting, diarrhea, hypersalivation, muscle tremors, loss of consciousness, and seizures. heat-induced illness can affect all major organ systems in the body because of denaturation of cellular proteins and enzyme activities, inappropriate shunting of blood, hypotension, decreased oxygen delivery, and lactic acidosis. cardiac dysrhythmias, interstitial and intracellular dehydration, intravascular hypovolemia, central nervous dysfunction, slough of gastrointestinal mucosa, oliguria, and coagulopathies can be seen as organ function declines. excessive panting can result in respiratory alkalosis. poor tissue perfusion results in a metabolic acidosis. loss of water in excess of solutes such as sodium and chloride can lead to a free water deficit and severe hypernatremia. a marked increase in pcv occurs because of the free water loss. severe abnormalities in electrolytes and ph can lead to cerebral edema and death. treatment goals for the patient with heat-induced illness are to lower the core body temperature and support cardiovascular, respiratory, renal, gastrointestinal, neurologic, and hepatic functions. at the scene the veterinarian or caretaker can spray the animal with tepid (not cold) water. immersion in cold water or ice baths is absolutely contraindicated. cold water and ice will cause extreme peripheral vasoconstriction, inhibiting the patient's ability to dissipate heat through conductive and convective cooling mechanisms. as a result, core body temperature will continue to rise despite the good intentions of well-doers at the scene. animals that present to the veterinarian that have been cooled to the point of hypothermia have a worse prognosis. once the animal has presented to the veterinarian, the goal is to cool the animal's body temperature with towels soaked in tepid water, cool intravenous fluids, and fans until the temperature has decreased to °f. organ system monitoring and support is based on the severity and duration of the heat stroke and the ability of the body to compensate and respond to treatment. management of the patient with heat-induced illness involves prompt aggressive cooling without being overzealous and creating iatrogenic hypothermia. administer cool intravenous crystalloid fluids to replenish volume and interstitial hydration and correct the patient's acid-base and electrolyte abnormalities. management consists of rule of twenty monitoring (see rule of ), taking care to evaluate, restore, and maintain a normal cardiac rhythm, blood pressure, urine output, and mentation. administer antibiotics if there are any signs of gastrointestinal bleeding that will predispose the patient to bacterial translocation. monitor baseline chemistry tests including a complete blood count, biochemical panel, platelet count, coagulation tests, and urinalysis. treat coagulopathies including dic aggressively and promptly (see also disseminated intravascular coagulation). severe changes in mentation including stupor or coma worsen a patient's prognosis. following initial therapy, monitor the patient for a minimum of to hours for secondary organ damage, including renal failure, myoglobinuria, cerebral edema, and dic. dogs that are going to die of heat-induced illness usually die within the first hours. animals that survive longer than hours have a more favorable prognosis. immediate treatment consists of cooling the patient with cooling measures as for hyperthermia and heat-induced illness (see the previous discussion), and eliminating the cause (i.e., exertion, anesthesia, or neuromuscular blockers such as succinylcholine). if the patient is under general anesthesia, hyperventilate the patient to help eliminate carbon dioxide and respiratory acidosis. administer dantrolene sodium ( to mg/kg iv) to stabilize the sarcoplasmic reticulum and decrease its permeability to calcium. animals with malignant hyperthermia should avoid any predisposing factors, including exertion, hyperthermia, and anesthesia. after an episode of malignant hyperthermia, administer crystalloid fluids intravenously to aid in the elimination of myoglobin. monitor renal function closely for myoglobinuria and pigment damage to the renal tubular epithelium. monitor and correct acid-base and electrolyte changes. walters jm: hyperthermia. in wingfield we, editor: the veterinary icu book, jackson, wyo, , teton newmedia. sometimes it is difficult to assess whether an animal has been bitten by a poisonous or nonpoisonous snake. in colorado, the bull snake closely resembles the prairie rattlesnake. both snakes make similar noise and can be alarming if noticed on a hike or in the backyard. whenever possible, identify the offending reptile but never risk being bitten. know what types of venomous creatures are in the geographic area of the practice. if an animal has been bitten by a nonpoisonous snake, usually the bite marks are small with multiple small tooth punctures, and the bite is relatively nonpainful. usually local reaction is negligible. however, large boas or pythons also can inflict large crushing injuries that can cause severe trauma, including bony fractures. treatment for a nonpoisonous snakebite involves clipping the bite wound and carefully cleaning the area with antimicrobial scrub solution. broad-spectrum antibiotics (e.g., amoxicillin-clavulanate, . mg/kg po q h) are indicated because of the extensive bacterial flora in the mouths of snakes. monitor all snakebite victims for a minimum of hours after the incident, particularly when the species of the offending reptile is in question. if clinical signs of envenomation occur, modify the patient's treatment appropriately and aggressively. the two major groups of venomous snakes in north america are the pit viper and the coral snake. all venomous snakes are dangerous. the severity of any given bite depends on the toxicity of the venom, the amount of venom injected, the site of envenomation, the size of the animal bitten, and the time from bite/envenomation to seeking appropriate medical intervention. the majority of reptile envenomations in the united states are inflicted by pit vipers, including the water moccasin (cottonmouth), copperhead, and numerous species of rattlesnakes. pit vipers are characterized by a deep pit located between the eye and nostril, elliptic pupils, and retractable front fangs (figure - ) . localized clinical signs of pit viper envenomation may include the presence of bleeding puncture wounds, local edema close to puncture wounds, immediate severe pain or collapse, edema, petechiae, and ecchymosis with subsequent tissue necrosis. systemic signs of pit viper envenomation may include hypotension, shock, coagulopathies, lethargy, weakness, muscle fasciculations, lymphangitis, rhabdomyolysis, and neurologic signs including respiratory depression and seizures. neurologic signs largely are associated with envenomation emergency management of specific conditions by the mojave and canebrake rattlesnakes, although a potent neurotoxin, mojave toxin a, also has been identified in other subspecies of rattlesnake. clinical signs of envenomation may take several hours to appear. hospitalize all suspected victims and monitor them for a minimum of hours. the severity of envenomation cannot be judged solely on the basis of local tissue reaction. first aid measures by animal caretakers do little to prevent further envenomation. the most important aspect of initiating therapy is to transport the animal to the nearest veterinary emergency facility. to determine whether an animal has been envenomated by a pit viper, examine a peripheral blood smear for the presence of echinocytes. echinocytes will appear within minutes of envenomation and may disappear within hours. other treatment should be initiated as rapidly and aggressively as possible, although controversy exists whether some therapies are warranted. the mainstay of therapy is to improve tissue perfusion with intravenous crystalloid fluids, prevent pain with judicious use of analgesic drugs, and when necessary, reverse or negate the effects of the venom with antivenin. because pit viper venom consists of multiple fractions, treat each envenomation as a complex poisoning. obtain vascular access and administer intravenous crystalloid fluids (one fourth of a calculated shock dose) according to the patient's perfusion parameters of heart rate, blood pressure, and capillary refill time (see also shock and fluid therapy). opioid analgesics are potent and should be administered at the time of presentation. (see also pharmacologic means to analgesia: major analgesics). diphenhydramine ( . to mg/kg im or iv) also can be administered to decrease the effects of histamine. famotidine, a histamine receptor antagonist, also can be administered ( . to mg/kg iv) to work synergistically with diphenhydramine. although antihistamines have no effect on the venom per se, they may have an effect on the tissue reaction to the venom and may prevent an adverse reaction to antivenin. the use of glucocorticosteroids is controversial. glucocorticosteroids (dexamethasone sodium phosphate [dex-sp], . to . mg/kg iv) may stabilize cellular membranes and inhibit phospholipase, an active component of some pit viper toxins. polyvalent antivenin is necessary in many cases of pit viper envenomation, except in most cases of prairie rattlesnake (crotalus viridis viridis) envenomation in colorado. a recent study demonstrated no difference in outcome with or without the use of antivenin in cases of prairie rattlesnake envenomation. clinically, however, patients that receive antivenin are more comfortable and leave the hospital sooner than those that do not receive antivenin. the exact dose of antivenin is unknown in small animal patients. administer a dose of at least vial of antivenin to neutralize circulating venom. mix antivenin with a swirling, rather than a shaking motion, to prevent foaming. mix the antivenin with a -ml bag of . % saline, and then administer it slowly over a period of hours. pretreat animals with diphenhydramine ( . to mg/kg im) before the administration of antivenin, and then monitor the animal closely for clinical signs of angioneurotic edema, urticaria, tachyarrhythmias, vomiting, diarrhea, and weakness during the infusion. administration of antivenin into the bite site is relatively contraindicated and ineffective because uptake is delayed, and systemic effects are the more life-threatening. management of pit viper envenomation largely involves maintenance of normal tissue perfusion with intravenous fluids, decreasing patient discomfort with analgesia, and negating circulating venom with antivenin. hydrotherapy to the affected bite site with tepid water is often soothing to the patient. the empiric use of antibiotics is controversial but is recommended because of the favorable environment created by a snakebite (i.e., impregnation of superficial gram-positive bacteria and gram-negative bacteria from the mouth of the snake into a site of edematous necrotic tissue). administer amoxicillin-clavulanate ( . mg/kg po q h, or cephalexin, mg/kg po q h). also consider administration of nsaids (carprofen, . mg/kg po q h). monitor the patient closely for signs of local tissue necrosis and the development of thrombocytopenia and coagulopathies including dic (see management of disseminated intravascular coagulation). treat coagulopathies aggressively to prevent end-organ damage. coral snakes are characterized by brightly colored bands encircling the body, with red and black separated by yellow. "red on black, friend of jack; red on yellow, kill a fellow." types of coral snakes include the eastern coral, texas coral, and sonoran coral snakes. clinical signs of coral snake envenomation may include small puncture wounds, transient initial pain, muscle fasciculations, weakness, difficulty swallowing/dysphagia, ascending lower motor neuron paralysis, miotic pinpoint pupils, bulbar paralysis, respiratory collapse, and severe hemolysis. clinical signs may be delayed for as long as hours after the initial bite. immediate treatment with antivenin is necessary in cases of coral snake envenomation before the clinical signs become apparent, whenever possible. support respiration during paralysis with mechanical ventilation. secure the patient's airway with a cuffed endotracheal tube to prevent aspiration pneumonia. clinical signs will progress rapidly once they develop. rapid administration with antivenin is the mainstay of therapy in suspected coral snake envenomation. respiratory and cardiovascular support should occur with mechanical ventilation and intravenous crystalloid fluids. keep the patient warm and dry in a quiet place. turn the patient every to hours to prevent atelectasis and decubitus ulcer formation. maintain cleanliness using a urinary catheter and closed urinary collection system. perform passive range of motion and deep muscle massage to prevent disuse atrophy of limb muscles and function. treat aspiration pneumonia aggressively with broad-spectrum antibiotics (ampicillin, mg/kg iv q h, with enrofloxacin, mg/kg iv q h, and then change to oral once tolerated and the patient is able to swallow) for weeks past the resolution of radiographic signs of pneumonia, intravenous fluids, and nebulization with sterile saline and coupage chest physiotherapy. several weeks may elapse before a complete recovery. the adult black widow spider (latrodectus spp.) can be recognized by a red to orange hourglass-shaped marking on the underside of a globous, shiny, black abdomen. the immature female can be recognized by a colorful pattern of red, brown, and beige on the dorsal surface of the abdomen. adult and immature females are equally capable of envenomation. the male is unable to penetrate the skin because of its small size. black widow spiders are found throughout the united states and canada. black widow spider venom is neurotoxic and acts presynaptically, releasing large amounts of acetylcholine and norepinephrine. there appears to be a seasonal variation in the potency of the venom, lowest in the spring and highest in the fall. in dogs, envenomation results in hyperesthesia, muscle fasciculations, and hypertension. muscle rigidity without tenderness is characteristic. affected animals may demonstrate clinical signs of acute abdominal pain. tonic-clonic convulsions may occur but are rare. in cats, paralytic signs predominate and appear early as a ascending lower motor neuron paralysis. increased salivation, vomiting, and diarrhea may occur. serum biochemistry profiles often reveal significant elevations in creatine kinase and hypocalcemia. myoglobinemia and myoglobinuria can occur because of extreme muscle damage. management of black widow spider envenomation should be aggressive in the cat and dog, particularly when the exposure is known. in many cases, however, the diagnosis is made based on clinical signs, biochemical abnormalities, and lack of other apparent cause. antivenin (one vial) is available and should be administered after pretreatment with diphenhydramine. if antivenin is unavailable, administer a slow infusion of calcium-containing fluid such as lactated ringer's solution with calcium gluconate while carefully monitoring the patient's ecg. the small brown nonaggressive spider is characterized by a violin-shaped marking on the cephalothorax. the neck of the violin points toward the abdomen. brown spiders are found primarily in the southern half of the united states but have been documented as far north as michigan. the venom of the brown spider has a potent dermatonecrolytic effect and starts with a classic bull's-eye lesion. the lesion then develops into an indolent ulcer into dependent tissues promoted by complement fixation and influx of neutrophils into the affected area. the ulcer can take months to heal and often leaves a disfiguring scar. systemic reactions are rare but can include hemolysis, fever, thrombocytopenia, weakness, and joint pain. fatalities are possible. immediate management of an animal with brown spider envenomation is difficult because there is no specific antidote and because clinical signs may be delayed until necrosis of the skin and underlying tissues becomes apparent through the patient's fur to days after the initial bite. dapsone has been recommended at a dose of mg/kg for days. surgical excision of the ulcer may be helpful if performed in the early stages of wound appearance. glucocorticosteroids may be of some benefit if used within hours of the bite. the ulcer should be left to heal by second intention. deep ulcers should be treated with antibiotics. bufo toad species (b. marinus, aka cane toad, marine toad, giant toad; and the colorado river toad or sonoran desert toad b. alvarius) can be associated with severe cardiac and neurotoxicity if an animal licks its skin. the severity of toxicity depends largely on the size of the dog. toxins in the cane toad, b. marinus, include catecholamines and vasoactive substances (epinephrine, norepinephrine, serotonin, dopamine) and bufo toxins (bufagins, bufotoxin, and bufotenine), the mechanism of which is similar to cardiac glycosides. clinical signs can range from ptyalism, weakness, ataxia, extensor rigidity, opisthotonus, and collapse to seizures. clinical signs associated with b. alvarius toxicity are limited largely to cardiac dysrhythmias, ataxia, and salivation. the animal should have its mouth rinsed out thoroughly with tap water even before presentation to the veterinarian. if the animal is unconscious or actively seizing and cannot protect its airway, flushing the mouth is contraindicated. once an animal presents to the veterinarian, the veterinarian should place an intravenous catheter and monitor the patient's ecg and blood pressure. attempt seizure control with diazepam ( . mg/kg iv) or pentobarbital ( to mg/kg iv to effect). ventricular dysrhythmias can be controlled first with esmolol ( . mg/kg). if esmolol is ineffective, administer a longer-acting parenteral β-antagonist such as propranolol ( . mg/kg iv). ventricular tachycardia also can be treated with lidocaine ( to mg/kg iv, followed by to µg/kg/minute iv cri). case management largely depends on supportive care and treating clinical signs as they occur. monitor baseline acid-base and electrolyte balance because severe metabolic acidosis may occur that should be treated with intravenous fluids and sodium bicarbonate ( . to meq/kg iv). monitor ecg, blood pressure, and mentation changes closely. control seizures and cardiac dysrhythmias. eubig pa: bufo species intoxication: big toad, big problem, vet med ( ) lizards of the family hemodermatidae are the only two poisonous lizards in the world. they are found in the southwestern united states and mexico. the venom glands are located on either side of the lower jaw. because these lizards are typically lethargic and nonaggressive, bite wounds are rare. the lizards have grooved teeth that introduce the venom with a chewing motion as the lizard holds tenaciously to the victim. the majority of affected dogs are bitten on the upper lip, which is very painful. there are no proven first aid measures for bites from gila monsters or mexican bearded lizards. the lizard can be disengaged by inserting a prying instrument in between the jaws and pushing at the back of the mouth. the teeth of the lizard are brittle and break off in the wound. topical irrigation with lidocaine and probing with a needle will aid in finding and removing the teeth from the victim. bite wounds will bleed excessively. irrigate wounds with sterile saline or lactated ringer's solution, and place compression on the affected area until bleeding ceases. monitor the patient for hypotension. establish intravenous access, and administer intravenous fluids according to the patient's perfusion parameters. antibiotic therapy is indicated because of the bacteria in the lizard's mouth. because no antidote is available, treatment is supportive according to patient signs. the majority of musculoskeletal emergencies are the result of external trauma, most commonly from motor vehicle accidents. blunt trauma invokes injury to multiple organ systems as a rule, rather than an exception. because of this, massive musculoskeletal injuries are assigned a relatively low priority during the initial triage and treatment of a traumatized animal. perform a rapid primary survey and institute any lifesaving emergency therapies. adhere to a crash plan or the abcs of resuscitation (see initial emergency examination, management, and triage). although musculoskeletal injuries are assigned a relatively lower priority, the degree of recovery from these injuries and financial obligation for fracture repair sometimes becomes a critical factor in a client's decision whether to pursue further therapy. one of the most important deciding factors is the long-term prognosis for the patient to have a good quality of life following fracture repair. the initial management of musculoskeletal injuries is important in ensuring the best chance for maximal recovery with minimal complications after definitive surgical fracture repair. this is particularly important for open fractures, spinal cord compromise, multiple fractures, open joints, articular fractures, physeal fractures, and concomitant ligamentous or neurologic compromise (box - ). immediately after the initial primary survey of a patient, perform a more thorough examination, including an orthopedic examination. multiple injuries often are observed in the patient that falls from height (e.g., "high-rise syndrome"), motor vehicle accidents, gunshot wounds, and encounters with other animals (e.g., "big-dog-little-dog"). address the most life threatening injuries, and palliate musculoskeletal injuries until more definitive repair can be attempted when the patient is more stable. in animals with the history of potential for multiple injuries, search thoroughly and meticulously for areas of injury to the spinal column, extremities, and for small puncture wounds. helpful signs that can provide a clue as to an underlying injury include swelling, bruising, abnormal motion, and crepitus (caused by subcutaneous emphysema or bony fracture). if the patient is alert, look for areas of tenderness or pain. in unconscious or depressed patients, reexamine the patient after the patient becomes more mentally alert. injuries often are missed during the initial examination in obtunded patients because of the early response and attenuation of pain. unconscious or immobile patients must have radiographic examination of the spinal column following stabilization and support. palpate the skull carefully for obvious depressions or crepitus that may be associated with a skull fracture. localization of the injury can be determined by motion in abnormal locations, swelling caused by hemorrhage or edema, pain during gentle movement or palpation, deformity, angular change, or a significant increase or decrease in normal range of motion of bones and joints. perform a rectal examination in all cases to palpate for pelvic fractures and displacement. once the diagnosis of a fracture or luxation has been confirmed, look for any evidence of skin lacerations or punctures near the fracture site. in long-haired breeds, clipping the fur near the fracture site often is necessary to perform a thorough examination of the area. if any wounds are found, the fracture is classified as an open fracture until proven otherwise. in some cases, the open fracture is obvious, with a large section of bone fragment protruding through the skin. in other cases, the puncture wound may be subtle, with only a small amount of blood or pinpoint hole in the skin surface. characteristics observed with open fractures include bone penetration, fat droplets or marrow elements in blood coming from the wound, subcutaneous emphysema on radiographs, and lacerations in the area of a fracture. protect the patient from further injury or contamination of wounds. excessive palpation to intentionally produce crepitus is inappropriate because it causes severe patient discomfort and has the potential to cause severe soft tissue and neurologic injury at the fracture site. sedation and analgesia aids in making the examination more comfortable for the patient and allows localization of the injury and comparison with the opposite extremity. higher-quality radiographs can be performed to determine the extent of the injury when the animal is sedated adequately and pain is controlled. sedate the patient judiciously with analgesic drugs. opioid drugs work well for orthopedic pain, produce minimal cardiorespiratory depression, and can be reversed with naloxone if necessary. handle the fracture site gently to avoid causing further pain and soft tissue injury at the fracture site. rough or careless handling of a fracture site can cause a closed fracture to penetrate through the skin and become an open fracture. cover open fractures immediately to prevent contamination of the fracture with nosocomial infection from the hospital. administer a first-generation cephalosporin (cephalexin, mg/kg po q h, or cefazolin, mg/kg iv q h). the bandage also serves to control hemorrhage and prevent desiccation of the bones and surrounding soft tissue structures. leave the initial bandages in place until the patient's cardiorespiratory status has been determined to be stable and more definitive wound management can occur in a clean, preferably sterile location. examine the neurologic status and cardiovascular status of the limb before and after treatment. determine the vascular status of the limb by checking the color and temperature of the limb, the state of distal pulses, and the degree of bleeding from a cut nail bed. in patients with severe cardiovascular compromise and hypotension caused by hemorrhagic shock, the viability of the limb may be in question until the cardiovascular status and blood pressure are normalized. reduction of the fracture or straightening of gross deformities may return normal vascularity to the limb. when checking neurologic status, examine for motor and sensory function to the limb. swelling may increase pressure on the nerves as they run through osteofascial compartments, resulting in decreased sensory or motor function, or neurapraxia. diminished function often returns to normal once the swelling subsides. serial physical examinations in the patient and response to initial stabilization therapy can lead to a higher index of suspicion that more occult injuries are present, such as a diaphragmatic hernia, perforated bowel, lacerated liver or spleen, or uroabdomen. to prevent ongoing trauma, reduce any fracture and then stabilize the site above and below the fracture. a modified robert jones splint or bandage often works well for fractures emergency management of specific conditions involving the distal extremities. fractures of the humerus or femur are difficult to immobilize without the use of spica or over-the-hip coaptation splints to prevent mobility. inappropriate bandaging of humerus or femur fractures can result in a fulcrum effect and worsen the soft tissue and neurologic injuries. further displacement of vertebral bodies or luxations can cause cord compression or laceration such that return to function becomes impossible. immediately place any patient with a suspected spinal injury on a flat surface, and tape down the animal to prevent further movement until the spine has been cleared by a minimum or two orthogonal radiographic views (lateral and ventrodorsal views performed as a cross-table x-ray technique). wounds associated with musculoskeletal trauma are common and include injury to the bones, joints, tendons, and surrounding musculature (box - ). major problems associated with these cases are the presence of soft tissue trauma that makes wound closure hazardous or impossible, because of the risk of infection. chronic deep infection of traumatized wounds can cause delayed healing and sequestrum to develop, particularly if there is avascular bone or cartilage within the wound. in the early management of an open fracture, the areas should be splinted without pulling any exposed bone back into the soft tissue. the wound should not be probed or soaked, as nosocomial bacteria and other external contaminants can be introduced into the wound, leading to severe infection. because of the risk of actually causing infection, probing, flushing, or replacing tissues back into the wound should be performed at the time of formal debridement when the patient is physiologically stable. immediate bactericidal antibiotic therapy with a first-generation cephalosporin should be started immediately to obtain adequate concentrations of antibiotics at the fracture site. the duration of antibiotic therapy should ideally be limited to - days to prevent the risk of superinfection. treatment of open musculoskeletal injury involves three considerations: initial inspection and wound debridement, stabilization and repair, and wound bandaging. emergency care when associated with a fracture, wound is created from the inside out by penetration of bone fragments through the skin or from a low-energy gunshot. simple or comminuted fracture pattern good stability of the two main bone segments treatment and prognosis are good and similar to those of a closed injury if wound is debrided and stabilized within to hours. when associated with a fracture, wound is created from the outside in. major deep injury with considerable soft tissue stripping from bone and muscle damage simple or comminuted fracture pattern prognosis is good if wound is debrided within hours of injury and provided rigid stabilization with a bone plate or external fixator. results from major external force severe damage and necrosis of skin, subcutaneous tissue, muscle, nerve, bone, tendon, and arteries soft tissue damage may vary from crush injury to shearing injury associated with bite wounds or low-speed automobile accidents. requires immediate and delayed sequential debridement and rigid external fixation can require prolonged healing times guarded prognosis initial inspection and wound debridement include the following steps: . after the patient's cardiovascular status has been stabilized and it has been determined that it can withstand anesthesia, place the animal under general anesthesia and remove the temporary splint. . keeping the wound covered, shave the surrounding fur. . remove the covering and then place sterile lubricant jelly over the wound. shave the fur to the edges of the wound margin. . wash away any entrapped fur and the lubricant jelly. . complete an antiseptic scrub of the surrounding skin. . if the wound is a small puncture (e.g., gunshot pellets or bites), probe the wound with a sterile hemostat. do a thorough debridement if tissues deep to the hole are cavitated. if not deep, create a hole for drainage. . flush the wound with a physiologic solution (lactated ringer's solution is preferred). . debride the wound from outward to inward. cut away damaged areas of skin and deeper tissues to open up underlying cavitations and tissue injury. . continuously irrigate with warm physiologic solution (lactated ringer's solution is preferred). the stream must be strong enough to flush debris out of the bottom of the wound. to accomplish this, attach a -gauge needle to a -ml syringe (will deliver psi). excise any obviously devitalized tissue. . do not remove any bone fragments that are firmly attached to soft tissue. do not cut into healthy soft tissue to find bullet or bone fragments, unless the bullet can cause injury to joints or nerve tissue. . do a primary repair of tendons and nerves if the wound is type i and recent (within hours of the initial injury). if the wound is too severe or if there is obvious infection, tag the ends of the tendons and nerves for later repair. it is best to stabilize and repair open fractures as soon as the patient's cardiovascular and respiratory status can tolerate general anesthesia, provided that adequate stabilization is possible. if this is not possible because of the level of experience of the surgeon or the lack of necessary equipment, it is best to perform wound management and place a temporary splint until definitive repair can be performed. wound bandaging is discussed in the section on bandaging techniques. structural injuries to the joints are common and can involve both ligaments and articular cartilage injuries. cartilage does not heal well; therefore, injuries involving articular cartilage can lead to a significant loss of function and degenerative joint disease (osteoarthritis). cartilage injuries that are superficial evoke a short-lived enzymatic and metabolic response that does not stimulate enough cellular growth to repair the defect. superficial lesions remain as defects but do not progress to chondromalacia or osteoarthritis. deep cartilage lacerations that extend to subchondral bone produce an exuberant healing response from the cells of the underlying cartilage. in many cases, this material undergoes degeneration and leads to osteoarthritis. impact injuries to surface cartilage can cause chondrocyte and underlying bone injury. these lesions rapidly progress to osteoarthritis; however, they may be totally or partially reversible. treatment of grade i injuries requires short-term coaptation splints and has a good prognosis. grade ii injuries require surgical treatment with a suture stent and consistent postoperative coaptation splints to heal and maintain good function. healing of grade iii injuries often is a problem, and suture stents or surgical reapproximation may be indicated. failure to immobilize joints that are frequently flexed (elbow and stifle) can result in late complications of ligament repair. ligamentous injuries of joints, particularly the collateral ligaments of the stifle, elbow, and hock, and carpal hyperextension injuries are commonly missed and may require surgical fixation, including arthrodesis (box - ). fractures in immature animals differ from those in adults in that young puppies and kittens have a great ability to remodel bone. remodeling is dependent on the age of the patient and the location of the fracture. the younger the puppy or kitten and the closer the fracture to the epiphysis or growth plate, the greater the potential for remodeling and the development of angular limb deformities. remodeling occurs more effectively in longlimbed breeds of dogs than in short-limbed breeds. fractures through the growth plate of immature animals may potentially cause angular limb deformities, joint dislocations or incongruity, and osteoarthritis. this form of injury is commonly observed in the distal ulnar growth plate and the proximal and distal radial growth plates. high-rise syndrome in cats is seen in cats that fall from a height usually greater than feet. it occurs most frequently in high-rise buildings in urban areas where cats lie on window ledges and suddenly fall out the window. the most common lesions observed in cats that fall from heights are thoracic injuries (rib and sternal fractures, pneumothorax, and pulmonary contusions) and facial and oral trauma (lip avulsions, mandibular symphyseal fractures, fractures of the hard palate, and maxillary fractures). limb and spinal cord fractures and luxations, radius and ulna fractures, abdominal trauma, urinary tract trauma, and diaphragmatic hernias are also common. the injuries sustained are often found in combination, rather than as an isolated injury of one area of the body. follow the mnemonic a crash plan when managing a cat suffering from high-rise syndrome, treating the animal immediately for shock. following cardiovascular and respiratory stabilization, evaluate thoracic and abdominal radiographs, including those of the spine. evaluate the bladder closely, making sure that the cat is able to urinate effectively. examine the hard palate, maxilla, and mandibular symphysis for fractures. palpate the pelvis and carefully manipulate all limbs to examine for fractures or ligamentous injuries. finally, perform a complete neurologic examination. patients that fall less than five stories often have a more guarded prognosis than patients that fall from higher levels. sometimes the owner witnesses the ingestion of a foreign body during play, such as throwing a stick or fetching a ball. cats tend to play with string or thread that becomes caught around the base of the tongue. in many cases, however, ingestion of the foreign object is not witnessed, and diagnosis is made based on clinical signs and physical examination. foreign bodies lodged in the oral cavity often cause irritation and discomfort, including difficulty breathing and difficulty swallowing. often, an animal paws at its mouth in an attempt to dislodge a stick or bones wedged across the roof of the mouth. irritation, inability to close the mouth, and blockage of the orpharynx can result in excessive drooling. the saliva may appear blood-tinged due to concurrent soft tissue trauma (figs - and - ) . obstruction of the glottis by a foreign body (e.g., tennis ball or toy) can result in cyanosis secondary to an obstructed airway and hypoxemia. in many cases, the object is small enough to enter the larynx but too large to be expelled. if a foreign object is lodged in the mouth for more than several days, halitosis and purulent discharge may be present. many animals are anxious at the time of presentation and may require sedation or a light plane of anesthesia to remove the foreign object. the animal may bite personnel and may have bitten the owner during his or her attempt to remove the object from the mouth en route to the hospital. propofol ( mg/kg iv) or a combination of propofol with diazepam ( . - mg/kg iv) is an excellent combination for a light plane of anesthesia. exercise caution when anesthetizing a patient with a ball lodged in the airway, as further compromise of respiratory function may occur and cause worsening of the hypoxemia. before inducing anesthesia, assemble all supplies necessary to remove the object. make sure that rigid towel clamps, sponge forceps, and bone forceps are on hand, because the foreign object is often very slippery with saliva. hemostats and carmalts may slip and not be useful in the removal of the foreign object. place a peripheral intravenous catheter to secure vascular access prior to anesthetic induction. have available the supplies necessary for an emergency tracheostomy, if the foreign object cannot be removed by usual methods. induce a light plane of anesthesia and then grasp the object with the sponge forceps or towel clamps, and extract. monitor the cardiorespiratory status of the animal at all times during the extraction process. if you are unable to remove the object, and if severe respiratory distress, including cyanosis, bradycardia, or ventricular dysrhythmias, develop, perform a tracheostomy distal to the site of obstruction. once the foreign body has been removed, administer supplemental flow-by oxygen until the animal awakens. if laryngeal edema or stridor on inspiration is present, administer a dose of dexamethasone sodium phosphate ( . mg/kg iv, im, sq) to decrease inflammation. the patient should be carefully monitored for hours, because noncardiogenic pulmonary edema can develop secondary to airway obstruction. esophageal foreign bodies pose a serious medical emergency. it is helpful if the owner witnessed ingestion of the object and noted rapid onset of clinical signs. in many cases, however, ingestion is not witnessed, and the diagnosis must be made based on clinical signs, thoracic radiographs, and results of a barium swallow. the most common clinical signs are excessive salivation with drooling, gulping, and regurgitation after eating. many animals will make repeated swallowing motions. some animals exhibit a rigid "sawhorse" stance, with reluctance to move immediately after foreign body ingestion and esophageal entrapment. after completing a physical examination, evaluate cervical and thoracic radiographs to determine the location of the esophageal obstruction. esophageal foreign objects are lodged most commonly at the base of the heart, the carina, or just orad to the lower esophageal sphincter. if the object has been lodged for several days, pleural effusion and pneumomediastinum may be present secondary to esophageal perforation. endoscopy is useful for both diagnosis and removal of the foreign object; however, it is invasive and requires general anesthesia ( fig. - ) . remove foreign objects lodged in the esophagus with a rigid or flexible endoscope after the patient has been placed under general anesthesia. evaluate the integrity of the esophagus both before and after removal of the material because focal perforation or pressure necrosis can be present. necrosis of the mucosa and submucosa of the esophagus often leads to stricture formation or perforation. attempt to retrieve the object with a flexible fiberoptic endoscope if available. rigid tube endoscopy can also be performed. in many cases, smooth objects that cannot be easily grasped can be pushed into the stomach and allowed to dissolve or may be removed by gastrotomy. if the foreign body is firmly lodged in the esophagus and cannot be pulled or pushed into the stomach, or if perforation has already occurred, the prognosis for return to function without strictures is not favorable. in such cases, referral to a surgical specialist is recommended for esophagostomy or esophageal resection. after removal of the object, carefully examine the esophagus and then administer gastroprotectant agents (famotidine, . mg/kg po bid; sucralfate slurry, . - . g/dog) for a minimum of to days. to rest the esophagus, the patient should receive nothing per os (npo) for to hours. if esophageal irritation or erosion is moderate to severe, a percutaneous gastrotomy tube should be placed for feeding until the esophagus heals. perform repeat endoscopy every days to evaluate the healing process and to determine whether stricture formation is occurring. persistent vomiting immediately or soon after eating is often associated with a gastric foreign body. in some cases, the owner knows that the patient has ingested a foreign body of some kind. in other cases, continued vomiting despite lack of response to conservative treatment (npo, antiemetics, gastroprotectant drugs) prompts further diagnostic procedures, including abdominal radiographs and bloodwork. obstruction to gastric outflow and vomiting of hydrochloric acid often cause a hypochloremic metabolic acidosis. radiopaque gastric foreign bodies may be observed on plain films. radiolucent cloth material may require a barium series to delineate the shape and location of the foreign body ( fig. - ) . treatment consists of removal with flexible endoscopy or a simple gastrotomy. most animals with uncomplicated gastric foreign bodies are relatively healthy, but any metabolic and electrolyte abnormalities should be corrected prior to anesthesia and surgery. small intestinal obstruction can be caused by foreign bodies, tumors, intussusception, volvulus, or strangulation within hernias. regardless of the cause, clinical signs of small intestinal obstruction depend on the location and degree of obstruction, and whether the bowel has perforated. clinical signs associated with a high small intestinal obstruction are usually more severe and more rapid in onset compared with partial or complete obstruction of the jejunum or ileum. complete obstructions that allow no fluid or chyme to pass are worse than partial obstructions, which can cause intermittent clinical signs interspersed with periods of normality (table - ). the most common clinical signs associated with a complete small intestinal obstruction are anorexia, vomiting, lethargy, depression, dehydration, and sometimes abdominal pain. early clinical signs may be limited to anorexia and depression, making a diagnosis challenging unless the owner has a suspicion that the animal ingested some kind of foreign object. obstructions cranial to the common bile duct and pancreatic papillae lead to vomiting of gastric contents, namely hydrochloric acid, and a hypochloremic metabolic alkalosis. obstructions caudal to the common bile duct and pancreatic papillae result in loss of other electrolytes and sometimes mixed acid-base disorders. eventually, all animals with small intestinal obstruction vomit and have fluid loss into dilated segments of bowel, leading to dehydration and electrolyte abnormalities. increased luminal pressure causes decreased lymphatic drainage and bowel edema. the bowel wall eventually becomes ischemic and may rupture. linear foreign bodies should be suspected in any vomiting patient, particularly cats. string or thread often is looped around the base of the tongue and can be visualized in many cases by a thorough oral examination. to look properly under the tongue, grasp the top of the animal's head with one hand, and pull the lower jaw open with the index finger of the opposite hand while pushing up the thumb simultaneously on the tongue in between the intermandibular space. thread and string can be observed lying along the ventral aspect of the tongue. in some cases, if a linear foreign body is lodged very caudally, it cannot be visualized without heavy sedation or anesthesia. linear foreign bodies eventually cause bowel obstruction and perforation of the intestines along the mesenteric border. the foreign material (e.g., string, thread, cloth, pantyhose) becomes lodged proximally, and the intestines become plicated as the body attempts to push the material caudally through the intestines ( fig. - ) . continued peristalsis eventually causes a sawing motion of the material and perforation of the mesenteric border of the intestines. once peritonitis occurs, the prognosis is less favorable unless prompt and aggressive treatment is initiated. reevaluate any patient that does not respond to conservative symptomatic therapy, performing a complete blood count, serum biochemical panel (including electrolytes), and abdominal radiographs. intestinal masses may be palpable on physical examination and are often associated with signs of discomfort or pain when palpating over the mass. radiography and abdominal ultrasound are the most useful diagnostic aids. plain radiographs may be diagnostic when the foreign object is radiodense or there is characteristic dilation or plication of bowel loops. as a rule of thumb, the width of a loop of small bowel should be no larger than twice the width of a rib. diagnosis of small intestinal obstruction or ileus can be based on the appearance of stacking loops of dilated bowel. comparison of the width of the bowel with the width of a rib is often performed. with mild dilation, the bowel width is three to four times the rib width; with extensive dilation, five to six times the rib width ( fig. - ) . in cases of linear foreign bodies, c-areas (comma-shaped areas) of gas trapped in the plicated bowel will appear stacked on one another. blunt, wedge-shaped areas of gas or square linear areas of gas adjacent to a distended bowel loop are characteristic of a foreign body lodged in the intestine. contrast radiography is indicated when confirmation of the suspected diagnosis is necessary and ultrasonography is not available. contrast material may outline the object or abruptly stop orad to the obstruction. the definitive treatment of any type of small intestinal foreign body is surgical removal. linear foreign bodies sometimes pass, but they should never be left untreated in a patient that is demonstrating clinical signs of inappetence, vomiting, lethargy, and dehydration. the timing of surgery is critical because the risk of intestinal perforation increases with time. prior to surgery, correct any acid-base and electrolyte abnormalities with intravenous fluid therapy. administer broad-spectrum antibiotics. perform an enterotomy or intestinal resection and anastomosis as soon as possible once the patient's acid-base and electrolyte status have been corrected. clinical signs of a foreign body in the large bowel are usually nonexistent. in most cases, if a foreign object has passed successfully through the small bowel, it will pass through the large bowel without incident unless bowel perforation and peritonitis occur. penetrating foreign bodies such as needles often cause localized or generalized peritonitis, abdominal pain, and fever. hematochezia may be present if the foreign object causes abrasion of the rectal mucosa. symptomatic patients should have abdominal radiographs performed. colonoscopy or exploratory laparotomy should be performed if survey radiographs are suggestive of a large intestinal obstruction or perforation. in most cases, large intestinal foreign bodies will pass without incident. surgery is required to treat perforations, peritonitis, or abscesses. emergency care figure - : after minutes, the barium has stopped moving and has reached a blunt, intraluminal intestinal foreign body. note that barium appears wedge-shaped or square at the site of the foreign body. foreign bodies in the rectum and anus often are the result of ingestion of bones, wood material, needles, and thread, or malicious external insertion. often the material can pass through the entire gastrointestinal tract and then get stuck in the anal ring. clinical signs include hematochezia and dyschezia with straining to defecate. diagnosis is made by visual examination of the item in the anus, or by careful digital palpation after heavy sedation or short-acting general anesthesia. radiography is helpful in locating needles that have penetrated the rectum and lodged in the perirectal or perinatal tissues. treatment consists of careful removal of the needle digitally or surgically. intussusception is the acute invagination of one segment of bowel (the intussusceptum) into another (the intussuscipiens). the proximal segment always invaginates into the distal segment of bowel. intussusception most commonly occurs in puppies and kittens less than year of age but can occur in an animal of any age with hypermotility of the small bowel, gastrointestinal parasites, and severe viral or bacterial enteritis. intussusception occurs primarily in the small bowel in the jejunum, ileum, and ileocolic junction. clinical signs include vomiting, abdominal discomfort, and hemorrhagic diarrhea. usually, hemorrhagic diarrhea is the first noticeable sign, and in puppies, may be due to parvoviral enteritis, with secondary intussusception. usually, the obstruction is partial with mild clinical signs. more serious clinical signs develop as the obstruction becomes more complete. differential diagnoses include hemorrhagic gastroenteritis, parvoviral enteritis, gastrointestinal parasites, intestinal foreign body, bacterial enteritis, and other causes of vomiting and diarrhea. the diagnosis of intussusception is often made based on palpation of a sausage-shaped firm, tubular structure in the abdomen accompanied by clinical signs and abdominal pain. plain radiographs may demonstrate segmental or generalized dilated segments of bowel, depending on the duration of the problem. ultrasonographs of the palpable mass resemble the layers of an onion, with hyperechoic intestinal walls separated by less echogenic edema. treatment consists of correction of the patient's acid-base and electrolyte abnormalities with intravenous fluids and surgical reduction or removal of the intussusception with resection and anastomosis. although enteroplication has been suggested, the technique has fallen out of favor because of the increased risk of later obstruction. the primary cause of intestinal inflammation and hypermotility must be identified and corrected. gastric dilatation can occur with or without volvulus in the dog. gastric dilatationvolvulus (gdv) occurs primarily in large-and giant-breed dogs with deep chests, such as the great dane, labrador retriever, saint bernard, german shepherd dog, gordon and irish setters, standard poodle, bernese mountain dog, and bassett hound. the risk of gdv increases with age; however, it can be seen in dogs as young as months. deep, narrow-chested breeds are more likely to develop gdv than dogs with broader chests. the overall mortality for surgically treated gastric dilatation-volvulus ranges from % to %, with most deaths occurring in patients that required splenectomy and partial gastrectomy. clinical signs of gdv include abdominal distention, unproductive vomiting or retching, lethargy, weakness, sometimes straining to defecate, and collapse. the owner may think that the animal is vomiting productively because of the white foamy froth (saliva) that is not able to pass into the twisted stomach. in some cases, there is a history of the dog's being fed a large meal or consuming a large quantity of water prior to the onset of clinical signs. instruct the owner of any patient with a predisposition for and clinical signs of gdv to transport the animal to the nearest veterinary facility immediately. physical examination often reveals a distended abdomen with a tympanic area on auscultation. in dogs with very deep chests, it may be difficult to appreciate abdominal distention if the stomach is tucked up under the rib cage. depending on the stage of shock, the patient may have sinus tachycardia with bounding pulses, cardiac dysrhythmias with pulse deficits, or bradycardia. the mucous membranes may appear red and injected or pale with a prolonged capillary refill time. the patient may appear anxious and attempt to retch unproductively. if the patient is nonambulatory at the time of presentation, the prognosis is more guarded. the definitive diagnosis of gdv is based on clinical signs, physical examination findings, and radiographic appearance of gas distention of the gastric fundus with dorsocranial displacement of the pylorus and duodenum (the so-called "double-bubble" or "popeye arm" sign) ( fig. - ) . in simple gastric dilatation without volvulus, there is gas distention of the stomach with anatomy appearing normal on radiography. with "food bloat," or gastric distention from overconsumption of food, ingesta is visible in the distended stomach ( fig. - ) . as soon as a patient presents with a possible gdv, place a large-bore intravenous catheter in the cephalic vein(s) and assess the patient's ecg, blood pressure, heart rate, capillary refill time, and respiratory function. obtain blood samples for a complete blood count, serum biochemistry profile, immediate lactate measurement, and coagulation tests before taking any radiographs. rapidly infuse a colloid (hetastarch or oxyglobin, ml/kg iv bolus) along with shock volumes of a crystalloid fluid (up to ml/kg/hour) (see section on shock). monitor perfusion parameters (heart rate, blood pressure, capillary refill time, and ecg) and titrate fluid therapy according to the patient's response. the use of short-acting glucocorticosteroids is controversial. glucocorticosteroids may help stabilize cellular membranes and decrease the mechanisms of ischemia-reperfusion injury, but no detailed studies have proved them to be beneficial versus not using glucocorticosteroids in the patient with gdv. attempt gastric decompression, either with placement of an orogastric tube or by trocharization. to place an orogastric tube, position the distal end of the tube at the level of the patient's last rib ( fig. - ) and place it adjacent to the animal's thorax; then put a piece of tape around the tube where it comes out of the mouth, once it is in place. put a roll of -inch tape in the patient's mouth behind the canine teeth and then secure the roll in place by taping the mouth closed around the roll of tape. lubricate the tube with lubricating jelly and slowly insert the tube through the center of the roll of tape into the stomach. the passing of the tube does not rule out volvulus. in some cases, the front legs of the patient need to be elevated, and the caudal aspect of the patient lowered (front legs standing on a table with back legs on the ground) to allow gravity to pull the stomach down to allow the tube to pass. once the tube has been passed, air within the stomach is relieved, and the stomach can be lavaged. the presence of gastric mucosa or blood in the efflux from the tube makes the prognosis more guarded. if an orogastric tube cannot be passed, clip and aseptically scrub the patient's lateral abdomen and then insert -gauge over-the-needle catheter. "pinging" the animal's side with simultaneous auscultation allows determination of the location that is most tympanic-that is, the proper location for catheter insertion. once intravenous fluids have been started in the animal, take a right lateral abdominal radiograph to document gdv. if no volvulus is present, the owner may elect for more conservative care, and the animal should be monitored in the hospital for a minimum of hours. because some cases of gdv intermittently twist and untwist, the owner should be cautioned that although the stomach is not twisted at that moment, a volvulus can occur at any time. if radiographs demonstrate food bloat, induce emesis (apomorphine, . mg/kg iv) or perform orogastric lavage under general anesthesia. documentation of gastric dilatation-volvulus constitutes a surgical emergency. figure - : example of "food bloat" with severe gastric distention caused by overconsump-following diagnosis of gdv, continue administration of intravenous fluids. serum lactate measurements greater than . mmol/l are associated with an increased risk of gastric necrosis, requirement for partial gastrectomy, and increased mortality. administer fresh frozen plasma ( ml/kg) to patients with thrombocytopenia or prolonged pt, activated partial thromboplastin time (aptt), or activated clotting time (act). cardiac dysrhythmias, particularly ventricular dysrhythmias, are common in cases of gdv and are thought to occur secondary to ischemia and proinflammatory cytokines released during volvulus and reperfusion. lidocaine ( - mg/kg followed by mcg/kg/minute iv cri) can be used to treat cardiac dysrhythmias preemptively that are associated with ischemia-reperfusion injury, or administration can be started when ventricular dysrhythmias are present. correct any electrolyte abnormalities, including hypokalemia and hypomagnesemia. the use of nonsteroidal antiinflammatory drugs (flunixin meglumine, carprofen, ketoprofen) that can potentially decrease renal perfusion and predispose to gastric ulcers is absolutely contraindicated. administer analgesic drugs (fentanyl, µ/kg iv bolus, followed by - µ/kg/hour iv cri; or hydromorphone, . mg/kg iv) before anesthetic induction. after carrying out a balanced anesthesia protocol, the patient should be taken immediately to surgery for gastric derotation and gastropexy. postoperatively, assess the patient's ecg, blood pressure, platelet count, coagulation parameters, and gastric function (see section on rule of twenty). if no resection is required, the animal can be given small amounts of water beginning hours after surgery. depending on the severity of the patient's condition, small amounts of a bland diet can be offered to hours postoperatively. continute supportive care with analgesia and crystalloid fluids until the patient is able to tolerate oral analgesic drugs (tramadol, - mg/kg po q - h). once the patient is ambulatory and able to eat and drink on its own, it can be released from the hospital; instruct the owner to feed the animal multiple small meals throughout the day for the first week. when the intestines twist around the root of the mesentery, a small intestinal or mesenteric volvulus occurs. the problem is most common in the young german shepherd dog, although it has been observed in other large and giant breeds. predisposing factors include pancreatic atrophy, gastrointestinal disease, trauma, and splenectomy. clinical signs of mesenteric volvulus include vomiting, hemorrhagic diarrhea, bowel distention, acute onset of clinical signs of shock, abdominal pain, brick-red mucous membranes (septicemia), and sudden death. diagnosis is based on an index of suspicion and the presence of clinical signs in a predisposed breed. plain radiographs often reveal grossly distended loops of bowel in a palisade gas pattern. in some dogs, multiple, tear-drop-shaped, gas-filled loops appear to rise from a focal point in the abdomen. usually, massive distention of the entire small bowel is observed ( fig. - ) . the presence of pneumoperitoneum or lack of abdominal detail secondary to the presence of abdominal fluid is characteristic of bowel perforation and peritonitis. in a patient with mesenteric volvulus, immediate aggressive action is necessary for the animal to have any chance of survival. treatment consists of massive volumes of iv crystalloid and colloid fluids (see section on iv therapy), broad-spectrum antibiotics (ampicillin, mg/kg iv qid, with enrofloxacin, mg/kg iv once daily), and surgical correction of the bowel. because of the massive release of proinflammatory cytokines, bacterial translocation, and ischemia, treatment for shock is of paramount importance (see sections on rule of twenty and shock). prognosis for any patient with mesenteric volvulus is poor. obstipation (obstructive constipation) is most common in the older cat. in cases of simple constipation, rehydrating the animal with intravenous fluids and stool softeners is often volvulus. this consistutes an immediate surgical emergency, and the prognosis is often poor. this condition is most common in young german shepherd dogs, but can be observed in any breed. sufficient for it to regain the ability to have a bowel movement. obstipation, however, is caused by adynamic ileus of the large bowel that eventually leads to megacolon. affected cats usually are anorectic, lethargic, and extremely dehydrated. treatment consists of rehydration with intravenous crystalloid fluids, correction of electrolyte abnormalities, enemas, and promotility agents such as cisapride ( . mg/kg po q - h). the use of phosphate enemas in cats is absolutely contraindicated because of the risk of causing acute, fatal hyperphosphatemia. in many cases, the patient should be placed under general anesthesia and manual deobstipation is performed with warm water soapy enemas and a gloved finger to relieve and disimpact the rectum. stool softeners such as lactulose and docusate stool sofener (dss) may also be used. predisposing causes of obstipation such as narrowing of the pelvic canal, perineal hernia, and tumors should be ruled out. adenocarcinoma is the most common neoplasm of the gastrointestinal tract that causes partial to complete obstruction. adenocarcinomas tend to be annular and constricting, and they may cause progressive obstruction of the lumen of the small or large bowel. siamese cats tend to have adenocarcinomas in the small intestine, whereas in dogs, the tumor tends to occur in the large intestine. clinical signs of adenocarcinoma are both acute and chronic and consist of anorexia, weight loss, and progressive vomiting that occur over weeks to months. effusion may be present if metastasis to peritoneal surfaces has occurred. diagnosis is based on clinical signs and physical examination findings of a palpable abdominal mass, radiographic evidence of an abdominal mass and small or large intestinal obstruction, or ultrasonographic evidence of an intestinal mass. treatment consists of surgical resection of the affected bowel segment. the prognosis for long-term survival ( - months) is good if the mass is completely resected and if other clinical signs of cachexia or metastasis are observed at the time of diagnosis. median survival is to weeks if metastasis to lymph nodes, liver, or the peritoneum are absent at the time of diagnosis. in dogs, the prognosis is more guarded. leiomyoma and leiomyosarcoma are tumors that can cause partial or complete obstruction of the bowel. clinical signs are often referred to progressive anemia, including weakness, lethargy, inappetence, and melena. hypoglycemia can be observed as a paraneoplastic syndrome, or due to sepsis and peritonitis secondary to bowel perforation. leiomyomas are most commonly observed at the ceco-colic junction or in the cecum. surgical resection and anastomosis is usually curative, and has a favorable prognosis. incarceration of a loop of bowel into congenital or acquired defects in the body wall can cause small bowel obstruction. pregnant females and young animals with congenital hernias are most at risk. rarely, older animals with perineal hernias and animals of any age with traumatic hernias can be affected. clinical signs are consistent with a small intestinal obstruction: anorexia, vomiting, lethargy, abdominal pain, and weakness. diagnosis is often made based on physical examination of a reducible or nonreducible mass in the body wall. hernias whose contents are reducible are usually asymptomatic. treatment consists of supportive care and rehydration, administration of broad-spectrum antibiotics, and surgical correction of the body wall hernia. in some cases, intestinal resection and anastomosis of the affected area is necessary when bowel ischemia occurs. the potential for bowel perforation should be suspected whenever there is any penetrating injury (knife, gunshot wound, bite wound, stick impalement) of the abdomen. injuries that result in bowel ischemia and rupture can also occur secondary to nonpenetrating blunt emergency care trauma or shear forces (e.g., big dog-little dog/cat). perforation of the stomach and small and large intestines can occur with use of nonsteroidal antiinflammatory drugs. diagnosis of bowel perforation first depends on the alertness to the possibility that the bowel may have been perforated or penetrated. as a general rule, all penetrating injuries of the abdomen should be investigated by exploratory laparotomy. diagnostic peritoneal lavage (dpl) can be performed; however, early after penetrating injury of the bowel, dpl may be negative or nondiagnostic until peritonitis develops. whenever any patient with blunt or penetrating abdominal trauma does not respond to initial fluid therapy, or responds and then deteriorates, the index of suspicion for bowel injury should be raised. the findings of pneumoperitoneum on abdominal radiographs or of intracellular bacteria, extracellular bacteria, bile pigment, bowel contents, and cloudy appearance of fluid obtained by abdominocentesis or diagnostic peritoneal lavage fluid (see sections on abdominocentesis and diagnostic peritoneal lavage) warrant immediate surgical exploration. treatment largely consists of stabilizing the patient's cardiovascular and electrolyte status with intravenous fluids, administration of broad-spectrum antibiotics, and definitive surgical exploration and repair of injured structures. prolapse of the rectum is observed most frequently secondary to parasitism and gastrointestinal viral infections in young puppies and kittens with chronic diarrhea. older animals with rectal prolapse often have an underlying problem such as a tumor or mucosal lesion that causes straining and dyschezia. the diagnosis of a rectal prolapse is made based on physical examination findings. the diagnosis of rectal prolapse is sometimes difficult to distinguish from small intestinal intussusception. in rare cases, the intussusception can invaginate through the large bowel, rectum, and anus. the two entities are distinguished from one another by inserting a lubricated thermometer or blunt probe into the cul-de-sac formed by the junction of the prolapsed mucosa and mucocutaneous junction at the anal ring. inability to insert the probe or thermometer indicates that the rectal mucosa is prolapsed. passage of the probe signifies that the prolapsed segment is actually the intussusceptum. treatment can be performed easily if the prolapse is acute and the rectal mucosa is not too irritated or edematous. the presence of severely necrotic tissue warrants surgical intervention. to reduce an acute rectal prolapse, after placing the patient under general anesthesia, lubricate the prolapsed tissue and gently push it back into the rectum, using a lubricated syringe or syringe casing. apply a loose purse-string suture, leaving it in place for a minimum of hours. de-worm the patient and administer stool softeners. if a rectal prolapse cannot be reduced, or if the tissue is nonviable, surgical intervention is warranted. in patients in which viable tissue does not stay reduced with a purse-string suture, a colopexy can be performed during a laparotomy. first, place tension on the colon to reduce the prolapse, and then suture the colon to the peritoneum of the lateral abdominal wall with two to three rows of - or - monofilament suture material. if the prolapsed tissue is nonviable, it must be amputated. place four stay sutures at -degree intervals through the wall of the prolapse at the mucocutaneous junction. resect the prolapse distal to the stay sutures and then reestablish the rectal continuity by suturing the seromuscular layers together in one circumferential line and the mucosal layers together in the other. replace the suture incision into the anal canal. following surgery, de-worm the patient and administer a stool softener and analgesic drugs. avoid using thermometers or other probes in the immediate postoperative period because they may disrupt suture lines. acute gastritis may be associated with a variety of clinical conditions, including oral hemorrhage, ingestion of highly fermentable nondigestable foods or garbage, toxins, foreign bodies, renal or hepatic failure, inflammatory bowel disease, and bacterial and viral infections. diarrhea often accompanies or follows acute gastritis. hemorrhagic gastroenteritis often occurs as a shock-like syndrome with a rapidly rising hematocrit level. clinical signs of gastritis include depression, lethargy, anterior abdominal pain, excessive water consumption, vomiting, and dehydration. differential diagnosis of acute gastritis includes pancreatitis, hepatic or renal failure, gastrointestinal obstruction, and toxicities (box - ). the diagnosis is often a diagnosis of exclusion of other causes (see preceding text). a careful and thorough examination of the vomitus may be helpful in arriving at a diagnosis. a complete blood count, serum biochemistry profile including amylase and lipase, parvovirus test (in young puppies), fecal flotation and cytology, abdominal radiographs (plain and/or contrast studies), and abdominal ultrasound may be warranted to rule out other causes of acute vomiting. while diagnostic tests are being performed, treatment consists of withholding all food and water for a minimum of hours. after calculating the patient's degree of dehydration, administer a balanced crystalloid fluid to normalize acid-base and electrolyte status. control vomiting with antiemetics such as metoclopramide, prochlorperazine, chlorpromazine, dolasetron, and ondansetron (table - ). if vomiting is accompanied by diarrhea, administer broad-spectrum antibiotics (cefazolin, mg/kg iv q h, with metronidazole, mg/kg iv q h; or ampicillin, mg/kg iv q h, with enrofloxacin, mg/kg iv q h) to decrease the risk of bacterial translocation and bacteremia/septicemia. although antacids (famotidine, ranitidine, cimetidine) do not have a direct antiemetic effect, their use can decrease gastric acidity and esophageal irritation during vomiting. if gastritis is secondary to uremia or nonsteroidal antiinflammatory drug use, administer gastroprotectant and antiemetic drugs (ranitidine, mg/kg po q h; sucralfate, . - g/dog po q h; or omeprazole ( . - mg/kg po q h) to decrease acid secretion and coat areas of gastric ulceration (table - ) . once food and water can be tolerated, the patient can be placed on an oral diet and medications, and intravenous fluids can be discontinued. do not use until a gastrointestinal obstruction has been ruled out. hemorrhagic gastroenteritis (hge) is an acute onset of severe hemorrhagic vomiting and diarrhea most commonly observed in young small-breed dogs (e.g., poodles, miniature dachshunds, miniature schnauzers) to years of age. clinical signs develop rapidly and include vomiting and fetid diarrhea with hemorrhage, often strawberry jam-like in appearance. the hematocrit can rise from % to %. often, the animal is extremely hypovolemic but has no apparent signs of abdominal pain. there is no known cause of hge, although clostridium perfringens, escherichia coli, campylobacter, and viral infections have been suggested but not consistently confirmed. other differential diagnoses of of hematemesis and hemorrhagic diarrhea include coronavirus, parvovirus, vascular stasis, sepsis, hepatic cirrhosis with portal hypertension, and other causes of severe shock. immediate treatment consists of placement of a large-bore intravenous catheter and replenishment of intravascular fluid volume with crystalloid fluids (up to ml/kg/hour), while carefully monitoring the patient's hematocrit and total protein. administer broad-spectrum antibiotics (ampicillin, mg/kg iv q h, and enrofloxacin mg/kg iv q h) because of the high risk of bacterial translocation and sepsis. control vomiting with antiemetic drugs. monitor the patient's platelet count and coagulation tests for impending disseminated intravascular coagulation (dic), and administer fresh frozen plasma and heparin, as needed (see section on disseminated intravascular coagulation). when vomiting has ceased for hours, offer the animal small amounts of water, and then a bland diet (e.g., boiled chicken and rice or boiled ground beef and rice mixed with low-fat cottage cheese). pancreatitis occurs most frequently in dogs but can occur in cats as well. in dogs, the onset of pancreatitis is sometimes preceded by ingestion of a fatty meal or the administration of drugs (e.g., potassium bromide or glucocorticoids). glucocorticoids can increase the viscosity of pancreatic secretions and induce ductal proliferation, resulting in narrowing and obstruction of the lumen of the pancreatic duct. pancreatitis can also occur following blunt or penetrating abdominal trauma, high duodenal obstruction causing outflow obstruction of the pancreatic papilla, pancreatic ischemia, duodenal reflux, biliary disease, and hyperadrenocorticism. in cats, acute necrotizing pancreatitis is associated with anorexia, lethargy, hyperglycemia, icterus, and sometimes acute death. chronic pancreatitis is more common in cats and results in intermittent vomiting, anorexia, weight loss, and lethargy. predisposing causes of chronic pancreatitis in cats include pancreatic flukes, viral infection, hepatic lipidosis, drugs, organophosphate toxicity, and toxoplasmosis. clinical signs of acute pancreatitis include sudden severe vomiting, abdominal pain, and lethargy. depending on the severity of pancreatic inflammation, depression, hypotension, and systemic inflammatory response syndrome (sirs) may be present. subacute cases may have minimal clinical signs. severe pancreatic edema can result in vascular changes and ischemia that perpetuates severe inflammation. hypovolemic shock and dic can also decrease pancreatic perfusion. severe pancreatic edema, autolysis, and ischemia lead to pancreatic necrosis. duodenal irritation is manifested as both vomiting and diarrhea. pain may be localized to the right upper abdominal quadrant or may be generalized if peripancreatic saponification occurs. differential diagnosis of pancreatitis is the same as for any other cause of vomiting. complications that occur in patients with severe pancreatitis include dehydration, acidbase and electrolyte abnormalities, hyperlipemia, hypotension, and localized peritonitis. hepatic necrosis, lipidosis, congestion, and abnormal architecture can develop. inflammatory mediators (bradykinin, phospholipase a, elastase, myocardial depressant factor, and bacterial endotoxins) stimulate the inflammatory cascade and can lead to sirs, with severe hypotension, clotting system activation, and dic. electrolyte imbalances and hypovolemia secondary to vomiting all can lead to multiple organ dysfunction syndrome (mods), and ultimately, death. if a patient survives an episode of acute pancreatitis, long-term sequelae can include diabetes mellitus. monitor patients with recurrent pancreatitis for clinical signs of polyuria, polydipsia, polyphagia, hyperglycemia, and glucosuria. the diagnosis of pancreatitis is based on the presence of clinical signs (which may be absent in cats), laboratory findings, and ultrasonographic evidence of pancreatic edema and increased peripancreatic echogenicity. serum biochemistry analyses can sometimes support a diagnosis of pancreatitis; however, serum amylase and lipase are often unreliable indicators of pancreatitis, depending on the chronicity of the process in the individual patient. both serum amylase and lipase are excreted in the urine. impaired renal clearance/ function can cause artifactual elevations of serum amylase and lipase in the absence of pancreatic inflammation. furthermore, serum lipase levels can be elevated as a result of gastrointestinal obstruction (e.g., foreign body). early in the course of the disease, levels can be two to six times normal, but they may decrease to within normal ranges at the time of presentation to the veterinarian. the transient nature of amylase elevation makes this test difficult to interpret, and it is not highly sensitive if a normal value is found. lipase levels also increase later in the course of the disease. amylase and lipase should be tested concurrently with the rest of the biochemistry profile. other changes often observed are elevations in bun and creatinine levels secondary to dehydration and prerenal azotemia, hyperglycemia, and hyperlipemia. hypocalcemia can occur secondary to peripancreatic fat saponification, and its presence warrants a more negative prognosis. a more specific measure is pancreatic lipase immunoreactivity, which becomes elevated in dogs and cats with pancreatitis. this test, combined with ultrasonographic or computed tomography evidence of pancreatitis, is the most sensitive and specific test available for making an accurate diagnosis. however, because the results of this test take time to obtain, animals must be treated in the meantime. abdominal effusion or fluid from diagnostic peritoneal lavage can be compared with serum amylase and lipase activity. abdominal lipase and amylase concentrations in the fluid greater than that in the peripheral blood are characteristic of chemical peritonitis associated with pancreatitis. wbc counts greater than cells/mm , the presence of bacteria, toxic neutrophils, glucose levels less than mg/dl, or lactate levels greater than that of serum are characteristic of septic peritonitis, and immediate exploratory laparotomy is warranted. if a biopsy sample obtained during laparotomy does not demonstrate inflammation, but this does not rule out pancreatitis, because disease can be focal in nature and yet cause severe clinical signs. abdominal radiographs may sometimes reveal a loss of abdominal detail or a ground glass appearance in the right upper quadrant. pancreatic edema and duodenal irritation can displace the gastric axis toward the left, toward the left with dorsomedial displacement of the proximal duodenum (the so-called "backwards " or "shepherd's crook" sign). ultrasonography and ct are more sensitive in making a diagnosis of pancreatitis. treatment of pancreatitis is largely supportive in nature and is designed to correct hypovolemia and electrolyte imbalances, prevent or reverse shock, maintain vital organ perfusion, alleviate discomfort and pain, and prevent vomiting (see section on rule of twenty). when treating pancreatitis in dogs, all food and water should be restricted. however, food should not be withheld from cats with chronic pancreatitis. give fresh frozen plasma to replenish alpha- -macroglobulins. administer antiemetics such as chlorpromazine (use with caution in a hypovolemic or hypotensive patient), dolasetron, ondansetron, or metoclopramide to prevent or control vomiting. analgesic drugs can be provided in the form of constant rate infusion (fentanyl, - µ/kg/hour iv cri, and lidocaine, - µ/kg/minute iv cri), intrapleural injection (lidocaine, - mg/kg q h), or intermittent parenteral injections (morphine, . - mg/kg sq, im; hydromorphone, . mg/kg im or sq). because the pancreas must be rested, consider using parenteral nutrition. acute hepatic failure may be associated with toxins, adverse reaction to prescription medication, and bacterial or viral infections. the most frequent clinical signs observed in a patient with acute hepatic failure are anorexia, lethargy, vomiting, icterus, bleeding, and cns depression or seizures (associated with hepatic encephalopathy). differential diagnosis and causes of acute hepatic failure are listed in box - . diagnosis of acute hepatic failure is based on clinical signs and biochemical evidence of hepatocellular (ast, alt) and cholestatic (alk phos, t bili, ggt) enzyme elevations. ultrasonography may be helpful in distinguishing the architecture of the liver, but unless a mass or abscess is present, cannot provide a specific diagnosis of the cause of the hepatic damage. management of the patient with acute hepatic failure includes correction of dehydration and acid-base and electrolyte abnormalities, as shown in the following list: • hypoalbuminemia: plasma or concentrated albumin. plasma also is an excellent source of clotting factors that can become depleted. • clotting abnormalities: vitamin k ( . mg/kg sq or po q - h) to • severe anemia: fresh or stored blood • gastric hemorrhage: gastroprotectant drugs (omeprazole, ranitidine, famotidine, cimetidine, sucralfate) • hypoglycemia: dextrose supplementation ( . %- %) • hepatic failure, particularly when hypoglycemia is present: broad-spectrum antibiotics (ampicillin mg/kg iv q h; with enrofloxacin, mg/kg iv q h) • hepatic encephalopathy: lactulose or betadine enemas • cerebral edema: mannitol ( . - . g/kg iv over to minutes) followed by furosemide ( mg/kg iv minutes later). deterioration of clinical signs may signify the development of cerebral edema. applewhite aa, cornell kk, selcer ba: diagnosis and treatment of intussusception in dogs. comp cont educ pract vet ( ) often, systemic hypertension is diagnosed when the animal is seen by the veterinarian because of some other clinical sign, such as acute blindness, retinal detachment, hyphema, epistaxis, and cns signs following intracranial hemorrhage. diagnosis of systemic hypertension is often difficult in the absence of clinical signs and without performing invasive or noninvasive blood pressure monitoring. normal blood pressure (bp) measurements in dogs and cats are listed in table - . hypertension is defined as a consistent elevation in systolic bp > mm hg, consistent diastolic bp > mm hg, and consistent mean arterial blood pressure > mm hg. the effects of systemic hypertension include left ventricular hypertrophy, cerebrovascular accident, renal vascular injury, optic nerve edema, hyphema, retinal vascular tortuosity, retinal hemorrhage, retinal detachment, vomiting, neurologic defects, coma, and excessive bleeding from cut surfaces. emergency care dog - - - cat - - - patients with systemic hypertension should have a thorough diagnostic work-up to determine the underlying cause. although uncommon, hypertensive emergencies can occur with pheochromocytoma, acute renal failure, and acute glomerulonephritis. sodium nitroprusside ( - µ/kg/minute iv cri) or diltiazem ( . - . mg/kg iv given slowly over minutes, followed by µ/kg/minute) can be used to treat systemic hypertension. with the use of sodium nitroprusside or diltiazem, monitor carefully for hypotension. diagnosis is based on consistent elevations in systolic, diastolic, and/or mean arterial bp. because many of the clinical signs associated with systemic hypertension involve hemorrhage into some closed cavity, other causes of hemorrhage, such as vasculitis, thrombocytopenia, thrombocytopathia, and hepatic or renal failure, should be investigated (see section on coagulation disorders). diagnostic testing is based on clinical signs and index of suspicion for an underlying disease and may include a complete blood count; urinalysis; urine protein:creatinine ratio; acth stimulation test; thoracic and abdominal radiographs; thoracic and abdominal ultrasound; tick serology; brain ct or mri; and assays of serum electrolytes, aldosterone concentration, t , endogenous tsh, plasma catecholamine, and growth hormone. management of systemic hypertension involves treatment of the primary underlying disorder, whenever possible. long-term adjunctive management includes sodium restriction in the form of cooked or prescription diets to decrease fluid retention. obese animals should be placed on dietary restrictions and undergo a weight reduction program. thiazide and loop diuretics may be used to decrease sodium retention and circulating blood volume. alpha-and beta-adrenergic blockers may be used, but they are largely ineffective as monotherapeutic agents for treating hypertension. calcium channel blockers and angiotensin-converting enzyme (ace) inhibitors are the mainstay of therapy in the treatment of hypertension in dogs and cats ( diabetic ketoacidosis (dka) is a potentially fatal and terminal consequence of unregulated insulin deficiency and possible glucagon excess. in the absence of insulin, unregulated lipolysis results in the beta-hydroxylation of fatty acids by abnormal hepatic metabolism. as a result, ketoacids-namely, acetoacetic acid, beta-hydroxybutyric acid, and acetoneare produced. early in the course of the disease, patients exhibit clinical signs associated with diabetes mellitus: weight loss, polyuria, polyphagia, and polydipsia. later, as ketoacids stimulate the chemoreceptor trigger zone, vomiting and dehydration occur, with resulting hypovolemia, hypotension, severe depression, abdominal pain, oliguria, and coma. at the time of presentation, often a strong odor of ketones (acetone) is present on the patient's breath. physical examination often reveals dehydration, severe depression or coma, and hypovolemic shock. in extreme cases, the patient exhibits a slow, deep kussmaul respiratory pattern in an attempt to blow off excess co to compensate for the metabolic acidosis. a serum biochemistry profile and complete blood count often reveal prerenal azotemia, severe hyperglycemia (blood glucose > mg/dl), hyperosmolarity (> mosm/kg), lipemia, hypernatremia (sodium > meq/l), elevated hepatocellular and cholestatic enzyme activities, high anion gap, and metabolic acidosis. although a whole body potassium deficit is usually present, the serum potassium may appear artifactually elevated in response to metabolic acidosis. with severe metabolic acidosis, potassium moves extracellularly in exchange for a hydrogen ion. phosphorus too moves intracellularly in response to acidosis, and serum phosphorus is usually decreased. hypophosphatemia > mg/dl can result in intravascular hemolysis. urinalysis often reveals + glucosuria, ketonuria, and a specific gravity of . or greater. the urine of all diabetic animals should be cultured to rule out a urinary tract infection or pyelonephritis. treatment of a patient with dka presents a therapeutic challenge. treatment is aimed at providing adequate insulin to normalize cellular glucose metabolism, correcting acidbase and electrolyte imbalances, rehydration and restoration of perfusion, correcting acidosis, providing carbohydrate sources for utilization during insulin administration, and identifying any precipitating cause of the dka. obtain blood samples for a complete blood count, and serum biochemistry electrolyte profiles. whenever possible, insert a central venous catheter for fluid infusion and procurement of repeat blood samples. calculate the patient's dehydration deficit and maintenance fluid requirements and give appropriate fluid and electrolytes over a period of hours. it is advisable to rehydrate patients with severe hyperosmolarity for a minimum of hours before starting insulin administration. use a balanced electrolyte solution (e.g., plasmalyte-m, normosol-r, lactated ringer's solution) or . % saline solution for maintenance and rehydration. balanced electrolyte solutions contain small amounts of potassium and bicarbonate precursors that aid in the treatment of metabolic acidosis. treat animals with severe metabolic acidosis with an hco − > meq/l or a ph < . with supplemental bicarbonate ( . - . meq/kg). add supplemental dextrose to the patient's fluids as a carbohydrate source during insulin infusion. both insulin and carbohydrates are necessary for the proper metabolism of ketone bodies in patients with dka. the rate and type of fluid and amount of dextrose supplementation will change according to the patient's blood glucose concentration. serum potassium will drop rapidly as the metabolic acidosis is corrected with fluid and insulin administration. measure serum potassium every hours, if possible, and supplement accordingly (see section on fluid therapy for chart of potassium supplementation). if the patient's potassium requirement exceeds meq/l, or if the rate of potassium infusion approaches . meq/ kg/hour in the face of continued hypokalemia, magnesium should be supplemented. magnesium is required as a cofactor for many enzymatic processes and for normal function of the na,k-atpase pump. hypomagnesemia is a common electrolyte disturbance in many forms of critical illness. replenishing magnesium (mgcl , . meq/kg/day iv cri) often helps to correct the refractory hypokalemia observed in patients with dka. patients with hypophosphatemia that approaches . mmol/l should receive potassium phosphate ( . - . mmol/kg/hour iv cri). when providing potassium phosphate supplementation, be aware of the additional potassium added to the patient's fluids, so as to not exceed recommended rates of potassium infusion. to determine the amount of potassium chloride (kcl) to add along with potassium phosphate (kpo ), use the following formula: meq k + derived from kcl = total meq of k + to be administered over hours − meq in which k + is derived from kpo clinical signs of severe hypophosphatemia include muscle weakness, rhabdomyolysis, intravascular hemolysis, and decreased cerebral function that can lead to depression, stupor, seizures, or coma. regular insulin can be administered either im or as a constant rate infusion in the treatment of patients with dka. subcutaneous insulin should not be administered. because of the severe dehydration present in most patients with dka, subcutaneous insulin is poorly absorbed and is not effective until hydration has been restored. in the low-dose intravenous method, place regular insulin ( . units/kg for a cat, and . units/kg for a dog) in ml of . % saline solution. run ml of this mixture through the intravenous line to allow the insulin to adsorb to the plastic tubing. administer the patient's insulin fluid rate according to blood glucose levels ( table - ) . adjust the patient's total fluid volume according to changes in the insulin fluid rate as necessary. in many cases, multiple bags of fluids are necessary because they must be changed when fluctuations in blood glucose concentrations occur in response to therapy. infusion of the insulin mixture should be in a separate intravenous catheter. to replenish hydration, use a second intravenous line for the more rapid infusion of non-insulin-containing fluids. to administer the regular insulin im, first give . unit/kg im and then re-check the patient's blood glucose every hour. additional injections of regular insulin ( . unit/kg other fluid type (ml/hour) > . % nacl - . % nacl + . % dextrose - . % nacl + . % dextrose - . % nacl + . % dextrose < . % nacl + % dextrose im) should be administered based on the patient's response to subsequent injections. once the patient's blood glucose falls to to mg/dl, add . % to % dextrose to the fluids to maintain the blood glucose concentration at to mg/dl. continue intramuscular injection of regular insulin ( . - . unit/kg q - h) until the patient is rehydrated, no longer vomiting, and able to tolerate oral fluids and food without vomiting. even in patients with intramuscular regular insulin therapy, a central venous catheter should be placed for frequent blood sample collection. as the patient begins to respond to therapy, monitor electrolytes, glucose, and acid-base status carefully. hypokalemia, hypophosphatemia, and hypomagnesemia can occur. when the patient's hydration and acid-base status has normalized and the patient is able to tolerate oral food and water, a longer-acting insulin can be administered as for treatment of a patient with uncomplicated diabetes. extreme hyperosmolarity can result in a coma, if uncorrected. in patients with diabetes mellitus, hyperglycemia and hypernatremia secondary to osmotic diuresis and free water loss can lead to severe hyperosmolarity. in dogs, normal serum osmolality is < mosm/l of serum. hyperosmolarity is expected when serum osmolality is > mosm/l. if equipment for determining serum osmolarity is not available, osmolarity can be calculated by the following formula: osm/l = (na + k) + (glucose/ ) + (bun/ . ) patients with severe dehydration, hyperglycemia, hypernatremia, and azotemia may experience cerebral edema without ketonemia. treatment is directed solely at rehydrating the patient and slowly reducing blood glucose levels using a hypotonic solution such as . % nacl + . % dextrose or % dextrose in water (d w). after the initial rehydration period, administer potassium supplementation conservatively. red blood cells and the brain absolutely depend on the oxidation of glucose for energy. hypoglycemia can be caused by various systemic abnormalities that can be related to intestinal malabsorption of nutrients, impaired hepatic glycogenolysis or gluconeogenesis, and inadequate peripheral utilization of glucose. clinical signs of hypoglycemia are extremely variable and can include weakness, tremors, nervousness, polyphagia, ataxia, tachycardia, muscle twitching, incoordination, visual disturbances, and generalized seizures. clinical signs typically occur when serum glucose levels are < mg/dl. the combination of the clinical signs listed previously, documentation of low serum glucose, and alleviation of clinical signs upon glucose administration is known as whipple's triad. whenever a patient presents with hypoglycemia, consider the following important factors: the age of onset, the nature of the hypoglycemic episode (transient, persisent, or recurrent) , and the pattern based on the patient's history . treatment of hypoglycemia is directed at providing glucose supplementation and determining any underlying cause. administer supplemental dextrose ( %- % dextrose, - ml/kg iv; or % dextrose, ml/kg po) as quickly as possible. do not attempt oral glucose supplementation in any patient having a seizure or if the airway cannot be protected. administer intravenous fluids (e.g., normosol-r, lactated ringer's solution, . % saline solution) with . %- % supplemental dextrose until the patient is eating and able to maintain euglycemia without supplementation. in some cases (e.g., insulinoma), eating or administration of supplemental dextrose can promote insulin secretion and exacerbate clinical signs and hypoglycemia. in cases of refractory hypoglycemia secondary to iatrogenic insulin overdose, glucagon ( mg/kg iv bolus, then - ng/kg/minute iv cri) can also be administered along with supplemental dextrose. to make a glucagon infusion of ng/ml, reconstitute ml ( mg/ml) of glucagon according to the manufacturer's instructions and add this amount to ml of . % saline solution. emergency care the diagnosis of eclampsia (puerperal tetany) is often made on the basis of history and clinical signs. clinical signs can become evident when total calcium decreases to < . mg/dl in dogs and < . mg/dl in cats. the disease is often observed in small, excitable dogs, and stress may play a complicating role in the etiology. in most bitches, the disease manifests itself to weeks after parturition. in some cases, however, clinical signs can develop before parturition occurs. hypophosphatemia may accompany hypocalcemia. clinical signs of hypocalcemia include muscle tremors or fasciculations, panting, restlessness, aggression, hypersensitivity, disorientation, muscle cramping, hyperthermia, stiff gait, seizures, tachycardia, a prolonged qt interval on ecg, polydipsia, polyuria, and respiratory arrest. treatment of eclampsia consists of slow, cautious calcium supplementation ( % calcium gluconate, . mg/kg iv over minutes). severe refractory tetanus can be controlled with intravenous diazepam. supportive care includes intravenous fluid administration and cooling (see section on hyperthermia and heat-induced illness). instruct the owner to give the patient oral calcium supplements (e.g., to tablets of tums bid-tid) after discharge from the hospital. also instruct the owner about how to wean the puppies, allowing the bitch to dry up, in order to prevent recurrence. recurrence with subsequent pregnancies is common, particularly in patients that receive calcium supplementation during gestation (table - ) . hypercalcemia can occur from a variety of causes. the gosh darn it mnemonic can be used to remember the various causes of hypercalcemia in small animal patients (box - ) . the gastrointestinal, renal, and nervous systems are most commonly affected, particularly when serum total calcium rises above . mg/dl. clinical signs of severe hypercalcemia include muscle weakness, vomiting, seizures, and coma. ecg abnormalities include prolonged pr interval, rapid qt interval, and ventricular fibrillation. the most serious clinical signs are often seen when hypercalcemia is observed in combination with hyperphosphatemia or hypokalemia. pay special attention to the "calcium × phosphorus product." if this product exceeds , dystrophic calcification can occur, leading to renal failure. renal complications include polyuria, polydipsia, dehydration, and loss of renal tubular concentrating ability. renal blood flow and the glomerular filtration rate (gfr) are impaired when serum total calcium exceeds mg/dl. the extent, location, and number of renal tubular injuries are the main factors in determining whether renal damage secondary to hypercalcemia is reversible or irreversible. emergency therapy of hypercalcemia is warranted when severe renal compromise, cardiac dysfunction, or neurologic abnormalities are present, or if no clinical signs occur but the calcium × phosphorus product exceeds . the treatment of choice is correction of the underlying cause of hypercalcemia, whenever possible. in some cases, the results of diagnostic tests take time, and emergency therapy should be initiated immediately, before a definitive cause of the hypercalcemia is found. emergency management of hypercalcemia consists of reduction of serum calcium levels. administer intravenous fluids ( . % saline solution) to expand extracellular fluid volume and promote calciuresis. to promote diuresis, initial intravenous fluid rates should approach two to three times maintenance levels ( - ml/kg/day). potassium supplementation may be required to prevent iatrogenic hypokalemia. administration of a loop diuretic such as furosemide ( - mg/kg iv) will promote calcium excretion. calcitonin ( iu/kg im q h for cats and iu/kg im q h for dogs) can be administered to decrease serum calcium levels. in severe refractory hypercalcemia secondary to cholecalciferol toxicity, more aggressive calcitonin therapy ( - iu/kg sq q - h) can be attempted. side effects of calcitonin treatment include vomiting and diarrhea. alternatively, bisphosphonates (pamidronate, . - . mg/kg iv) are useful in rapidly reducing serum calcium concentrations. glucocorticosteroids reduce calcium release from the bone, decrease intestinal absorption of calcium, and promote renal calcium excretion. administer glucocorticosteroids only after the underlying cause of hypercalcemia has been determined and appropriate therapy started. because many forms of neoplasia can result in hypercalcemia as a paraneoplastic syndrome, empiric use of glucocorticosteroids can induce multiple drug resistance, making the tumor refractory to the effects of chemotherapeutic agents. hypoadrenocorticism is most commonly observed in young to middle-aged female dogs, but it can occur in animals of any age, gender, and breed. clinical signs, which are referable to deficiency in glucocorticoid (cortisol) and mineralocorticoid (aldosterone) hormones, may develop slowly over time, leading to a waxing and waning course; acute clinical signs occur when > % of the adrenal functional reserve has been destroyed. in such cases, complete adrenocortical collapse can result in an addisonian crisis. lack of aldosterone causes a lack of renal sodium and water retention, and impaired potassium excretion. the most significant clinical signs associated with hypoadrenocorticism are depression, lethargy, weakness, anorexia, shaking, shivering, vomiting, diarrhea, weight loss, abdominal pain, weakness, hypotension, dehydration, and inappropriate bradycardia (box - ) . the diagnosis of hypoadrenocorticism is made based on the patient's clinical signs in combination with electrolyte abnormalities that include hyperkalemia, hyponatremia, and hypochloremia. serum sodium concentration ( - meq/l) is often greatly reduced, and serum potassium is elevated (> . meq/l). a sodium:potassium ratio of < is characteristic of hypoadrenocorticism, although not exactly pathognomonic. electrocardiographic changes associated with hyperkalemia include inappropriate bradycardia, absence of p waves, elevated spiked t waves, and widened qrs complexes. other more variable bloodwork abnormalities include a lack of a stress leukogram, eosinophilia, hypoglycemia, hyperphosphatemia, hypercalcemia, azotemia, and hypocholesterolemia. a definitive diagnosis of hypoadrenocorticism is based on an adrenocorticotropic hormone (acth) stimulation test. in patients with hypoadrenocorticism, baseline cortisol levels are usually low, with a lack of appropriate cortisol release after administration of acth analogue. rarely, animals with "atypical" hypoadrenocorticism lose glucocorticoid secreting ability from the zona fasciculata, but retain mineralocorticoid secretory ability from the zona glomerulosa. atypical addisonian patients have normal serum electrolytes but still have clinical signs of vomiting, diarrhea, weakness, lethargy, inappetence, muscle wasting, and weight loss. the diagnosis is more difficult in such cases because of the presence of normal electrolytes. an acth stimulation test should be considered, particularly in predisposed breeds. treatment of hypoadrenocorticism includes placement of a large-bore intravenous catheter, infusion of intravenous crystalloid fluids ( . % saline solution), and replenishment of glucocorticoid and mineralocorticoid hormones. administer dexamethasone or dexamethasone-sodium phosphate ( . - . mg/kg iv). dexamethasone will not interfere with the acth stimulation test, unlike other longer-acting steroids (e.g., prednisolone, methylprednisolone sodium succinate, triamcinolone). depending on the severity of the patient's condition, consider monitoring using the rule of twenty. administer antiemetics and gastroprotectant drugs to treat nausea, vomiting, and hematemesis. give the patient broad-spectrum antibiotics (ampicillin, mg/kg iv q h) if hematochezia or hemorrhagic diarrhea is present. if severe gastrointestinal blood loss occurs, whole blood, packed red blood cells, or fresh frozen plasma may be required. control hypoglycemia with . %- . % dextrose. use sodium bicarbonate, regular insulin with dextrose, or calcium gluconate to correct severe hyperkalemia with atrial standstill (see section on atrial standstill). chronic therapy for hypoadrenocorticism consists of mineralocorticoid and glucocorticosteroids supplementation for the rest of the animal's life. mineralocorticoid supplementation can be in the form of desoxycorticosterone pivalate (docp) ( . mg/kg im) or fludrocortisone acetate ( . mg/ . - kg body weight daily). fludrocortisone acetate possesses both mineralocorticoid and glucocorticoid activities and can be used as the sole daily treatment of hypoadrenocorticism. (because fludrocortisone is poorly absorbed in some dogs, it may not completely normalize electrolyte abnormalities in these animals.) docp is primarily a mineralocorticoid. give supplemental glucocorticosteroids in the form of prednis(ol)one ( - . mg/kg/day). in dogs, iatrogenic hypoadrenocorticism can be caused by abrupt discontinuation of glucocorticosteroid treatment. long-term glucocorticosteroid supplementation can downregulate the pituitary gland's excretion of endogenous acth and the zona fasciculata's ability to excrete cortisol. however, the zona glomerulosa's ability to secrete aldosterone does not appear to be affected. clinical signs of iatrogenic hypoadrenocorticism include inability to compensate for stress, weakness, lethargy, vomiting, diarrhea, and collapse. treatment of iatrogenic hypoadrenocorticism is the same as for naturally occurring disease. following immediate emergency treatment, the patient should be weaned slowly from exogenous glucocorticosteroid supplementation. severe hyperthyroidism can manifest as a medical emergency as a result of hypermetabolism. clinical signs in affected cats with severe thyrotoxicosis include fever, severe tachycardia (heart rate > bpm), vomiting, hypertension, congestive heart failure with pulmonary edema, and fulminant collapse. clinical signs typically are manifested as an end-stage of chronic debilitation associated with hyperthyroidism and are often preceded by polyphagia, weight loss, cardiac murmur, polyuria/polydipsia (pu/pd), vomiting, and diarrhea. treatment of thyrotoxicosis includes antagonizing the adrenergic activity by administration of a beta-adrenergic blocker (esmolol, ( - µ/kg/minute, or propranolol, . mg/ kg/hour). administration of glucocorticosteroids (dexamethasone, mg/kg) may inhibit the conversion of thyroxine (t ) to the active form triiodothyronine (t ) and decrease peripheral tissue responsiveness to t , effectively blocking its effects. correct hypoglycemia with supplemental dextrose ( . %). use care to avoid overhydration in a patient with cardiac failure or insufficiency. start the patient on methimazole as quickly as possible and consider the use of radioactive iodine therapy. to maintain cerebral perfusion pressure, blood pressure must be normalized. if other concurrent injuries are suspected (e.g., pulmonary contusions), administer synthetic colloid fluids (dextran- , - ml/kg iv, or hetastarch, - ml/kg iv) to normalize blood pressure. although the use of colloids is controversial because of their potential to leak into the calvarium, the benefits of reestablishing cerebral perfusion far outweigh the risks of their use. hypertonic saline ( . % nacl, - ml/kg iv) can also be administered over to minutes to expand intravascular volume. maintain blood glucose within normal reference ranges whenever possible, because hyperglycemia is a negative prognostic indicator in cases of head trauma. if tremors or seizures cause hyperthermia or increased metabolism, active cooling of the patient is warranted (see sections on hyperthermia and heat-induced injury). all patients with head trauma should receive care and monitoring based on the rule of twenty (see section on rule of twenty). examine the patient's level of consciousness, response to various stimuli, pupil size and reactivity to light, physiologic nystagmus, and cranial nerve deficits. in dogs, damage to the midbrain often produces coma and decerebrate rigidity. initial consciousness followed by a unconsciousness or stupor usually involves an injury to the brainstem. brainstem lesions can be caused by compressive skull fractures, extradural or subdural hematomas, or herniation through the foramen magnum from cerebral edema (box - ) . the patient's pupil size and response to light can be used to localize a diagnosis and give a rough prognosis for severity of disease and possibility for return to function. pupils can be normal in size, mydriatic, or miotic. whenever a pupil appears miotic, direct ocular emergency care unconscious with no response to noxious stimuli injury with uveitis or secondary miosis due to brachial plexus injury should be ruled out. the eyes should always be examined to rule out ocular trauma. in a patient with head trauma, a change from dilated to constricted to normal pupil size is suggestive of improvement in clinical function. bilateral mydriatic pupils that are unresponsive to light in an unconscious animal are a grave prognostic sign and usually indicate an irreversible severe midbrain contusion. bilateral miotic pupils with normal nystagmus and ocular movements are associated with diffuse cerebral or diencephalic lesions. miotic pupils that become mydriatic indicate a progressive midbrain lesion with a poor prognosis. unilateral, slowly progressive pupillary abnormalities in the absence of direct ocular injury are characteristic of brainstem compression or herniation caused by progressive brain swelling. asymmetric pupils are seen in patients with rostral brainstem lesions and can change rapidly. unresponsive pupils that are seen in the midposition occur with brainstem lesions that extend into the medulla and are a grave sign. visual deficits are common with intracranial injury. lesions that are less severe and limited to the cerebrum produce contralateral menace deficits with normal pupillary light response. bilateral cerebral edema can cause blindness with a normal response to light if the midbrain is not disturbed. a patient that is severely depressed and recumbent may not respond to menacing gestures, even when visual pathways are intact. ocular, optic tract, optic nerve, or optic chiasm lesions can interfere with vision and the pupillary light response. brainstem contusion and cerebral edema may produce blindness and dilated unresponsive pupils due to disturbance of the oculomotor area. examine all cranial nerves carefully. cranial nerve abnormalities can indicate direct contusion or laceration of the neurons in the brainstem or where they exit the skull. cranial nerves that are initially normal then later lose function indicate a progressively expanding lesion. when specific cranial nerve deficits are present, the prognosis is considered guarded. clinical signs such as rolling to one side, torticollis, head tilt, and abnormal nystagmus are usually associated with petrosal bone or cerebellomedullary lesions that produce vestibular neuron dysfunction. fractures of the petrosal temporal bone often cause hemorrhage and cerebrospinal fluid (csf) leak from the external ear canal. if the lesion is limited to the membranous labyrinth, the loss of balance will be toward the injured side and the quick phase of the nystagmus will be toward the injured side. normal physiologic nystagmus requires that the pathway is between the peripheral vestibular neurons and the pontomedullary vestibular nuclei to the nuclei of the cranial nerves that innervate the extraocular muscles (iii, iv, vi). severe brainstem lesions disrupt this pathway. disruption of the pathway is manifested as an inability to produce normal physiologic nystagmus by moving the patient's head from side to side. in patients with severe central nervous system depression, this reflex may not be observed. next, assess postural changes and motor function abilities. a loss of the normal oculocephalic ("dolls-eye") reflex is an early sign of brainstem hemorrhage and a late sign of brainstem compression and herniation. any intracranial injury may be accompanied by a concurrent cervical spinal cord injury. handle animals with such injuries with extreme care to avoid causing further damage. whenever there is uncertainty whether a spinal cord lesion exists, strap the patient down to a flat surface and obtain radiographs of the spine. at least two orthogonal views may be required to see fractures; however, do not manipulate the patient until radiography has been completed. crosstable views, in which the bucky is turned perpendicular to the patient's spine, with a radiograph plate secured behind the patient, may be required to minimize patient motion. in patients with cerebral lesions, hemiparesis usually resolves within to days. evaluation of cranial nerve function at frequent intervals may reveal an initial injury or a progressively expanding lesion in the brain. signs of vestibular disorientation, marked head tilt, and abnormal nystagmus occur with contusions of the membranous labyrinth and fracture of the petrous temporal bone. hemorrhage and cerebrospinal fluid otorrhea may be visible from the external ear canal. rolling movements indicate an injury to the cerebellar-medullary vestibular system. respiratory dysfunction and abnormal respiratory patterns are sometimes observed with severe head injury. lesions of the diencephalon produce cheyne-stokes respirations, in which the patient takes progressively larger and larger breaths, pauses, then takes progressively smaller and smaller breaths. mesencephalic lesions cause hyperventilation and can result in respiratory alkalosis. medullary lesions result in a choppy, irregular respiratory pattern. clinical signs of respiratory dysfunction in the absence of primary respiratory damage indicate a guarded prognosis. after injury, seizures may be associated with intracranial hemorrhage, trauma, or an expanding intracranial mass lesion. immediately begin medical therapy to control the seizure. administer diazepam ( . mg/kg iv or . - . mg/kg/hour iv cri) to treat seizures. if diazepam is not effective in combination with other treatments to control intracranial edema, consider giving pentobarbital . loading doses of phenobarbital ( - mg/kg iv divided into or doses, given every to minutes) may be beneficial in preventing further seizures. severe refractory seizures or decreased mentation may be associated with cerebral edema and increased intracranial pressure. mannitol, an osmotic diuretic, is effective at reducing cerebral edema ( . - . g/kg iv over to minutes). mannitol also acts as a free radical scavenger that can inhibit the effects of cerebral ischemia-reperfusion injury. mannitol works synergistically with furosemide ( mg/kg iv given minutes after the mannitol infusion). corticosteroids have not been demonstrated to be beneficial in the treatment of head trauma and may induce hyperglycemia. hyperglycemia has been shown to be a negative prognostic indicator in cases of head trauma. also, glucocorticoids can suppress immune system function and impair wound healing. because of the known risks and lack of known benefits of glucocorticosteroids, their use in treatment of head trauma is contraindicated. the prognosis for any patient with severe head trauma is guarded. management of head trauma patients may include intense nursing care for a period of weeks to months, depending on the presence and extent of concurrent injuries. if progressive loss of consciousness occurs, surgery for decompression of compressive skull injuries should be considered. the most common injury associated with head trauma in small animals is a contusion with hemorrhage in the midbrain and pons. subdural or extradural hemorrhage with space-occupying blood clots is uncommon. diagnostic tests of head trauma may include skull radiographs, ct, and mri of the brain. special studies can help detect edema and hemorrhage in the brain and brainstem, and aid in making an accurate diagnosis and prognosis. a cerebrospinal fluid tap is contraindicated in patients with head trauma because of the risk of causing a rapid decrease in intracranial pressure and brainstem herniation. if a compressive skull fracture is present, the patient should be stabilized for surgery to remove the compression. surgery to alleviate increased intracranial pressure is rarely performed in veterinary medicine because of the poor prognosis and results. in some cases, when a lesion can be localized to one area, -to -cm burr holes can be placed through the skull over the affected area of the cerebrum, exposing the underlying brain tissue. blood clots can be removed through the holes. the bone flap may or may not be replaced, depending on the surgeon's preference and the degree of brain swelling. spinal cord injuries may be associated with trauma, disk rupture, fractures, and dislocation of the spinal column. proceed with caution when moving a patient with suspected spinal cord injury. avoid flexion, extension, and torsion of the vertebral column. all animals that are unconscious following a traumatic event should be considered to have cervical or thoracolumbar spinal injury until proved otherwise by radiography, ct, or mri. the animal should be moved onto a flat surface (e.g., board, door, window, picture frame) and taped down to prevent motion and further displacement of vertebrae. sedation with analgesics or tranquilizers may be necessary to keep the animal immobile and to minimize patient motion. whenever possible, avoid the use of narcotics in patients with head trauma because of the risk of increasing intracranial pressure. as in other emergencies, the abcs emergency care should be evaluated, and the patient treated for shock, hemorrhage, and respiratory compromise. once the cardiovascular and respiratory systems have been evaluated and stabilized, a more thorough neurologic examination can be performed. protrusion of an intervertebral disk indicates that the disk is bulging into the vertebral canal as a result of dorsal shifting of the nuclear pulposus disk material. disk extrusion refers to the rupture of the outer disk membrane and extrusion of the nuclear material into the vertebral column. in dogs and cats, there are intervertebral disks that potentially can cause a problem. chondrodystrophic breeds of dogs are predisposed to endochondral ossification and include the dachshund, shih tzu, french bulldog, bassett hound, welsh corgis, american spaniel, beagle, lhasa apso, and pekingese. initial examination of the patient with suspected intervertebral disk disease includes identifying the neuroanatomic location of the lesion based on clinical signs and neurologic deficits and then establishing a prognosis. the neurologic examination should be carried out without excessive manipulation of the animal. the presence of pain, edema, hemorrhage, or a visible deformity may localize an area of vertebral injury. once an area of suspected lesion is localized based on physical examination findings, take radiographs to establish a diagnosis and to institute therapy. in most cases, the animal must receive a short-acting anesthestic for proper radiographic technique and to prevent further injury. lateral and crosstable ventrodorsal (vd) or dorsoventral (dv) radiographs require less manipulation of the animal compared with traditional vd and dv projections. myelography is often required to delineate the location of the herniated disk material. prognosis in spinal cord injury depends on the extent of the injury and the reversibility of the damage. perception of noxious stimuli, or the presence of "deep pain," by the animal when the stimulus is applied caudal to the level of the lesion is a good sign. to apply a noxious stimulus, apply firm pressure to a toe on one of the rear limbs using a thick hemostat or a pair of pliers. flexion or withdrawl of the limb is simply a local spinal reflex, and should not be perceived as a positive response to or patient perception of the noxious stimulus. turning of the head, vocalization, dilation of the pupils, change in respiratory rate or character, or attempts to bite are behaviors that are more consistent with perception of the noxious stimulus. absence of perception of the noxious stimulus ("loss of deep pain") is a very poor prognosis for return to function. focal lesions are usually associated with vertebral fractures and displacement of the vertebral canal. focal lesions in one or more of the spinal cord segments from t to t can cause complete dysfunction of the injured tissue as a result of concussion, contusion, or laceration. the degree of structural damage cannot be determined from the neurologic signs alone. transverse focal lesions result in paraplegia, with intact pelvic limb spinal reflexes and analgesia of the limbs and body caudal to the lesion. clinical signs in patients with spinal injury are summarized in table - . carefully evaluate the cardiovascular and respiratory status of patients with spinal injuries. immediately address specific injuries such as pneumothorax, pulmonary contusions, hypovolemic shock, and open wounds. if there is palpable or radiographic evidence of a vertebral lesion causing compressive injury, surgery is the treatment of choice unless the displacement has compromised most or all of the vertebral canal. displacements through % to % of the vertebral canal are associated with a poor prognosis, particularly if deep pain is absent caudal to the lesion. in the absence of a radiographic lesion and in the presence of continued neurologic deficits, an mri or ct scan or myelography is warranted to localize a potentially correctable lesion. surgical exploration can be considered: with the objectives of providing spinal cord decompression by hemilaminectomy or laminectomy with removal of disk material or blood clots, realign and stabilize the vertebral column, and perform a meningotomy, if necessary. place the patient on a backboard or other rigid surface, taped down for transport and sedated, to be transported to a surgical specialist. the presence of worsening or ascending clinical signs may signify ascending-descending myelomalacia and is characteristic of a very poor prognosis.in acute spinal trauma, the use of glucocorticoids has been the mainstay of therapy; however, controversy exists about whether they actually offer any benefit. traditional glucocorticosteroid therapy is listed in box - . more recently, the use of propylene glycol has proved to be beneficial in the treatment of acute traumatic herniated disk. high-dose glucocorticoids should only be used for the first hours after initial injury. side effects of glucocorticosteroid therapy include gastric and intestinal ulceration. the prophylactic use of gastroprotectant drugs will not prevent gastrointestinal ulcer formation; however, if signs of gastrointestinal ulcer are present, institute gastroprotectant therapy. management of the patient with spinal cord injury includes aggressive nursing care and physical therapy. many patients with spinal cord injury have little to no control over bladder function, which results in chronic dribbling or retention of urine and overdistention of the urinary bladder with overflow incontinence. urinary bladder retention can lead to urinary tract infection, bladder atony, and overflow incontinence. manual expression of the bladder several times a day may be enough to keep the bladder empty. alternatively, place a urinary catheter to maintain patient cleanliness and to keep the bladder decompressed. (see section on urinary catheterization). paralytic ileus and fecal retention are frequent complications of spinal cord injury. to help prevent constipation, provide highly digestable foods and maintain the patient's hydration with oral and intravenous fluids. mild enemas or stool softeners can also be used to treat fecal retention. to prevent decubital ulcer formation, turn the patient every to hours, and use clean, dry, soft padded bedding. apply deep muscle massage and passive range of motion exercises to prevent disuse atrophy of the muscles and dependent edema. the radial nerve innervates the extensor muscles of the elbow, carpus, and digits. the radial nerve also supplies sensory innervation to the distal craniolateral surface of the forearm and the dorsal surface of the forepaw. injuries to the radial nerve at the level of the elbow emergency care cranial to c spastic tetraplegia or tetraparesis hyperreflexive all four limbs severe injury can result in death from respiratory failure. c -t tetraparesis or tetraplegia depressed thoracic limb spinal reflexes (lower motor neuron) hyperreflexive pelvic limbs (upper motor neuron) t -t horner' syndrome (prolapsed nictitans, enophthalmos, and miosis) t -l schiff-sherrington syndrome (extensor rigidity of thoracic limbs, flaccid paralysis with atonia, areflexia, and analgesia of pelvic limbs) result in an inability to extend the carpus and digits. as a result, the animal walks and bears weight on the dorsal surface of the paw. there is also loss of cutaneous sensation, which leads to paw injury. injuries to the radial nerve above the elbow (in the shoulder area) results in an inability to extend the elbow and bear weight on the affected limb. it can take weeks before the full extent of the injury and any return to function are manifested. the animal may need to be placed in a carpal flexion sling or have eventual amputation if distal limb injury or self-mutilation occurs. the sciatic nerve primarily innervates the caudal thigh muscles that flex the stifle and extend the hip. the tibial branch of the sciatic nerve innervates the caudal leg muscles that extend the tarsus and flex the digits. the tibial nerve provides the sole cutaneous sensory innervation to the plantar aspect of the paw and digits. the peroneal branch of the sciatic nerve provides the sole sensory cutaneous innervation to the dorsal surface of the paw ( table - ) . sciatic nerve injury may occur with pelvic fractures, particularly those that involve the body of the ileum at the greater ischiatic notch, or with sacroiliac luxations that contuse the l and l spinal nerves that pass ventral to the sacrum to contribute to the sciatic nerve. with sciatic nerve injury, there is decreased stifle flexion and overflexion of the hock (tibial nerve), and the animal walks on the dorsal surface of the paw (peroneal nerve). clinical signs of tibial or peroneal damage are seen with femur fractures or with inadvertent injection of drugs into the caudal thigh muscles. the femoral nerve innervates the extensor muscles of the stifle. the saphenous branch of the femoral nerve provides the sole cutaneous innervation to an area on the medial distal thigh, the leg, and the paw. the femoral nerve is protected by muscles and is rarely injured in pelvic fractures. clinical signs of femoral nerve injury are inability to support weight on the pelvic limb, absence of a patellar reflex, and analgesia in the area of cutaneous innervation. coma is complete loss of consciousness, with no response to noxious stimuli. in some animals that present in a coma or stuporous state, the immediate cause will be apparent. in other cases, however, a careful and thorough diagnostic work-up must be performed. a coma scale devised to assist in the clinical evaluation of the comatose patient is shown in table - . whenever an animal presents in a comatose state, immediately secure the emergency management of specific conditions c -t nerve roots radial nerve paralysis musculocutaneous nerve inability to flex the elbow axillary or thoracodorsal dropped elbow nerve median and ulnar nerves loss of cutaneous sensation on the caudal surface of the forearm and palmar and lateral surfaces of the paw; inability to flex the carpus and digits c -t nerve roots radial, median, or ulnar nerve injury c -c nerve roots musculocutaneous, suprascapular, and axillary injury c -t horner's syndrome (miosis, enophthalmos, and prolapsed nictitans) airway by placing an endotracheal tube (see section on endotracheal intubation). if necessary, provide respiratory assistance, or at a minimum, supplemental oxygen. control existing hemorrhage and treat shock, if present. take a careful and thorough history from the owner. make careful note of any seizure, trauma, or toxin exposure, and whether prior episodes of coma have ever occurred. perform a careful physical examination, taking note of the patient's temperature, pulse, and respiration. an elevated temperature may suggest the presence of systemic infection, such as pneumonia or hepatitis, or a brain lesion with loss of hypothalamic thermoregulatory control. very high temperatures associated with shock and coma are often observed in animals with heat stroke (see section on heat stroke and heat-induced illness). circulatory collapse or barbiturate overdose can produce coma and hypothermia. abnormal respiratory patterns also may be observed in a comatose patient. hypoventilation may occur with elevated intracranial pressure or barbiturate overdose. rapid respiratory rate may be associated with pneumonia, metabolic acidosis (dka, uremia), or brainstem injury. examine the skin for any bruises or external trauma. examine the mucous membranes and make note of color and capillary refill time. icterus with petechiae or ecchymotic hemorrhage in a comatose patient may be associated with end-stage hepatic failure and hepatic encephalopathy. smell the patient's breath for the odor of ketones that may signify dka or end-stage hepatic failure. motor activity normal gait, normal spinal reflexes hemiparesis, tetraparesis, or decerebrate activity recumbent, intermittent extensor rigidity recumbent, constant extensor rigidity recumbent, constant extensor rigidity with opisthotonus recumbent, hypotonia of muscles, depressed or absent spinal reflexes normal papillary reflexes and oculocephalic reflexes slow pupillary light reflexes and normal to reduced oculocephalic reflexes bilateral unresponsive miosis with normal to reduced oculocephalic reflexes pinpoint pupils with reduced to absent oculocephalic reflexes unilateral, unresponsive mydriasis with reduced to absent oculocephalic reflexes bilateral, unresponsive mydriasis with reduced to absent oculocephalic reflexes occasional periods of alertness and responsive to environment depression of delirium, capable of responding to environment but response may be inappropriate semicomatose, responsive to visual stimuli semicomatose, responsive to auditory stimuli semicomatose, responsive only to repeated noxious stimuli comatose, unresponsive to repeated noxious stimuli *neurologic function is assessed for each of the three categories and a grade of to is assigned according to the descriptions for each grade. the total score is the sum of the three category scores. this scale is designed to assist the clinician in evaluating the neurologic status of the craniocerebral trauma patient. as a guideline and according to clinical impressions, a consistent total score of to represents a grave prognosis, to a poor to guarded prognosis, and to a good prognosis. (modified from the glasgow coma scale used in humans.) from shores a: craniocerebral trauma. in kirk rw, ed: current veterinary therapy x. small animal practice. philadelphia, wb saunders, , p . finally, conduct a complete neurologic evaluation. the presence of asymmetric neurologic signs may suggest an intracranial mass lesion (e.g., hemorrhage, neoplasia, injury). usually, toxicities or metabolic disturbances (e.g., dka, hepatic encephalopathy) cause symmetric clinical signs of neurologic dysfunction, with cerebral signs predominating. in hepatic encephalopathy, pupils are usually normal in size and responsive to light. in toxicities, the pupils are abnormal in size and may be unresponsive to light. obtain a complete blood count, serum biochemistry profile, urinalysis, and specific tests for glucosuria and ketonuria. findings of a drastically elevated blood glucose with glucosuria, ketonuria, and high specific gravity are characteristic of dka. fever and uremic encephalopathy are characterized by severe azotemia with a low urine specific gravity. if barbiturate intoxication is suspected, save urine for later toxin analysis. evaluate urine sediment for calcium oxalate crystalluria that may indicate ethylene glycol toxicity. calculate plasma osmolality (see following section) to check for nonketotic hyperosmolar diabetes mellitus. elevated blood ammonia levels may be associated with hepatic encephalopathy. in uncontrolled diabetes mellitus, hyperosmolarity can result in clinical signs of disorientation, prostration, and coma. plasma osmolarity can be calculated from the formula: mosm/l = (na + k) + (glucose/ ) + (bun/ . ) clinical signs of hyperosmolarity can occur when the plasma osmolarity exceeds mosm/l. treatment of dka or nonketotic hyperosmolar syndrome is aimed at reducing ketoacid production, stimulating carbohydrate utilization, and impeding peripheral release of fatty acids. the treatment of choice is rehydration and provision of supplemental regular insulin and a carbohydrate source (see section on diabetic ketoacidosis). during ketosis, insulin resistance may be present. slow rehydration with . % saline solution or other balanced crystalloid fluids (e.g., normosol-r, plasmalyte-m, lactated ringer's solution), should occur, with the goal of rehydration over to hours. too rapid rehydration can result in cerebral edema and exacerbation of clinical signs. hepatic encephalopathy (he) is characterized by an abnormal mental state associated with severe hepatic insufficiency. the most common cause of he is congenital or acquired c o m a portosystemic shunts. acute hepatic destruction can also be caused by toxins, drugs, or infectious causes. the treatment of he is considered a medical emergency (table - ) . absorption of ammonia and other nitrogenous substances from the gastrointestinal tract is thought to be one of the complicating factors in he. prevent absorption of ammonia and other nitrogenous substances from the gastrointestinal tract by restricting dietary protein to % to % for dogs, and to % to % (on a dry matter basis) for cats. dietary protein should be from a nonanimal plant source (e.g., soybean) whenever possible. caloric requirements are met with lipids and carbohydrates. also prescribe cleansing enemas to rid the colon of residual material, and antibiotic therapy to reduce gastrointestinal tract bacteria. neomycin ( mg/kg q h) can be administered as a retention enema. metronidazole ( . mg/kg po, q - h) or amoxicillin-clavulanate ( . mg po q h) can also be administered. administer lactulose ( . - . ml q h for cats; . - ml q h for dogs) to trap ammonia in the colon to prevent absorption (table - ) . administer lactulose orally to an alert animal, or as a retention enema to a comatose animal. if lactulose is not available, betadine retention enemas will change colonic ph and prevent ammonia absorption. a side effect of lactulose administration (po) is soft to diarrheic stool. a seizure is a transient disturbance of brain function that is sudden in onset, ceases spontaneously, and has a tendency to recur, depending on the cause. most seizures are generalized and result in a loss of consciousness and severe involuntary contraction of the skeletal muscles, resulting in tonic-clonic limb activity and opisthotonus. mastication, salivation, urination, and defecation are common. partial (petit mal) seizures range from limited limb activity, facial muscle twitching, and episodic behavioral abnormalities to brief loss of consciousness. similar clinical signs also can occur with syncopal episodes. conduct a careful cardiac examination in any patient with a history of petit mal seizures. seizures of any form constitute a medical emergency, particularly when they occur in clusters, or as status epilepticus. most seizures are of short duration and may have subsided by the time the animal is presented for treatment. whenever a seizure occurs, however, it is important that the animal does not inadvertently injure itself or a bystander. it is important to evaluate whether the patient has a coexisting disease that can predispose it to seizures, such as hepatic failure, uremia, diabetes mellitus, hypoglycemia, toxin exposure, insulin-secreting tumors, and thiamine deficiency. many toxins are responsible for clinical signs of tremors or seizures (see section on poisons and toxins). treatment of a primary disease entity can help control seizures, in some cases, provided that the underlying cause is investigated and treated. status epilepticus, a state of continuous uncontrolled seizure activity, is a medical emergency. when an animal is in a state of status epilepticus, immediately place a lateral or medial saphenous intravenous catheter and administer diazepam ( . mg/kg iv) to help control the seizure. in most cases, the seizure must be controlled before a diagnostic workup is attempted. whenever possible, however, blood samples should be collected before administration of any anticonvulsant agent because of the risk of incorrect test results. for example, the propylene glycol carrier in diazepam can cause a false-positive ethylene glycol test using an in-house testing kit. whenever possible, check blood glucose levels, particularly in young puppies or kittens, to evaluate and treat hypoglycemia as a cause of seizures. if hypoglycemia exists, administer % dextrose ( g/kg iv). if diazepam partially controls the status epilepticus, administer a constant rate infusion ( . mg/kg/hour in % dextrose in water). diazepam is sensitive to light, and the bag and infusion line must be covered to prevent degradation of the drug. if diazepam fails to control status epilepticus, give pentobarbital ( - mg/kg iv to effect). the animal's airway should be intubated and protected while the patient is kept in the drug-induced coma. protracted cases of seizures may require mannitol and furosemide therapy to treat cerebral edema. administer intravenous fluids (balanced crystalloid at maintenance doses [see section on intravenous fluid therapy]). the patient should be turned every to hours to emergency care prevent atelectasis. insert a urinary catheter for cleanliness, and place the animal on soft dry padded bedding to prevent decubital ulcer formation. depending on the length of time that the patient is rendered unconscious, apply passive range of motion exercises and deep muscle massage to prevent disuse atrophy of the muscles and dependent or disuse edema. monitor the patient's oxygenation and ventilation status by arterial blood gas measurement or pulse oximetry and capnometry (see section on blood gas, pulse oximetry, and capnometry). administer supplemental oxygen to any patient that is hypoxemic secondary to hypoventilation or other causes. severe refractory seizures can result in the development of neurogenic pulmonary edema. lubricate the animal's eyes every hours to prevent drying out and corneal abrasions. depending on the cause of the seizure, administer phenobarbital at a loading dose of to mg/kg iv given in four to five injections, every to minutes; make sure that the patient is rousable in between injections). seizures in cats often are associated with structural brain disease. the occurrence of partial focal seizures is unequivocally associated with a focal cerebral lesion and acquired structural brain disease. an initial high frequency of seizures is also a strong indication that structural brain disease is present. seizure activity in cats may occur as mild generalized seizures or complex partial seizures and may be associated with systemic disorders such as feline infectious peritonitis virus, toxoplasmosis, cryptococcus infection, lymphosarcoma, meningiomas, ischemic encephalopathy, and thiamine deficiency. thiamine deficiency in the cat can be a medical emergency characterized by dilated pupils, ataxic gait, cerebellar tremor, abnormal oculocephalic reflex, and seizures. treatment consists of administration of thiamine ( mg/day) for three days. steffen f, grasmueck s: propofol for treatment of refractory seizures in dogs and a cat with intracranial disorders. j small anim pract ( ) ( ) ( ) ( ) . j am vet med assoc ( ): [ ] [ ] [ ] [ ] [ ] [ ] . an ocular emergency is any serious condition that causes or threatens to cause severe pain, deformity, or loss of vision. treat ocular emergencies immediately, within to several hours after the emergency, whenever possible (box - , - ). to assess the location and degree of ocular injury, perform a complete ocular examination. in some cases, short-acting sedation or general anesthesia in conjunction with topical local anesthetic may be necessary to perform the examination, because of patient discomfort and blepharospasm. the equipment listed in box - may be necessary and may be invaluable in making an accurate diagnosis. to perform a systematic and thorough ocular examination, first obtain a history from the owner. has there been any prior incident of ocular disease? is there any history of trauma or known chemical irritant or exposure? did the owner attempt any irrigation or medical techniques prior to presentation? when was the problem first noticed? has it changed at all since the owner noticed the problem? after a history has been obtained, examine the patient's eyes for discharge, blepharospasm, or photophobia. if any discharge is present, note its color and consistency. do not attempt to force the eyelids open if the patient is in extreme discomfort. administer a short-acting sedative and topical local anesthetic such as . % proparacaine. note the position of the globe within its orbit. if the eye is exophthalmic, strabismus and protrusion of the third eyelid are often visible. exposure keratitis may be present. in cases of retrobulbar or zygomatic salivary gland inflammation, the patient will resist opening the mouth and exhibit signs of discomfort or pain. note any swelling, contusions, abrasions, or lacerations of the eyelids. note whether the lids are able to close completely and cover the cornea. if a laceration of the lid is present, determine the depth of the laceration. palpate the orbit for fractures, swelling, pain, crepitus, and cellulitis. examine the cornea and sclera for penetrating injury or foreign material. the use of lid retractors or small forceps can be very helpful in these cases. if a wound appears to penetrate completely into the globe, look for loss of uveal tissue, lens, or vitreous. do not put any pressure on the globe, because intraocular herniation may result. examine the conjunctiva for hemorrhage, chemosis, lacerations, and foreign bodies. examine the superior and inferior conjunctival cul-de-sacs for foreign material. in such cases, placement of a topical anesthetic and use of a moistened cotton swab is invaluable to sweep the conjunctival fornix to pick up foreign bodies. use a small, fine-tipped forceps to retract the third eyelid away from the globe and examine behind the third eyelid for foreign bodies. next, examine the cornea for opacities, ulcers, foreign bodies, abrasions, or lacerations. place a small amount of fluroescein stain mixed with sterile water or saline on the dorsal sclera. close the eye to disperse the stain over the surface of the cornea, then flush gently with sterile saline irrigation. examine the cornea again for any defects. a linear defect perpendicular to the long axis of the eye should alert the clinician to investigate the conjunctiva for dystechia. record the pupil size, shape, and response to light (both direct and consensual). examine the anterior chamber and note its depth and whether hyphema or aqueous flare are present. is the lens clear and is it in the normal position? lens luxation can cause the lens tissue to touch the cornea and cause acute corneal edema. measure intraocular pressure with a schiotz tonometer or tonopen. finally, dilate the pupil and examine the posterior chamber using a direct or indirect ophthalmoscope to look for intraocular hemorrhage, retinal hemorrhage, retinal detachment, tortuous retinal vessels, optic neuritis, and inflammation. the basic surgical instruments listed in box - may be useful in the treatment of ocular lacerations and other ophthalmic injuries: bite wounds and automobile trauma commonly cause lacerations and abrasions of the lid margins. the lids can be considered to be two-layer structures, with the anterior composed of the skin and orbicularis muscle and the posterior layer composed of the tarsus and conjunctiva. the openings of the meibomian glands in the lid margin form the approximate line separating the lids into anterior and posterior segments. splitting the lid into these two segments facilitates the use of sliding skin flaps to close wound defects, if necessary. clean and thoroughly but gently irrigate the wound with sterile saline solution before attempting any lid laceration repair. use sterile saline solution to irrigate the wound and conjunctiva. a % povidone-iodine scrub can be used on the skin, taking care to avoid getting any scrub material in the soft tissues of the eye. drape the eye with an adhesive ocular drape, if possible, to prevent further wound contamination. trim the ragged wound edges, but be very conservative with tissue debridement. leave as much tissue as possible to insure proper wound contracture with minimal lid deformity. close a small lid wound with a figure-of-eight or two-layered simple interrupted suture of absorbable suture material or nylon in the skin. the lid margins must be absolutely apposed to prevent postoperative lid notching. direct blunt trauma to the eye can cause severe ecchymosis because of the excellent vascular supply of the eyelids. other associated ocular injuries such as orbital hemorrhage, proptosis, and corneal laceration may also occur. trauma, allergic reactions, inflammation of the sebaceous glands (hordeolum), thrombocytopenia, and vitamin k antagonist rodenticide intoxication can all cause ecchymoses of the lids. treat eyelid ecchymoses initially with cool compresses, followed by warm compresses. resorption of blood can occur from to days after the initial insult. ocular allergies respond well to topical application (dexamethasone ophthalmic ointment q - h) and systemic administration of glucocorticosteroids, along with cool compresses. in order to fully assess the conjunctiva for abnormalities, it may be necessary to carefully dissect it away from the underlying sclera. when performing this dissection, do not place undue pressure on the globe because of the risk of herniation of the intraocular contents through a scleral wound. repair large conjunctival lacerations with - absorbable sutures, using an interrupted or continuous pattern. carefully approximate the margins of the conjunctiva to prevent formation of inclusion cysts. when large areas of the conjunctiva have been damaged, advancement flaps may be required to close the defect. subconjunctival hemorrhage is a common sequela of head trauma, and it may also be observed in various coagulopathies. by itself, it is not a serious problem but may signify severe underlying intraocular damage. a complete ocular examination is indicated. other causes of subconjunctival hemorrhage include thrombocytopenia, autoimmune hemolytic anemia, hemophilia, leptospirosis, vitamin k antagonist rodenticide intoxication, severe systemic infection or inflammation, and prolonged labor (dystocia). uncomplicated subconjunctival hemorrhage usually clears on its own within days. if the conjunctiva is exposed because of swelling and hemorrhage, administer a topical protective triple antibiotic ophthalmic ointment every to hours until the conjunctival hemorrhage resolves. toxic, acid, and alkaline chemical injuries to the eye can sometimes occur. the severity of the injury caused by ocular burns depends on the concentration, type, and ph of the chemical and on the duration of exposure. weak acids do not penetrate biologic tissue very well. the hydrogen ion precipitates the protein upon contact and therefore provides some protection to the corneal stroma and intraocular contents. precipitation of corneal proteins produces a ground-glass appearance in the cornea. alkaline solutions and very strong acids penetrate tissues rapidly, causing saponification of the plasma membrane, denaturation of collagen, and vascular thrombosis within the conjunctiva, episclera, and anterior uvea. severe pain, blepharospasm, and photophobia are produced by exposure of free nerve endings in the corneal epithelium and conjunctiva. severe alkaline burns cause an increase in intraocular pressure. intraocular prostaglandins are released, and the intraocular aqueous ph increases, producing changes in the blood-aqueous barrier and secondary uveitis. uveitis with anterior synechia formation, eventual chronic glaucoma, phthisis, secondary cataract, and corneal perforation can occur. healing of the corneal epithelium is usually accomplished by neovascularization and sliding and increased mitosis of the corneal epithelium. severe stromal burns within the cornea heal by degradation and removal of necrotic debris, followed by replacement of the collagen matrix and corneal epithelial cells. the release of collagenase, endopeptidase, and cathepsins from polymorphonuclear cells serves to cause further corneal breakdown. in severe cases, only pmns may be present, and fibroblasts may never invade the corneal stroma. all chemical burns should be washed copiously with any clean aqueous solution available. if any sticky paste or powder is adherent to the conjunctival sac, remove it with moist cotton swabs and irrigation. begin mydriasis and cycloplegia by topical application of % atropine ophthalmic drops or ointment. start antibiotic therapy with triple antibiotic ophthalmic ointment or gentocin ointment every to hours. treat secondary glaucomas with topical carbonic anhydrase inhibitors. to avoid fibrinous adhesions and symblepharon formation, keep the conjunctival cul-de-sacs free of proteinaceous exudate that can form adhesions. analgesics are required for pain. oral nonsteroidal antiinflammatory agents such as carprofen, ketoprofen, meloxicam, or aspirin are recommended. persistent epithelial erosions may require a conjunctival flap left in place for to weeks or placement of a topical collagen shield (contact lens). topical antibiotics, mydriatics, and lubricants (lacrilube or puralube ointment) should also be used. strong acid or alkali burns can result in severe corneal stromal loss. in the past, topical n-acetylcysteine ( % mucomyst) has been recommended. this treatment is very painful. other treatments are also available, such as ethylenediaminetetraacetic acid (edta) ( . m solution) and patient serum to inhibit mammalian collagenase activity. to prepare patient serum, obtain to ml of whole blood from the patient. spin it down in a serum separator tube after a clot forms and then place the serum in a red-topped tube on the patient's cage. (the contents of the tube are viable for days without refrigeration.) apply the serum topically to the affected eye every to hours. avoid using topical steroids because they inhibit fibroblast formation and corneal healing. in severe cases, if conjunctival swelling and chemosis also are present, antiinflammatory doses of oral steroids can be administered short-term. oral steroids and nonsteroidal antiinflammatory drugs should never be administered to the patient concurrently, because of the risk of gastrointestinal ulcer and perforation. corneal abrasions are associated with severe pain, blepharospasm, lacrimation, and photophobia. animals with such intense pain are often difficult to examine until analgesia has been administered. topical use of proparacaine ( . % proparacaine hydrochloride) is usually sufficient to permit relaxation of the eyelids so that the eye can be examined. using a focal source of illumination and an eye loupe, examine the cornea, inferior and superior conjunctival fornixes, and medial aspect of the nictitans for foreign bodies. place a sterile drop of saline on a fluorescein-impregnated strip and touch the superior conjunctiva once to allow the stain to spread onto the surface of the eye. irrigate the eye to remove excess stain and then examine the corneal surface for any areas of stain uptake. if an area of the cornea persistently remains green, there is damage to the corneal epithelium in that area. initial treatment consists of application of a topical mydriatic ( drop of % atropine in affected eye q h) to prevent anterior synechiae and improve cycloplegia. triple antibiotic ointment is the treatment of choice (a / -inch strip in the affected eye q h) until the ulcer heals. in some cases, nonhealing ulcers (e.g., boxer ulcer, indolent ulcer) form in which the epithelial growth does not adhere to the underlying cornea. gently debride the loose edges of the ulcer/erosion with a cotton swab and topical anesthesia. more severe cases in which only minimal healing has occurred after days of treatment require grid keratectomy, in which a -gauge needle is used to gently scratch the surface of the abrasion or ulcer in the form of a grid to promote neovascularization. apply a topical anesthetic before performing the procedure. a collagen contact lens also may be required to promote wound healing. all corneal abrasions should be reevaluated in hours, and then every to days thereafter until they have healed. acute infectious keratitis secondary to bacterial infection is characterized by mucopurulent ocular discharge, rapidly progressing epithelial and corneal stromal loss, inflammatory cellular infiltrates into the corneal stroma, and secondary uveitis, often with hypopyon formation. confirmation of infectious keratitis is based on corneal scrapings and a positive gram stain. initial treatment for bacterial keratitis consists of systemic antibiotics and topical ciprofloxacin ( . % eyedrops or ointment). penetrating injuries through the cornea may result in prolapse of intraocular contents. frequently, pieces of uveal tissue or fibrin effectively but temporarily seal the defect and permit the anterior chamber to re-form. avoid manipulation of these wounds until the animal has been anesthetized, as struggling or excitement can promote loss or dislodgement of the temporary seal and cause the intraocular contents to be extruded. superficial corneal lacerations need not be sutured and can be treated the same as a superficial corneal ulcer or abrasion. if the laceration penetrates more than % the thickness of the cornea, or extends more than to mm, it should be sutured. when placing sutures in the cornea, it is helpful to use magnification. referral to a veterinary ophthalmologist is advised. if a veterinary ophthalmologist is not available, use - or - silk, collagen, or nylon sutures on a micropoint spatula-type needle. use a simple interrupted suture pattern and leave the sutures in place for a minimum of weeks. because many corneal lacerations are jagged and corneal edema forms, most of the wound edges cannot be tightly juxtaposed. in such cases, pull a conjunctival flap across the wound to prevent leakage of aqueous fluid. never suture through the full thickness of the cornea; rather, the suture should pass through the mid-third of the cornea. following closure of the corneal wound, the anterior chamber must be re-formed to prevent anterior synechia formation with secondary glaucoma. taking care to avoid iris injury, use a -or -gauge needle to insert sterile saline at the limbus. any defect in the suture line will be apparent because of leakage of the fluid from the site and should be repaired. incarceration of uveal tissue in corneal wounds is a difficult surgical problem. persistent incarceration of uveal tissue can result in development of a chronic wick in the cornea, a shallow anterior chamber, chronic irritation, edema, vascularization of the cornea, and intraocular infection that can lead to panophthalmitis. referral to a veterinary ophthalmologist is strongly recommended. the most common foreign bodies associated with ocular injuries in small animals are birdshot, bb pellets, and glass. the site of intraocular penetration of the foreign bodies may be obscured by the eyelids. a foreign body entering the eye may penetrate the cornea and fall into the anterior chamber or become lodged in the iris. foreign bodies may occasionally penetrate the lens capsule, producing cataracts. some metallic high-speed foreign bodies may penetrate the cornea, iris, and lens to lodge in the posterior wall of the eye or vitreous chamber. direct visualization of a foreign body is the best means of localization. examination of the eye with an indirect ophthalmoscope or biomicroscope (if available) is invaluable for locating foreign bodies. indirect visualization of the ocular foreign body can also be achieved through radiographic techniques. three separate views should be obtained to determine the plane of location of the foreign object. ct or mri may prove useful, although scatter from the foreign body may make it difficult to directly visualize with these techniques. ocular ultrasound is perhaps the most useful and refined radiographic technique for locating intraocular foreign bodies. before removing any foreign body from the eye, the risk and surgical danger of removing it must be weighed against the risks of leaving it in place. metallic foreign bodies in the anterior chamber are much easier to remove than nonmagnetic ones. attempted removal of foreign objects from the vitreous chamber of the eye has consistently produced poor results. for the best chance of recovery, ocular foreign bodies should be removed by a veterinary ophthalmologist whenever possible. blunt trauma to the globe can result in luxation or subluxation of the lens. the subluxated lens may move anteriorly and make the anterior chamber more shallow. trembling of the iris (iridodonesis) may be noticed when the lens is subluxated. in complete luxation, the lens may fall totally into the anterior chamber and obstruct aqueous outflow, causing secondary glaucoma. alternatively, the lens may be lost into the vitreous cavity. luxation of the lens is almost always associated with rupture of the hyaloid membrane and herniation of the vitreous through the pupillary space. emergency surgery for lens luxation is required if the lens is entirely within the anterior chamber or incarcerated within the pupil, causing a secondary pupillary block glaucoma. acute elevation in intraocular pressure can cause vision loss within hours; thus, lens removal should be accomplished as quickly as possible. referral to a veterinary ophthalmologist is recommended. severe trauma to the globe or a direct blow to the head can result in retinal or vitreous hemorrhage. there may be large areas of subretinal or intraretinal hemorrhage. subretinal hemorrhage assumes a discrete globular form, and the blood appears reddish-blue in color. the retina is detached at the site of hemorrhage. superficial retinal hemorrhage may assume a flame-shaped appearance, and preretinal or vitreous hemorrhage assumes a bright-red amorphous appearance, obliterating the underlying retinal architecture. retinal and vitreous hemorrhage secondary to trauma usually resorbs spontaneously over a -to -week period. unfortunately, vitreous hemorrhage, as it organizes, can produce vitreous traction bands that eventually produce retinal detachment. expulsive choroid hemorrhage can occur at the time of injury and usually leads to retinal detachment, severe visual impairment, and total loss of vision. treatment of vitreal and retinal hemorrhage includes rest and correction of factors that may predispose to intraocular hemorrhage. more complicated cases may require vitrectomy performed by a veterinary ophthalmologist. hyphema refers to blood in the anterior chamber of the eye. the most common traumatic cause of hyphema is an automobile accident. hyphema may also present because of penetrating ocular wounds and coagulopathies. blood within the eye may come from the anterior or posterior uveal tract. trauma to the eye may result in iridodialysis or a tearing of the iris at its root, permitting excessive bleeding from the iris and ciliary body. usually, simple hyphema resolves spontaneously in to days and does not cause vision loss. loss of vision following bleeding into the anterior chamber is associated with secondary ocular injuries such as glaucoma, traumatic iritis, cataract, retinal detachment, endophthalmitis, and corneal scarring. treatment of hyphema must be individualized, but there are severe general principles of treatment. first, stop ongoing hemorrhage and prevent further bleeding whenever possible. this may involve correction of the underlying cause, if a coagulopathy is present. next, aid in the elimination of blood from the anterior chamber, control secondary glaucoma, and treat associated injuries, including traumatic iritis. finally, detect and treat any late complications of glaucoma. in most cases of traumatic hyphema, little can be done to arrest or prevent ongoing hemorrhage. it is best to restrict the animal's activity and prohibit exertion. rebleeding can occur within days, and intraocular pressure must be monitored closely. after to days, the blood in the anterior chamber will change color from a bright red to bluish-black ("eight-ball hemorrhage"). if total hyphema persists and intraocular pressure rises despite therapy, surgical intervention by a veterinary ophthalmologist may be necessary. the primary route of escape of rbcs from the anterior chamber is via the anterior drainage angle. iris absorption and phagocytosis play a minor role in the removal of blood from the anterior chamber. because of the associated traumatic iritis in hyphema, topical administration of a glucocorticoid ( % dexamethasone drops or % prednisolone drops) is advised to control anterior chamber inflammation. a cycloplegic agent ( % atropine) should also be used. the formation of fibrin in the anterior chamber of the eye secondary to hemorrhage can produce adhesions of the iris and secondary glaucoma (see section on glaucoma secondary to hyphema) by blocking the trabecular network. hyphema secondary to retinal detachment (collie ectasia syndrome) and end-stage glaucoma are extremely difficult to treat medically and have a poor prognosis. proptosis of the globe is common secondary to trauma, particularly in brachycephalic breeds. proptosis of the globe in dolichocephalic breeds requires a greater degree of initiating contusion than the brachycephalic breeds because the orbits are so much deeper. therefore, secondary damage to the eye and cns associated with proptosis of the globe may be greater in the collie or greyhound than in the pug. when proptosis occurs, carefully evaluate the cardiovascular system for evidence of hypovolemic or hemorrhagic shock. examine the respiratory and neurologic systems. be sure to establish an airway and treat shock, if present. control hemorrhage and stabilize the cardiovascular system before attempting to replace the globe within its orbit or perform enucleation. during the initial management of the cardiovascular and respiratory systems, the eye should be covered with an ophthalmic grade ointment or sponges soaked in sterile saline to prevent the globe from drying out. proptosis of the globe can be associated with serious intraocular problems including iritis, chorioretinitis, retinal detachment, lens luxation, and avulsion of the optic nerve. stain the surface of the eye with fluorescein to look for topical abrasions or ulcers. carefully examine the sclera, cornea, and conjunctiva for penetrating injuries that may allow aqueous leakage. evaluate the size, location, and response to light of the pupil. a reactive pupil is better than a mydriatic fixed pupil. topical administration of a mydriatic (atropine %) to prevent persistent miosis and synechia formation is indicated, along with topical and oral antibiotics and oral analgesic therapy. reposition the proptosed globe with the patient under general anesthesia. make a lateral canthotomy incision to widen the palpebral fissure. lavage the globe with sterile saline irrigation to remove any external debris. place a copious amount of triple antibiotic ophthalmic ointment on the surface of the eye and then gently press the globe into the orbit using the flat side of a scalpel handle or a moistened sterile surgical sponge. do not probe the retro-orbital space with a needle or attempt to reduce intraocular pressure by paracentesis. when the globe is replaced in the orbit, close the lateral canthotomy incision with simple interrupted sutures. place three non-penetrating mattress sutures in the lid margins but do not draw them together. tighten the lid sutures through small pieces of a red rubber catheter or length of intravenous extension tubing to prevent the sutures from causing lid necrosis. leave the medial canthus of the eye open in order to allow topical treatment. postoperative treatment is directed at preventing further iritis and preventing infection. administer systemic broad-spectrum antibiotics (clavamox, . mg/kg po bid) and analgesic drugs. apply topical triple antibiotic ophthalmic ointment ( / inch in affected eye q - h) and atropine ( % in affected eye q h) to prevent infection, cycloplegia, and anterior synechiae. antiinflammatory doses of systemic steroids can also be added to the treatment if severe periorbital inflammation is present. systemic steroids should never be used in conjunction with nonsteroidal antiinflammatory drugs, because of the risk of gastrointestinal ulceration and perforation. the sutures should remain in place for a minimum of weeks. after this time, remove the sutures and inspect the globe. if proptosis recurs, repeat the treatment. following proptosis, strabismus is common secondary to periorbital muscle injury. even after extensive treatment, vision in the eye may still be lost. nonvisual eyes can remain in place, but phthisis may develop. carbonic anhydrase inhibitors such as acetazolamide and dichlorphenamide decrease aqueous secretion and may effectively reduce intraocular pressure if the trabecular outflow is still functioning at % of its capacity. an eye with a poorly functional trabecular outflow system will respond poorly to therapy with carbonic anhydrase inhibitors. osmotic agents such as mannitol or glycerol may be helpful in controlling glaucoma secondary to hyphema. reduction in vitreous chamber size can make the anterior chamber deeper and may allow increased aqueous outflow. evacuation of blood or blood clots from the anterior chamber is not advisable unless the glaucoma cannot be controlled medically or there is no indication after a prolonged period of time that blood is being resorbed. tissue plasminogen activator (t-pa) has proved to be useful in may be helpful in lysing blood clots and preventing excessive fibrin formation. the t-pa is reconstituted to make a solution of µ/ml, which is then frozen at − °c in . -ml aliquots. the thawed, warmed reconstituted t-pa is injected into the anterior chamber. blind probing of the anterior chamber of the eye and surgical intervention in an attempt to remove blood clots can cause serious complications such as rebleeding, lens luxation, iris damage, and damage to the corneal epithelium, and therefore is not advised. acute glaucoma is a rise in intraocular pressure that is not compatible with normal vision. glaucoma may present as early acute congestive or noncongestive glaucoma, or as end-stage disease. cardinal signs of glaucoma are a sudden onset of pain, photophobia, lacrimation, deep episcleral vascular engorgement, edematous insensitive cornea, shallow anterior chamber depth, dilated unresponsive pupil, loss of visual acuity, and buphthalmia. intraocular pressure usually exceeds mm hg but may be normal or only slightly increased if glaucoma is secondary to anterior uveitis. most forms of clinical glaucoma in dogs are secondary to some other intraocular problem. primary glaucoma is recognized in some breeds, including the bassett hound, cocker spaniel, samoyed, bouvier des flandres, and some terrier breeds either from goniodysgenesis or a predisposition to lens luxation. other common causes of acute glaucoma are anterior uveitis and intumescent lens secondary to rapid cataract development, particularly in dogs with diabetes mellitus. treatment involves investigation of the underlying cause of the sudden rise in intraocular pressure and rapid reduction in intraocular pressure. permanent visual impairment is often associated with chronically buphthalmic globes or the presence of rippling or striae formation on the cornea. referral to a veterinary ophthalmologist is recommended. if the eye is still visual and not buphthalmic, the prognosis is favorable, depending on the cause of the acute glaucoma. treatment to reduce intraocular pressure consists of improving aqueous outflow, reducing intraocular volume with osmotic agents, and reducing aqueous formation (table - ). the use of topical mydriatic agents in acute glaucoma is contraindicated because of the risk of making lens luxation or anterior uveitis worse. referral to a veterinary ophthalmologist for emergency surgery is indicated in cases of iris bombe, intumescent lens, or lens subluxation. administer osmotic agents to reduce the size of the vitreous body and the amount of aqueous. osmotic agents create an osmotic gradient between the intraocular fluids and the emergency management of specific conditions vascular bed, thus allowing osmotic removal of fluid independent of the aqueous inflow and outflow systems. if no other treatments are available, oral glycerol ( %, . ml/kg or . g/kg) can be used to effectively reduce intraocular pressure. an adverse side effect of oral glycerol treatment is protracted vomiting. do not use glycerol in a diabetic patient. mannitol ( - g/kg iv over hour) also effectively reduces intraocular pressure but does not cause vomiting. carbonic anhydrase inhibitors can be used to reduce intraocular volume by reducing aqueous production. oral administration of dichlorphenamide, methazolamide, and acetazolamide ( - mg/kg) is usually not very effective alone in reducing aqueous volume and intraocular pressure and also can cause metabolic acidosis. topical carbonic anhydrase inhibitors appear to be more effective (dorzolamide, trusopt) when used in conjunction with topical beta-blockers (timolol, . % or . % solution q h). the most effective treatment for acute pressure reduction is use of a topical prostaglandin inhibitor (latanaprost). usually just one or two drops effectively reduces intraocular pressure in the emergency stages, until the patient can be referred to a veterinary ophthalmologist the following day. many clinical conditions that are presented as emergencies may be due in part or wholly to the presence of a neoplasm. paraneoplastic signs are summarized in table - . prompt identification of the neoplasia combined with knowledge of treatment, expected response to therapy, and long-term prognosis can aid owners and practitioners in making appropriate treatment decisions. hemorrhage or effusion can occur in any body cavity as a result of the presence of benign or malignant tumors. tumors secrete anticoagulants to allow angiogenesis to grow unchecked. hemorrhage often occurs as a result of rupture of a neoplasm or invasion of a neoplasm into a major vascular structure. effusion may be the result of direct fluid production by the mass or may be due to obstruction of lymphatic or venous flow. hemorrhagic effusions in the abdominal cavity occur most commonly with neoplastic masses of the spleen or liver. the most common causes are hemangiosarcoma and hepatocellular carcinoma. clinical signs associated with acute abdominal hemorrhage, regardless of the cause, are related to hypovolemic shock and decreased perfusion and include pale mucous membranes, tachycardia, anemia, lethargy, and acute collapse. treatment for abdominal hemorrhage includes placement of a large-bore peripheral cephalic catheter and starting one fourth of a shock dose ( ml/kg/hour for dogs, and ml/kg/hour for cats) of intravenous crystalloid fluids, taking care to carefully monitor perfusion parameters of heart rate, capillary refill time, mucous membrane color, and blood pressure. administer intravenous colloids such as dextran- , hetastarch, and oxyglobin ( - ml/kg iv bolus) to restore intravascular volume and normotension. treat severe anemia with whole blood or packed rbcs to improve oxygen-carrying capacity and oxygen delivery (see sections on transfusion medicine and treatment of shock). confirm the presence of hemoabdomen abdominocentesis (see section on abdominocentesis). the presence of nonclotting hemorrhagic effusion is consistent with free blood. packed cell volume of the fluid is usually the same or higher than that of the peripheral blood. an abdominal compression bandage can be placed while further diagnostics are being performed. in cases of acute hemoabdomen, obtain right lateral, left lateral, and ventrodorsal or dorsoventral thoracic radiographs to help rule out obvious metastasis. monitor the patient's ecg and correct dysrhythmias as necessary (see section on cardiac dysrhythmias). surgery is indicated once the patient is stabilized. in some cases, hemorrhage is so severe that the patient should be taken immediately to surgery. when recommending surgery for a hemorrhaging intraabdominal mass, it is important to discuss likely diagnoses and long-term prognosis with the owner. hemangiosarcoma usually involves the spleen or liver or both. the presence of free abdominal hemorrhage is associated with a malignant tumor in % of cases. even when free abdominal hemorrhage is not present, the tumor is malignant in % of cases. approximately % (two thirds) of masses in the spleen are malignant (hemangiosarcoma, lymphoma, mast cell tumor, malignant fibrous histiocytoma, leiomyosarcoma, fibrosarcoma), and approximately one third are benign (hematoma, hemangioma). hepatocellular carcinoma usually affects one liver lobe (usually the left), and surgery is the treatment of choice. with complete surgical excision, median survival in dogs is longer than days. if diffuse disease is observed at the time of surgery, the prognosis is poor. nonhemorrhagic effusions are associated with mesothelioma, lymphoma, carcinomatosis, or any mass that causes vascular or lymphatic obstruction. clinical signs of respiratory distress and abdominal distention with nonhemorrhagic effusions are usually slowly progressive in onset and not as severe as those observed with hemorrhage. treatment is usually aimed at identification of the underlying cause. obtain a fluid sample via thoracocentesis or abdominocentesis. to obtain further cells for cytologic evaluation, aspirate fluid from the thoracic or abdominal mass with ultrasound guidance. cytologic evaluation of the fluid will often elucidate the causative tumor type. an abdominal ultrasound can determine the degree of metastasis. perform therapeutic abdominocentesis or thoracocentesis if the effusion is causing respiratory difficulty. rapid re-accumulation of the fluid potentially can cause hypoproteinemia and hypovolemic shock. mesothelioma is a rare tumor most commonly observed in urban environments. in humans, mesothelioma has been associated with exposure to asbestos. it is sometimes difficult to differentiate between reactive mesothelial cells and malignant mesothelial cells. treatment is aimed at controlling the neoplastic effusion. intracavitary cisplatin has been demonstrated to slow rates of fluid re-accumulation, but is largely a palliative therapy. lymphoma is another tumor type that can cause thoracic or abdominal effusion. cytologic evaluation of the fluid usually reveals abundant lymphoblasts. treatment with multiagent chemotherapy protocols, with or without adjunctive radiation therapy, can prevent tumor remission and stop fluid accumulation. carcinomatosis occurs as a result of diffuse seeding of the abdominal cavity with malignant carcinomas and has a poor prognosis. carcinomatosis may occur de novo or from metastasis of a primary tumor. treatment consists of fluid removal when respiratory difficulty occurs, with or without intracavitary cisplatin as a palliative measure. cisplatin should never be used in cats due to fatal acute pulmonary edema. clinical signs of hemorrhagic thoracic effusion include acute respiratory distress, anemia, hypovolemic or cardiogenic shock, and collapse. hemorrhagic thoracic effusions are rare in association with neoplastic effusions. a notable exception is intrathoracic hemorrhage in young dogs with osteosarcoma of the rib. hemorrhage can result when a primary lung tumor erodes through a vessel. hemangiosarcoma of the lungs or right auricular area can also result in hemorrhagic thoracic effusion. in many cases, hemorrhage may be confined to the pericardial sac with a right auricular mass, causing a globoid cardiac silhouette on thoracic radiographs. treatment consists of pericardiocentesis (see section on pericardial effusion and pericardiocentesis) and placement of a pericardial window, or the mass may be removed if it is in the right auricular appendage and resectable. although surgery can resolve clinical signs of right-sided heart failure, metastatic disease often develops soon afterward. nonhemorrhagic thoracic effusion is more common than hemorrhagic thoracic effusion, and is caused most commonly by mesothelioma, lymphoma, carcinomatosis, and thymoma. clinical signs develop gradually and include respiratory difficulty, cyanosis, and cough. supplemental oxygen should be administered. in many cases, thoracocentesis can be therapeutic and diagnostic. obtain thoracic radiographs both before and after thoracocentesis to determine whether a mass effect is present. following identification of a cause, definitive therapy can be instituted. mesotheliomas are rare and are associated with diffuse serosal disease. they are more common in dogs than in cats. effusions caused by mesotheliomas can affect the pleural or pericardial cavities. treatment is directed at removing effusion fluid and controlling reaccumulation with use of intracavitary platinum compounds, carboplatin, and cisplatin can be used in dogs. (cisplatin and carboplatin should never be used in cats.) chemical or physical pleurodesis may be helpful in controlling reaccumulation of fluid, but it is very painful in small animal patients. thoracic effusion secondary to lymphoma often is associated with an anterior mediastinal mass. t-cell lymphoma is the most common type of mediastinal mass observed in dogs. b-cell lymphoma is associated with a decreased response to chemotherapy and shorter survival times. treatment consists of combination chemotherapy with or without radiation therapy to decrease mass size. carcinomatosis is a diffuse disease of the pleural cavity that often is a result of metastasis from a primary pulmonary carcinoma or mammary adenocarcinoma. treatment is similar to that for mesothelioma and is aimed at controlling the effusion and delaying its recurrence. thymomas have been documented in both dogs and cats. dogs most commonly present with a cough, while cats present with clinical signs of respiratory distress and a restrictive respiratory pattern associated with the presence of pleural effusion. an anterior mediastinal mass is often observed on thoracic radiographs. in some cases, the pleural effusion must be drained via thoracocentesis before a mass is visible. ultrasound-guided aspiration and cytologic evaluation of the mass reveal a malignant epithelial tumor with small lymphocytes and mast cells. prognosis is good if the tumor can be completely excised. treatment consists of surgical removal with or without presurgical radiation therapy to shrink the mass. paraneoplastic syndromes of myasthenia gravis have been documented in dogs with thymomas. if megaesophagus or aspiration pneumonia is present, the prognosis is more guarded because of the high rate of complications. obstructive lesions affecting the urinary tract can be extramural (intra-abdominal, pelvic, or retroperitoneal) or intramural (urethral, bladder, or urethral wall) . transitional cell carcinoma is the most common type of bladder tumor observed in dogs. prostatic adenocarcinoma, or neoplasia of the sublumbar lymph nodes (lymphoma, adenocarcinoma from apocrine gland adenocarcinoma), also can cause urethral obstruction. treatment is aimed at relieving the obstruction and then attempting to identify the cause of the disease. to alleviate the obstruction, pass a urinary catheter whenever possible. perform cystocentesis only as a last resort because of the risk of seeding the peritoneal cavity with tumor cells if transitional cell carcinoma is the cause of the obstruction. institute supportive therapy including intravenous fluids and correction of electrolyte abnormalities. plain radiographs may reveal a mass lesion or may not be helpful without double contrast cystography. abdominal ultrasound is more sensitive in identifying a mass lesion in the urinary bladder. masses in the pelvic urethra are difficult to visualize with ultrasonography. double contrast cystourethrography is preferred. once the patient is stabilized, biopsy or surgery is indicated to identify the cause of the mass and attempt resection. urine tests for transitional cell carcinoma are available for identification of transitional cell carcinoma in the dog. complete surgical excision of transitional cell carcinoma or removal of benign tumors of the urinary bladder yields a favorable prognosis. poorer prognosis is seen with incomplete excision. many transitional cell carcinomas are located in the trigone region of the bladder and cannot be completely excised. the nonsteroidal antiinflammatory drug piroxicam is helpful in alleviating clinical signs for a reported -month median survival. in some dogs, cisplatin and carboplatin may delay recurrence of transitional cell carcinoma. tumors of the prostate gland are always malignant and occur with equal frequency in castrated and uncastrated male dogs. diagnosis of prostatic tumors is based on ultrasonographic evidence of a mass effect or prostatomegaly and on transrectal or transabdominal aspiration or biopsy. surgery, chemotherapy, and radiation therapy generally are unrewarding over the long term, although palliative radiation therapy may relieve clinical signs for to months. luminal tumors of the gastrointestinal tract typically cause obstruction, with slowly progressive clinical signs including vomiting, inappetence, and weight loss, or with acute severe protracted vomiting. extraluminal obstructive lesions usually arise from adhesions, or strangulation may occur, resulting in obstruction. perforation of the mass through the gastric or intestinal wall can cause peritonitis. treatment consists of initial stabilization and rehydration, evaluation for evidence of metastasis, and surgical resection of the affected area in cases of adenocarcinoma, leiomyoma, leiomyosarcoma, and obstructive or perforated lymphoma. gastric and intestinal adenocarcinoma are the most common gastrointestinal tumors observed in dogs. affected animals typically have a history of anorexia, weight loss, and vomiting. obtain an abdominal ultrasound before performing any surgery. fine needle aspirates of the mass and adjacent lymph nodes are usually diagnostic and can determine whether there is local metastasis. many tumors are not resectable, and metastasis occurs in approximately % of cases. dogs with smaller tumors that can be resected typically have longer survival times. leiomyosarcomas occur in the intestines of dogs, and carry a more favorable prognosis than adenocarcinoma if the mass can be completely resected. with complete resection, the average survival time is longer than year. the paraneoplastic syndrome of hypoglycemia has been observed with this tumor type. gastrointestinal lymphoma is the most common tumor of the gastrointestinal tract observed in cats. in comparison, it is relatively rare in dogs. unless there is complete obstruction or perforation of the gastrointestinal tract, surgical treatment for gastrointestinal lymphoma is not indicated. rather, multiple chemotherapy drugs are used in combination to achieve remission and resolution of the clinical signs of anorexia, weight loss, and vomiting. treatment responses unfortunately are poor. mast cell tumors of the gastrointestinal tract typically are manifested as gastrointestinal ulceration and hemorrhage in up to % of patients. the gastrointestinal hemorrhage that occurs with mast cell tumors results from increased acid secretion as a result of histamine receptor stimulation. treatment consists of histamine or proton pump inhibition (ranitidine, famotidine, cimetidine, or omeprazole). bowel perforation is a rare complication. many chemotherapy agents exert their effects on rapidly dividing normal and neoplastic cells. normal tissues that are commonly affected include the bone marrow, gastrointestinal tract, skin and hair follicles, and reproductive organs. some drugs have unique organspecific toxicities that must be monitored. knowledge and recognition of the expected type and onset of complications can alleviate their severity by rapid treatment, when complications occur (see table - ) . neutropenia is the most common bone marrow toxicity observed secondary to chemotherapy in small animal patients (table - ) . in most cases, the neutropenia is dose-dependent. the nadir, or lowest neutrophil count, is typically observed to days after chemotherapy treatment. once the nadir occurs, bone marrow recovery is observed, with an increase in circulating neutrophils within to hours (table - ) . treatment of myelosuppression is largely supportive to treat or prevent sepsis. prophylactic antibiotics are recommended in the afebrile patient with a neutrophil count < /µl. acceptable antibiotics include trimethoprim-sulfa and amoxicillin-clavulanate. granulocyte-colony stimulating factor (g-csf) (e.g., neupogen) is a recombinant human product that stimulates the release of neutrophils from the bone marrow, and its use shortens the recovery time following myelosuppressive drug therapy. disadvantages of g-csf include antibody production in response to the drug within weeks of use and its high cost. to prevent ongoing neutropenia, subsequent chemotherapy dosages should be decreased by %, and the interval in between treatments increased. whenever possible, overlap of myelosuppressive drugs should be avoided. acute gastrointestinal toxicity can occur within to hours after administration of cisplatin and actinomycin d. in many cases, pretreatment with the antiemetics metoclopramide, butorphanol, chlorpromazine, dolasetron or ondansetron can prevent chemotherapyinduced nausea and vomiting. vomiting can also occur as a delayed side effect to days after treatment with doxorubicin (adriamycin), actinomycin d, methotrexate, and cytoxan. in delayed reactions, vomiting and diarrhea are caused by damage to intestinal crypt cells. treatment consists of administration of antiemetics, intravenous fluids, and a bland highly digestible diet. doxorubicin also can cause hemorrhagic colitis within to days of administration. treatment includes a bland diet, metronidazole, and tylosin tartrate (tylan powder). emergency care mild to none not observed vincristine (low-dose), l-asparaginase, glucocorticosteroids moderate - days melphalan, cisplatin, mitoxantrone, actinomycin d severe - days doxorubicin, cyclophosphamide, vinblastine paralytic ileus can be observed to days after administration of vincristine. this side effect is more common in humans than animals and can be treated with metoclopramide once a gastrointestinal obstruction has been ruled out. cardiotoxicity doxorubicin (adriamycin) causes a dose-dependent dilative cardiomyopathy when the cumulative dose reaches to mg/m . in many cases, however, clinical signs do not occur until the cumulative dose is mg/m . the myocardial lesions are irreversible. treatment of cardiac dysrhythmias is dependent on the type of dysrhythmia (see section on treatment of dysrhythmias). discontinue doxorubicin and administer diuretics and positive inotropic therapy for dilative cardiomyopathy in order to delay the progression of congestive heart failure (see sections on treatment of congestive heart failure). if abnormalities are shown on electrocardiography performed before beginning therapy, substitute liposome-encapsulated doxorubicin or mitoxantrone substituted in the chemotherapy protocol. cardioprotectant drugs such as vitamin e, selenium, and n-acetyl cysteine have shown some promise in the prevention of doxorubicin-induced cardiotoxicity. cyclophosphamide can cause a sterile hemorrhagic cystitis. damage to the urinary bladder mucosa and vessels is caused by the toxic metabolite acrolein. clinical signs of sterile hemorrhagic cystitis include a history of cyclophosphamide administration, stranguria, hematuria, and pollakiuria. treatment for sterile hemorrhagic cystitis is discontinuation of the drug, treatment of any underlying urinary tract infection with antibiotic therapy based on susceptibility testing, and intravesicle drug administration. in extremely refractory cases, surgical debridement and cauterization of the bladder mucosa may be necessary. prevention of sterile hemorrhagic cystitis includes emptying the bladder frequently and administering the drug in the morning. concurrent administration of prednisone can induce polyuria and polydipsia. if sterile hemorrhagic cystitis occurs, chlorambucil can be substituted as a chemotherapeutic agent. anaphylactic reactions have been observed with the administration of l-asparaginase, adriamycin, etoposide, and paclitaxel. the risk of anaphylaxis increases with repeated administration, although in some animals anaphylaxis will occur on the first exposure to the drug. treatment consists of administration of epinephrine, diphenhydramine, famotidine, and glucocorticosteroids, as with any other life-threatening allergic reaction (see section on treatment of allergic reactions). to decrease the risk of an adverse reaction, give diphenhydramine ( . mg/kg im) to minutes before drug administration. slowing the rate of intravenous infusion also can decrease the chance of an anaphylactic reaction. cisplatin can cause a fatal irreversible pulmonary edema in cats, even at low dosages. -fluorouracil ( -fu) can cause a severe neurotoxicity in cats that results in ataxia and seizures. never use cisplatin or -fu in cats. poisoning cases benefit from a rapid, organized approach. key points in this approach are giving appropriate advice over the telephone, being able to access information sources, and providing appropriate treatment. there are only a few classes of poisons that account for the majority of toxicities reported in dogs and cats. every veterinarian should develop a familiarity with the clinical management of rodenticide and insecticide toxicity and be prepared with antidotes on hand. beyond the most common toxins, the spectrum of possibilities is endless, and the veterinarian must rely on appropriate information resources. it is important to have available a comprehensive source of pharmaceutical and plant identification resources. remarkably, considering the myriad of potentially toxic substances to which an animal can be exposed, relatively few specific antidotes are commonly used in veterinary medicine. because of the lack of specific antidotes, the veterinarian must treat each toxicity with general methods of poison management, applying basic critical care in the treatment of specific clinical signs associated with the poison exposure or toxicity. the adage "treat the patient, not the poison" often comes into play when the exact toxic substance is unknown, or has no specific antidote. before an animal arrives, the staff should be prepared to ask specific questions over the phone, and provide initial advice for clients, particularly if the animal lives some distance from the hospital (box - .) it is important to have access to a database of information on toxic substances. thousands of potentially toxic substances are available on the market today. the american society for the prevention of cruelty to animals (aspca) animal poison control center provides direct access to veterinary toxicologists hours a day, days a year. for additional information, call the nearest veterinary school or emergency center (box - ). also, see section for a table of emergency hotlines. check your local telephone book for a poison control center listing under emergency numbers, usually found on the front cover. although these numbers are for human poisonings, they have access to extensive poison and toxin databases and can potentially provide useful information for veterinarians, particularly regarding antidotal substances suitable for out of the ordinary toxins and human medications. information on the toxic ingredients in thousands of medications, insecticides, pesticides, and other registered commercial products has been confidentially placed by the government in these poison control centers. as new products are marketed, information regarding toxin ingredients is forwarded to the centers. various e-mail discussion lists can serve as an informative resource for practitioners, but access generally requires an initial subscription and may have the disadvantage of delayed *do not keep the client on the telephone for too long. lengthy histories can be performed once the animal is at your hospital and you have started to initiate treatment. † hair dressing products sometimes have hydrogen peroxide as a % w/v; this concentration is not suitable for induction of emesis. is your animal breathing or does it have respiratory difficulty? what is the color of the gums or tongue? is your animal able to walk? is there any vomiting, diarrhea, trembling, or seizures? does it appear lethargic or hyperactive? what is the substance that your animal ingested (was exposed to)? did you witness the ingestion or exposure? how much did the animal consume? how long ago was the exposure? was the substance swallowed, or is it on the animal's skin or eyes? how is the patient acting? how long has the animal been acting that way? or when was the last time you saw your animal act normally? . first aid instructions for the client: induce vomiting at home and save the vomitus. never induce vomiting if the patient is depressed, appears comatose, or is actively seizing. if the animal has ingested a caustic substance (strong alkali or acids) or a petroleum-based product (kerosene or turpentine), never recommend induction of emesis. hydrogen peroxide ( % w/v † ) ml = tsp/ lb of body weight can repeat once if no vomiting occurs after minutes . remind the owner to bring a sample of the toxin and the vomitus in with the patient. . advise the owner to transport the patient as rapidly as possible to the nearest veterinary hospital. response times. they are useful for ideas on standard and long-term therapy, but not emergency stabilization. an exception to this is the veterinary interactive network (vin), which posts message board communications. previous communications from veterinarians who treated a case with the same poison/toxin can be accessed with a subscription. many manufacturers operate an information service about their products. if the product label or name is available, check for a telephone number that may route you to a specialist. there are six essential steps in treating toxicities: . performing a physical examination . stabilizing the patient's vital signs . taking a thorough history . preventing continued absorption of the toxin . administering specific antidotes when available . facilitating clearance or metabolism of the absorbed toxin it is most important to provide symptomatic and supportive care both during and following emergency treatment. immediately on presentation, perform a brief but thorough physical examination. obtain a minimum database as well as serum, urine, or orogastric lavage samples for later toxicologic analyses. it is important at this time to systematically evaluate the patient's physical status, focusing particularly on the toxins most common to a particular geographic location and the organ systems most commonly affected by toxins in veterinary medicinenamely, the neurologic and gastrointestinal tracts. a checklist is useful when performing a complete physical examination (box - ). the minimium database includes a urine sample, packed cell volume, total protein, serum urea, and serum glucose. the information obtained from these simple cage-side tests is useful for determining dehydration, hemoconcentration, azotemia (renal or prerenal), and hypo-or hyperglycemia. when appropriate, obtain samples for serum biochemistry profiles, serum electrolytes, blood gases, serum osmolality, a complete hemogram, and coagulation profiles. samples of serum, urine, and any vomitus or orogastric lavage contents should be collected and saved for later toxicologic analyses as required later. stabilization of vital signs includes four major goals of treatment: maintain respiration, maintain cardiovascular function, control cns excitation, and control body temperature. in any patient with clinical signs of respiratory distress or respiratory dysfunction, supplemental oxygen should be administered via flow-by, oxygen hood, oxygen cage, nasal, nasopharyngeal, or transtracheal oxygen sources. ventilatory assistance may be necessary. irritant or corrosive substances can cause damage to the oropharyngeal mucosa to such an extent that airway obstruction occurs. when necessary, a temporary tracheostomy should be performed. arterial blood gases, pulse oximetry, and capnometry may be required to monitor oxygenation and ventilation. at the time of presentation, immediately place an intravenous catheter for administration of intravenous fluids, inotropes, antiarrhythmics, and antidotes, if necessary. the initial fluid of choice is a balanced crystalloid solution such as normosol-r, plasmalyte-m, or lactated ringer's solution. fluid therapy can later be changed based on the patient's acidbase and electrolyte status. some toxins can cause severe dysrhythmias and hyper-or hypotension. monitor blood pressure and perform ecg and correct any abnormalities according to standard therapy (see sections on hypotension and cardiac dysrhythmias). what is the pupil size? what is the pupil reactivity to light? is the ocular examination normal? what is the sensitivity to light or sound? nose: is it moist, dry, bubbling, or frothy, or caked with dirt? throat: are there any characteristic odors on the breath? are there any traces of foreign material on the tongue or in the crevices of the teeth or gums? are there petechiae or ecchymosis on the gums or bleeding from the gumline? what is the mucous membrane color? is it normal and pink, or dark red (injected), pale, or icteric? what is the capillary refill time? is it fast, normal, or slow? what is the patient's heart rate? are there any pulse deficits or dysrhythmias auscultated? what is the patient's blood pressure? what is the quality of the femoral pulse? is it synchronous with the heart rate, or are there dropped pulses? is the pulse bounding, normal, thready, or not palpable? what is the patient's electrocardiogram? what is the patient's respiratory rate? what is the patient's respiratory character? is it normal, fast, shallow, or labored? what do you hear on thoracic auscultation? do you hear harsh airway sounds or pulmonary crackles? what is the patient's rectal temperature? is there excessive salivation? is there evidence of vomiting or diarrhea? is abdominal palpation painful? do the intestinal loops feel normal, or are they fluid-filled or gas-filled? what is the color and consistency of the feces? is there a palpable urinary bladder? is there urine production? what is the color of the urine? peripheral lymph nodes should be normal in poisonings. some toxins cause hemolysis, methemoglobinemia, heinz body anemia, and coagulopathies. whole blood, fresh frozen plasma, packed rbcs, or hemoglobin-based oxygen carriers should be available and used if necessary. treat methemoglobinemia with a combination of ascorbic acid and n-acetylcysteine. many toxins affect the cns, producing clinical signs of excitation and/or seizures. diazepam is the drug of choice for most but not all seizures and tremors. if an animal has cns excitation secondary to the ingestion of selective norepinephrine reuptake inhibitors, avoid using diazepam, as it can potentially exacerbate clinical signs. muscle relaxants such as guaifenesin or methocarbamol may be required to control muscle spasm and tremors associated with some toxicities. consider animals that are in status epilepticus because of toxin exposure at high risk. such patients may not require the full dose of anesthetics or sedatives for seizure control. give phenobarbital ( - mg/kg iv) or pentobarbital ( - mg/kg iv to effect) for longer-term management of seizures. core body temperature can easily increase or decrease secondary to increased muscle activity or coma. animals may present as hypo-or hyperthermic, depending on the toxin ingested and the stage of toxicity. manage hypothermia with circulating hot water or hot air blankets, or place bubble wrap or saran wrap around the animal's peripheral extremities. manage hyperthermia by placing lukewarm wet towels on the patient until the rectal temperature has decreased to . °c ( °f). (see section on of hyperthermia and heat-induced illness). if sedatives or anesthetics have been used, initial hyperthermia may initially resolve due to hypothalamic loss of thermoregulatory control, cool water bathing should not be performed. when the patient is first presented to the veterinarian, have the owner complete a toxicologic history form (figure - ) while the animal is being initially assessed and vital signs are being stabilized. when initial stabilization of vital signs has been accomplished, the veterinarian can discuss the patient's history with the owner. in urgent situations, the veterinarian should obtain a brief history as an initial procedure (box - ). knowing when the animal was last seen as normal provides a time frame in which the toxic substance was most likely accessed, allowing differential diagnoses to be ranked in some order of probability by rate of onset. in eliciting a history from the owner about the animal's access to poisons, it is important not to take anything for granted. many owners do not realize how poisonous some substances can be, such as insecticide products, garbage, cleaning chemicals, and over-the-counter drugs commonly used by humans. many owners will deny that an animal could have ingested anything that might be toxic, not wanting to believe that the source of the toxin is within their household or property, particularly if recreational drug exposure is suspected. it is useful to phrase questions in a neutral fashion-for example, "is such-and-such present on the premises?" rather than "could the dog have eaten such-and-such?" if recreational drug exposure is suspected, another way to question the owners is to ask whether they have had any guests in their house recently that may have had such-and-such (e.g., marijuana, cocaine, methamphetamine). this approach serves to minimize the suggestion of any bias or preconceptions. when questioning an owner about recent events, it is useful to realize and acknowledge that disruption in the household routine is a distinct factor in the occurrence accidents, including poisonings. examples of such disruptive events include moving from the house, family member is ill or in the hospital, and renovations or recent construction. while these events are occurring, the safeguards followed by a normally careful owner may be disrupted. often, doors or gates may be left open, animals may be outside instead of inside (or vice versa), and inexperienced people may be pet-sitters. once owners are made aware of the importance of assessing such risks, they are often able to provide insight into otherwise baffling circumstances. various methods can be used to remove toxins from the gastrointestinal tract, including emesis, orogastric lavage, cathartics, and enemas. adsorbents, ion exchange resins, or precipitating or chelating agents may be used. removal of a toxic substance from the body surface may be necessary, depending on the toxin.the use of both emesis and orogastric lavage is less and less frequent in human medicine because of the risk of aspiration pneumonia and doubts about their efficacy. currently, management of poisonings in human medicine relies heavily on the use of activated charcoal combined with sorbitol as a cathartic, when appropriate, and supportive critical care. it should be emphasized, however, that the majority of poisonings in humans are due to drug overdoses (illicit or otherwise) (which have a relatively small volume and rapid absorption), for which this treatment is appropriate. furthermore, adoption of the approach rests on the availability of a hospital intensive care infrastructure, which is not always available in veterinary practice. induce emesis if the animal's physiology and neurologic status are stable (i.e., does not have respiratory depression or is not actively seizing, obtunded, unable to swallow or protect its airway). do not administer the same emetic more than twice. if the emetic doesn't work after two doses, give a different emetic or perform orogastric lavage under general anesthesia. emetics are strictly contraindicated for toxicity from petroleum-based products and corrosives because of the risk of aspiration pneumonia and further esophageal damage. emetics may also be of little value if poisons with antiemetic properties have been ingested, such as benzodiazepines, tricyclic antidepressants, and marijuana (table - ) . various emetics traditionally have been recommended for use in veterinary medicine. many have fallen out of favor because of the risk of causing adverse consequences and side effects. apomorphine ( . mg/kg iv or in the conjunctival sac) remains the standard but is less useful in certain situations in which the poison causes cns excitation or stimulation. it is ineffective in cats. other emetics include xylazine and hydrogen peroxide. do not use table salt because of the risk of severe oropharyngeal irritation and hypernatremia. do not use mustard powder or dishwashing liquid detergent because of the risk of severe oropharyngeal, esophageal, and gastric irritation. orogastric lavage is described in detail in the section on emergency procedures gastric lavage is contraindicated in treatment of toxicity from petroleum-based compounds and acid/alkali ingestion. the procedure can be messy but is very effective if performed within to hours of ingestion of the poison. to prevent aspiration, the patient should be placed under general anesthesia. keep the animal's head lowered during the procedure to prevent aspiration of stomach contents into the trachea. it is sometimes helpful to put the animal in both right and left lateral recumbency to allow complete emptying of gastric contents. repeat the procedure until the fluid runs clear from the stomach. in some cases in which solid material has been ingested, this process can take a long time, so be prepared with a large volume of warm water. following successful evacuation and lavage, administer a slurry of activated charcoal through the orogastric tube before removing it. keep the endotracheal tube cuffed and in place until the animal is semi-conscious, is starting the fight the tube, and is visibly able to swallow and protect its airway. • when was the animal last seen as normal? • what clinical signs developed? • how fast did the clinical signs develop? • when was the onset of clinical signs? • what is the animal's activity level? • does the animal have access to any poisonous substances? • this includes known toxins or chemicals, over-the-counter or prescription medications (including the owner's), and recreational drugs. enemas are useful to facilitate the action of cathartics and in cases in which the poison is a solid material (e.g., compost, snail bait, garbage) (box - ). it is best to use just lukewarm water. commercially available phosphate enema solutions can cause severe electrolyte disturbances (hyperphosphatemia, hyponatremia, hypocalcemia, and hypomagnesemia) and acid-base abnormalities (metabolic acidosis); therefore, they are absolutely contraindicated in small animal patients. use nonsterile nonspermicidal water-soluble lubricants (k-y jelly) old intravenous fluid bag enema bag -to -ml syringe fluid warm water, with or without hand or liquid dish soap the fluid volume required depends on the size of the animal and the state of its lower gastrointestinal tract. as with orogastric lavage, continue the procedure until the water runs clear. if difficulty is encountered emptying the lower gastrointestinal tract, repeat the enema in or hours, rather than be overzealous on the first attempt. cathartics are useful for hastening gastrointestinal elimination of toxins, and they are particularly useful for elimination of most solid toxicants (e.g., compost, garbage, snail baits). cathartics can be used in conjunction with activated charcoal. do not use magnesium-based cathartics in patients with cns depression, because hypermagnesemia can worsen this disorder and also cause cardiac rhythm disturbances (table - ) . activated charcoal ( - ml/kg) is the safest and to date the most effective adsorbent for the treatment of ingested toxins. activated charcoal can be administered after emesis or orogastric lavage or can be administered as the sole treatment. various preparations are available on the market, including dry powder, compressed tablets, granules, liquid suspensions, and concentrated paste preparations. commercially available products are relatively inexpensive and should be used whenever possible for ease of administration. vegetableorigin activated charcoal is the most efficient adsorbent and binds compounds with weak, nonionic bonds. some preparations are combined with sorbitol to provide simultaneous administration of an adsorbent and a cathartic; this combination has been shown to be most efficacious. repeated administration of activated charcoal every to hours has been shown to be beneficial in the management of a toxin that undergoes enterohepatic recirculation. administration of an oily cathartic or mixing the activated charcoal with food only serves to reduce the absorptive surface of the activated charcoal and therefore is not recommended. in general, substances that are very soluble and are rapidly absorbed are not well adsorbed by activated charcoal, including alkalis, nitrates, mineral acids, ethanol, methanol, ferrous sulfate, ammonia, and cyanide. kaolin and bentonite are clays that have been used as adsorbents. both are usually less effective than activated charcoal. however, they are reported to be better adsorbents than activated charcoal for the herbicide paraquat. ion exchange resins can ionically bind certain drugs or toxins. cholestyramine is one such resin, commonly used in human medicine to bind intestinal bile acids and thereby decrease cholesterol absorption. its application in toxicology extends to the absorption of fat-soluble toxins such as organochlorine and certain acidic compounds such as digitalis. ion exchange resins also have been used to delay or reduce the absorption of phenylbutazone, warfarin, chlorothiazide, tetracycline, phenobarbital, and thyroid preparations. precipitating, chelating, and diluting agents precipitating, chelating, and diluting agents are used primarily in the management of heavy metal intoxications, such as alkaloids or oxalates. they work by binding preferentially to the metal ion and creating a more soluble complex that is amenable to renal excretion. those chelating agents in common usage are calcium edta, deferoxamine, and d-penicillamine. calcium edta and deferoxamine should both be on hand in the veterinary hospital because they are necessary to treat zinc and iron toxicity, respectively, both of which have a short window of opportunity for therapeutic intervention. d-penicillamine has a wide application for a number of metal toxicities but tends to be used for long-term chronic therapy because it can be administered orally. various agents used for nonspecific dilution of toxins, including milk of magnesia and egg whites, although old-fashioned, still have wide application in many cases in which low-grade irritants have been ingested. bathing the animal is an important aspect of treatment for topical exposures to toxins such as insecticidal products, petroleum-based products, and aromatic oils. bathing an animal is not an innocuous procedure. to avoid hypothermia and shock, use warm water at all times. actively dry the animal to further minimize the risk of hypothermia. when bathing the animal, use rubber gloves and a plastic apron to avoid exposure to noxious agents. in most cases, a mild dishwashing soap is appropriate. medicated or antibacterial shampoos are less appropriate in this situation. for petroleum-based products in particular, dawn dishwashing liquid that "cuts the grease" works well to remove the oils. if dawn is not available, mechanics' hand cleaners or coconut oil-based soaps can be used instead. as a general principle, best results are obtained by barely wetting the patient's fur until the detergent is worked well into the fur, keeping the amount of water to a minimum until ready for the rinse. oil-based paint is best removed by clipping rather than by attempting removal with solvents, because solvents are also toxic. to remove powder products, brush and vacuum the animal before bathing it to eliminate further toxic exposure. with caustic alkaline or acidic products, the primary treatment is to dilute and flush the skin with warm water; do not attempt neutralization. neutralization can cause an exothermic reaction that causes further damage to the underlying tissues. eliminating poison from the eyes for ocular exposures, irrigate the eyes for a minimum of to minutes with warm (body temperature) tap water or warmed . % sterile saline solution. the use of neutralizing substances is not recommended because of the risk of causing further ocular damage. following adequate irrigation, treat chemical burns of the eyes with lubricating ointments and possibly a temporary tarsorrhaphy. atropine may be indicated as a cycloplegic agent. systemic nonsteroidal antiinflammatory drugs can be used to control patient discomfort. daily follow-up examinations are required because epithelial damage may be delayed, especially with alkali burns, and it is difficult to predict the final extent of ocular damage. topical glucocorticosteroids are contraindicated if the corneal epithelium is not intact. if severe conjunctival swelling is present with a corneal ulcer, parenteral glucocorticosteroids can be administered to help alleviate inflammation, but nonsteroidal antiinflammatory drugs should not be used simultaneously due to the risk of gastrointestinal ulceration or perforation. whenever possible, administer specific antidotes to negate the effects of the toxin and prevent conversion of the substance to the toxic metabolite. three categories of agents are used in the management of poisonings. the first category is specific antidotes. unfortunately, few specific antidotes are available for use in veterinary medicine. some "classic" toxins and antidotes are now considered to be rare, such as curare and physostigmine, thallium and prussian blue, and fluoride and calcium borogluconate. these and a few others have been omitted from the table. the second, broader category of antidotes includes those drugs used in the symptomatic management of clinical signs, which are part of our routine veterinary stock. drugs such as atropine, sedatives, steroids, antiarrhythmics, and beta-blockers fall into this category. the third category comprises nonspecific decontaminants such as activated charcoal, cathartics, and emetics. these were discussed previously. many patients benefit from efforts to enhance clearance or metabolism of the absorbed toxins. some specific therapies have been developed for this purpose, including -methylpyrazole for ethylene glycol toxicity and specific antibodies such as digibind (digoxin immune fab [ovine]) for digitalis toxicity. other strategies are aimed at promoting renal excretion. renal excretion strategies include diuresis, ion trapping, and peritoneal dialysis or hemodialysis (see section on peritoneal dialysis). diuresis and ion trapping are applicable to a large number of toxins and are discussed here in more detail. other toxins respond to urine acidification and urine alkalinization. enhancing renal excretion of substances is most useful for those organic substances that are present in significant concentrations in the plasma. substances that are non-ionic and lipid-soluble, such as certain herbicides, are likely to be less affected by attempts to promote rapid renal elimination. before starting diuresis or ion trapping, intravenous fluid therapy should be adequate as determined by normal central venous pressure, urine output, and mean arterial blood pressure. if any of these values are less than normal, use other measures to ensure adequate renal perfusion, including but not limited to a constant rate infusion of dopamine. simple fluid diuresis can influence the excretion of certain substances. the use of mannitol as an osmotic diuretic may reduce the passive reabsorption of some toxic substances in the proximal renal convoluted tubule by reducing water reabsorption. dextrose ( %) can be used as an osmotic diuretic. furosemide can be used to promote diuresis, but again, there is no substitute for intravenous fluid therapy. the use of mannitol, dextrose, and furosemide is contraindicated in hypotensive or hypovolemic patients. take care to avoid causing dehydration with any diuretic; central venous pressure monitoring is strongly recommended. ion trapping is based on the principle that ionized substances do not cross renal tubular membranes easily, and are not well reabsorbed. if the urinary ph can be changed so that the toxin's chemical equilibrium shifts to its ionized form, then that toxin can be "trapped" in the urine and excreted. alkaline urine favors the ionization of acidic compounds, and acidic urine favors the ionization of alkaline compounds. those toxins that are amenable to ion trapping are mostly weak acids and weak bases. ammonium chloride can be used to promote urinary acidification. contraindications to the use of ammonium chloride include a preexisting metabolic acidosis, hepatic or renal insufficiency, and hemolysis or rhabdomyolysis leading to hemoglobinuria or myoglobinuria. signs of ammonia intoxication include cns depression and coma. when performing urine acidification, frequently check the serum potassium concentration and urine ph. urine alkalinization can be performed with use of sodium bicarbonate. contraindications to the use of sodium bicarbonate include metabolic alkalosis (particularly with concurrent use of furosemide), hypocalcemia, and hypokalemia. as with urine acidification, monitor the serum potassium concentration and urine ph frequently. the major steps in management of poisonings discussed here must be accompanied by application of the fundamentals of critical care. respiratory and cardiovascular support have been discussed previously. renal and gastrointestinal function and analgesia are particularly important in the management of the poisoning patient. maintenance of renal perfusion is a priority in the poisoning patient. fluid, electrolyte, and acid-base balance must be controlled and be accurate. poisoning patients are at particularly high risk for renal damage and acute renal failure, whether by primary toxic insult to the renal parenchyma or by acute or prolonged renal hypoperfusion. for this reason, a protocol that aims at preventing oliguria and ensuing renal failure is one of the therapeutic strategies that should be routinely employed. this protocol is described in box - . gastrointestinal protectant drugs may be indicated for the management of those poisons that are gastrointestinal irritants or ulcerogenic. commonly used gastroprotectant drugs include cimetidine, ranitidine, famotidine, omeprazole, sucralfate, and misoprostol. antiemetics may be used to suppress intractable vomiting. metoclopramide is commonly used, and it is the drug of choice for centrally mediated nausea. antiemetics that work by different mechanisms can be used in combination as necessary. examples are dopamine -receptor antagonists such as prochlorperazine, -hydroxytryptamine antagonists such as ondansetron and dolasetron, and h- receptor antagonists such as diphenhydramine and meclizine. analgesics are more appropriate to treat poisonings than once thought. common effects of poisons including severe gastroenteritis and topical burns or ulcerations may warrant the use of analgesics. longer-acting analgesics such as morphine, hydromorphone, and buprenorphine are particularly useful. nutritional support may be necessary in the form of enteral or parenteral feeding in patients that have esophageal or gastric damage or that need to be sedated for long periods of time. endoscopy may be useful in assessing the degree of esophageal and gastric damage, particularly after ingestion of caustic substances. introduction: acetaminophen (paracetamol) is the active ingredient in tylenol and many over-thecounter cold products. acetaminophen is converted to n-acetyl-p-benzoquinonimine in the liver, a toxic substance that can cause oxidative injury of red blood cells and hepatocytes. clinical signs of acetaminophen toxicity include respiratory distress from lack of oxygen-carrying capacity, cyanosis, methemoglobinemia (chocolate-brown appearance of the blood and mucous membranes), lethargy, vomiting, and facial and paw swelling (cats). the toxic dose of acetaminophen is > mg/kg for dogs, and mg/kg for cats. treatment of acetaminophen toxicity includes induction of emesis or orogastric lavage if the substance has been ingested within minutes. activated charcoal should also be administered. in cases of severe anemia, give supplemental oxygen along with a packed rbc transfusion. administer intravenous fluids to maintain renal and hepatic perfusion. n-acetylcysteine, vitamin c, and cimetidine are the treatments of choice for methemoglobinemia in patients with acetaminophen toxicity. introduction: hydrochloric, nitric, and phosphoric acids cause chemical burns through contact with the skin and/or eyes. localized superficial coagulative necrosis occurs upon contact. usually, the patient's skin is painful to the touch or the animal may lick or chew at an irritated area that is not visible under the haircoat. if the chemical is swallowed, do not induce emesis or perform orogastric lavage, because of the risk of worsening esophageal irritation. rinse the patient's skin and eyes with warm water or warm saline for a minimum of / hour. use analgesics and treat corneal ulcers (see section on corneal ulcers) as required. do not attempt chemical neutralization, because of the risk of causing an exothermic reaction and worsening tissue injury. aflatoxin (aspergillus flavus) is found in moldy feed grains. clinical signs of toxicity occur after ingestion and include vomiting, diarrhea, and acute hepatitis; abortion may occur in pregnant bitches. treatment of suspected aflatoxin ingestion consists of gastric decontamination, administration of activated charcoal, intravenous fluids, and hepatic supportive care (s-adenosyl methionine [same], milk thistle). drinking (ethanol), rubbing (isopropyl), and methyl (methanol) alcohols can be harmful if ingested ( . to . g/kg po). all cause disruption of neuronal membrane structure, impaired motor coordination, cns excitation followed by depression, and stupor that can lead to cardiac and respiratory arrest, depending on the amount ingested. affected animals may appear excited and then ataxic and lethargic. contact or inhalant injury can occur, causing dermal irritation and cutaneous hyperemia. methanol also can cause hepatotoxicity. and diarrhea result from muscarinic overload. nicotinic overload produces muscle tremors. toxicity can result in seizures, coma, and death. and cause severe irritation and corrosion of the mucous membranes and skin. some compounds also can cause clinical signs similar to those observed with anticholinesterase compounds, including muscle tremors, seizures, paralysis, and coma. methemoglobinemia can occur. signs of ethylene glycol intoxication and renal impairment or failure, a negative test for the presence of calcium oxalate crystalluria means that there is no more ethylene glycol in the patient's serum because it has all been metabolized. cats are very sensitive to the toxic effects of ethylene glycol. in many cases, cat may have ingested a toxic dose, but because the sensitivity of the assay is low, test results will be negative. lack of treatment can result in death. there are three phases of ethylene glycol intoxication. in the first to hours after ingestion (stage i), the patient may appear lethargic, disoriented, and ataxic. in stage ii ( to hours following ingestion), the patient improves and appears clinically normal. in stage iii ( to hours following ingestion), the patient demonstrates clinical signs of renal failure (polyuria and polydipsia) that progress to uremic renal failure (vomiting, lethargy, oral ulceration). finally, seizures, coma, and death occur. crosses, old english sheepdogs, and some terriers. clinical signs of ivermectin toxicity include vomiting, ataxia, hypersalivation, agitation, tremors, hyperactivity, hyperthermia, hypoventilation, coma, seizures, signs of circulatory shock, bradycardia, and death. clinical signs often occur within to hours after ingestion or iatrogenic overdose. blood ivermectin levels can be measured, but diagnosis is often made based on clinical signs and knowledge of exposure in predisposed breeds. there is no known antidote. the clinical course can be prolonged for weeks to months before recovery occurs. to treat known exposure, induce emesis or perform orogastric lavage if the substance was ingested was within hour of presentation and the patient is not symptomatic. administer activated charcoal. control seizures with phenobarbital, pentobarbital, or propofol administered as intermittent boluses or as a constant rate infusion. diazepam, which potentially can worsen central nervous stimulation, is contraindicated. administer intravenous fluids to maintain perfusion and hydration, and treat hyperthermia. supportive care may be necessary, including supplemental oxygen (or mechanical ventilation, if necessary), frequent turning of the patient and passive range-of-motion exercises, placement of a urinary catheter to maintain patient cleanliness and monitor urine output, lubrication of the eyes, and parenteral nutrition (see section on rule of twenty). specific antidotes used to treat ivermectin toxicity include physostigmine and picrotoxin. physostigmine therapy was beneficial in some patients for a short period; picrotoxin caused severe violent seizures and therefore should be avoided. introduction d-limonene and linalool are components of citrus oil extracts used in some flea control products. the toxic dose is unknown, but cats appear to be very sensitive to exposure. clinical signs of toxicity include hypersalivation, muscle tremors, ataxia, and hypothermia. treatment of d-limonene and linalool exposure includes treatment of hypothermia, administration of activated charcoal to prevent further absorption, and careful, thorough bathing to prevent further dermal exposure. lead is ubiquitous, and is found in some paints, car batteries, fishing equipment/ sinkers, and plumbing materials. lead can be toxic at doses of mg/kg. if more than than - mg/kg of lead is ingested, death can occur. lead causes toxicity by inhibiting sulfur-containing enzymes, leading to increased rbc fragility, and cns damage. clinical signs of hyperexcitability, dementia, vocalization, seizures, and lower motor neuron polyneuropathy can occur. affected animals may appear blind, or vomiting, anorexia, and constipation or diarrhea may occur. if lead toxicity is suspected, blood and urine lead levels can be measured. treatment of lead toxicity is supportive and is directed at treatment of clinical signs. control seizures with diazepam or phenobarbital. if cerebral edema is present, administer mannitol ( . - . g/kg iv), followed by furosemide ( mg/kg iv minutes after mannitol). sodium or magnesium sulfate should be administered as a cathartic. initiate chelation therapy with dimercaprol, penicillamine, or calcium edta. if a lead object is identified in the gastrointestinal tract on radiographs, remove the object using endoscopy or exploratory laparotomy. hyperthermia, that occurs within - minutes of ingestion. diarrhea and convulsions can develop. if hyperthermia is severe, renal failure secondary to myoglobinuria and disseminated intravascular coagulation can result. delayed hepatic failure has been described days after initial recovery. if metaldehyde toxicosis is suspected, analysis of urine, serum, and stomach contents is warranted. to treat metaldehyde toxicity, procure and maintain a patent airway and control cns excitation and muscle tremors. if an animal has just ingested the metaldehyde and is not symptomatic, induce emesis. if clinical signs are present, perform orogastric lavage. both emesis and orogastric lavage should be followed by administration of one dose of activated charcoal. administer intravenous fluids to control hyperthermia, prevent dehydration, and correct acid-base and electrolyte abnormalities. methocarbamol is the treatment of choice to control muscle tremors. diazepam can be used to control seizures if they occur. introduction mushroom ingestion most commonly causes activation of the autonomic nervous system, resulting in tremors, agitation, restlessness, hyperexcitability, and seizures. in some cases slud (salivation, lacrimation, urination, and defecation) is seen. some mushrooms (amanita spp.) also can cause hepatocellular toxicity. clinical signs include vomiting, anorexia, lethargy, and progressive icterus. hemoglobinuria and pigment damage of the renal tubular epithelium. heinz bodies may be observed on cytologic evaluation of the peripheral blood smear. paint in a sorbitol or glycerol carrier. when large quantities of these osmotically active sugars are ingested, osmotic shifts of fluid cause a sudden onset of neurologic or gastrointestinal signs, including ataxia, seizures, and osmotic diarrhea caused by massive fluid shifts into the gastrointestinal tract. the loss of water in excess of solute can result in hypernatremia, a free water deficit, and increased serum osmolality. following orogastric lavage, treatment of ingestion includes administering warm water enemas to help speed the movement of the paintballs through the gastrointestinal tract. do not administer activated charcoal (usually in a propylene glycol carrier), because the compound's cathartic action will pull more fluid into the gastrointestinal tract. baseline electrolytes should be obtained and then carefully monitored. if severe hypernatremia develops, administer hypotonic solutions such as . % nacl + . % dextrose or % dextrose in water after calculating the patient's free water deficit. because of the large volume of fluid loss, intravenous fluid rates may seem excessive but are necessary to normalize acid-base, electrolyte, and hydration status. in most cases, these patients can survive if the problem is recognized promptly and corrected with careful electrolyte monitoring, aggressive decontamination strategies, and intravenous fluid support. introduction paraquat, a dipyridyl compound, is the active ingredient in some herbicides. the ld of paraquat is - mg/kg. paraquat initially causes cns excitation. it also causes production of oxygen-derived free radical species in the lungs, that can lead to the development of acute respiratory distress syndrome. initial clinical signs include vomiting, diarrhea, and seizures. within to days, clinical signs associated with severe respiratory distress and acute respiratory distress syndrome (ards) can develop, leading to death. chronic effects include pulmonary fibrosis, if the patient survives the initial toxicity period. the prognosis for paraquat toxicity is generally unfavorable. to treat paraquat ingestion, remove the toxin from the gastrointestinal tract as rapidly as possible after ingestion. there are no known antidotes. if the compound was ingested within the past hour and the animal is able to protect its airway, induce emesis. otherwise, perform orogastric lavage. activated charcoal is not as effective as clay or bentonite adsorbents for removing this particular toxin. early in the course of paraquat toxicity, oxygen therapy is contraindicated because of the risk of producing oxygen-derived free radical species. later, oxygen therapy, including mechanical ventilation, is necessary if ards develops. experimentally, free radical scavengers (n-acetyl cysteine, vitamin c, vitamin e, same) have been shown to be useful in preventing damage caused by oxygen-derived free radical species. hemoperfusion may be useful in eliminating the toxin, if it is performed early in the course of toxicity. pennyroyal oil is an herbal flea control compound that contains menthofuran as its toxic compound. menthofuran is hepatotoxic and may cause gastrointestinal hemorrhage and coagulopathies. to treat toxicity, administer a cathartic and activated charcoal and antiemetic and gastroprotectant drugs, and thoroughly bathe the animal to prevent further dermal exposure. petroleum distillates: see fuels phenobarbital: see barbiturates phenylcyclidine (angel dust) introduction phenylcyclidine (angel dust) is an illicit recreational drug that causes both cns depression and excitation, decreased cardiac output, and hypotension. to treat phenylcyclidine toxicity, place an intravenous catheter, and administer intravenous fluids and antiarrhythmic drugs to maintain organ perfusion. administer supplemental oxygen, and administer diazepam to control seizures. urine alkalinization can help eliminate the compound. phenylephrine is an α-adrenergic agonist in many over-the-counter decongestant preparations. clinical signs of intoxication include mydriasis, tachypnea, agitation, hyperactivity, and abnormal flybiting and staring behavior. tachycardia, bradycardia, hypertension, hyperthermia, and seizures can occur. to treat phenylephrine toxicity, place an intravenous catheter and give intravenous fluids to maintain hydration, promote diuresis, and treat hyperthermia. administer prazosin or sodium nitroprusside to treat hypertension, antiarrhythmic drugs as necessary, and diazepam to control seizures. phenylpropanolamine has both αand β-adrenergic agonist effects, and is used primarily in the treatment of urinary incontinence in dogs. the drug was taken off of the market for use in humans because of the risk of stroke. clinical signs of phenylpropanolamine intoxication include hyperactivity, hyperthermia, mydriasis, tachyarrhythmias or bradycardia, hypertension, agitation, and seizures. to treat toxicity, administer prazosin or nitroprusside to control hypertension, a betablocker (esmolol, propranolol, atenolol) to control tachyarrhythmias, diazepam to control seizures, and intravenous fluids to maintain hydration and promote diuresis. urine acidification may aid in facilitating excretion. if bradycardia occurs, do not use atropine. pseudoephedrine is an αand β-adrenergic agonist that is a component of many over-thecounter decongestants and is used in the manufacture of crystal methamphetamine. clinical signs of toxicity include severe restlessness, tremors, mydriasis, agitation, hyperthermia, tachyarrhythmias or bradycardia, hypertension, and seizures. to treat toxicity, administer activated charcoal, intravenous fluids to promote diuresis and treat hyperthermia, chlorpromazine to combat α-adrenergic effects, a beta-blocker (propranolol, esmolol, atenolol) to treat β-adrenergic effects, and cyproheptadine (per rectum) to combat serotoninergic effects. piperazine is a gaba agonist, and causes cervical and truncal ataxia, tremors, seizures, coma, and death. salt used for thawing ice commonly contains calcium chloride, a compound that has a moderate toxic potential. calcium chloride produces strong local irritation and can cause gastroenteritis and gastrointestinal ulcers if ingested. respiratory emergencies consist of any problem that impairs delivery of oxygen to the level of the alveoli or diffusion of oxygen across the alveolar capillary membrane into the pulmonary capillary network. decreased respiratory rate or tidal volume can result in hypoxia and buildup of carbon dioxide, or hypercarbia, leading to respiratory acidosis. conditions most frequently encountered result in airflow obstruction, prevention of normal lung expansion, interference with pulmonary gas exchange (ventilation-perfusion mismatch), and alterations of pulmonary circulation. evaluation of the patient with respiratory distress is often challenging, because the most minimal stress can cause rapid deterioration, or even death in critical cases. careful observation of the patient from a distance often allows the clinician to determine the severity of respiratory distress and localize the lesion based on the patient's respiratory pattern and effort. animals in respiratory distress often have a rapid respiratory rate (> breaths per minute). as respiratory distress progresses, the patient may appear anxious and start openmouth breathing. the animal often develops an orthopneic posture, characterized by neck extension, open-mouthed breathing, and elbows abducted or pulled away from the body. cyanosis of the mucous membranes often indicates extreme decompensation. clinical signs of respiratory distress can develop acutely, or from decompensation of a more chronic problem that was preceded by a cough, noisy respirations, or exercise intolerance. localization of the cause of respiratory distress is essential to successful case management. in any patient with clinical signs of respiratory distress, the differential diagnosis should include primary pulmonary parenchymal disease, airway disease, thoracic cage disorders, congestive heart failure, dyshemoglobinemias (carbon monoxide, methemoglobin), and anemia. careful observation of the patient's respiratory pattern can aid in making a diagnosis of upper airway disease/obstruction, primary pulmonary parenchymal disease, pleural space disease, and abnormalities of the thoracic cage. it is often helpful to rest a hand on the patient and breathe along with the patient's effort, to confirm the periods of inhalation and exhalation. the pharynx, larynx, and extrathoracic trachea comprise the upper airway. obstructive lesions are associated with a marked inspiratory wheeze or stridor and slow deep inspiratory effort. auscultation of the larynx and trachea may reveal more subtle obstructions of normal air flow. stridor can usually be auscultated without the use of a stethoscope. lung sounds are usually normal. the neck should be carefully palpated for a mass lesion, tracheal collapse, and subcutaneous emphysema. subcutaneous emphysema suggests tracheal damage or collapse secondary to severe trauma. in some cases, there is a history of voice, or bark, change secondary to laryngeal dysfunction. differential diagnosis is usually based on the patient's signalment, history, and index of suspicion of a particular disease process. differential diagnoses of upper airway obstruction are listed in box - . diseases of the pleural space often are associated with a restrictive respiratory pattern. inspiratory efforts are short, rapid, and shallow, and there is often a marked abdominal push. the pattern has been referred to as a choppy "dysynchronous" respiratory pattern. depending on the disease present, lung sounds may be muffled ventrally and enhanced dorsally. percussion of the thorax reveals decreased resonance if fluid is present. increased resonance is present with pneumothorax. decreased compressibility of the anterior thorax may be present with an anterior mediastinal mass lesion, particularly in cats and ferrets. a pneumothorax or diaphragmatic hernia is commonly associated with evidence of trauma, with or without rib fractures. respiratory distress due to hemothorax may be exacerbated by anemia. differential diagnoses for patients with evidence of pleural cavity disease include pneumothorax, diaphragmatic hernia, neoplasia, and various types of pleural effusion. primary pulmonary parenchymal disease can involve the intrathoracic airways, alveoli, interstitial space, and pulmonary vasculature. a rapid, shallow, restrictive respiratory pattern may be observed with a marked push on exhalation, particularly with obstructive airway disease such as chronic bronchitis (asthma) in cats. crackles or wheezes are heard on thoracic auscultation. differential diagnoses for pulmonary parenchymal disease include cardiogenic and noncardiogenic pulmonary edema, pneumonia, feline bronchitis (asthma), pulmonary contusion, aspiration pneumonitis, pulmonary thromboembolism, neoplasia, infection (bacterial, fungal, protozoal, viral) , and/or chronic bronchitis. other abnormal respiratory patterns may be evident, and warrant further consideration. tachypnea present in the absence of other signs of respiratory distress can be a normal response to nonrespiratory problems, including pain, hyperthermia, and stress. a restrictive respiratory pattern with minimal thoracic excursions can be associated with diseases of neuromuscular function, including ascending polyradiculoneuritis, botulism, and tick paralysis. if adequate ventilation cannot be maintained by the patient, mechanical ventilation may be indicated. kussmaul respiration manifests as very slow, very deep respirations when a metabolic acidosis is present. this type of respiratory pattern typically is observed in patients with severe diabetic ketoacidosis and renal failure in a compensatory attempt to blow off carbon dioxide. cheyne-stokes respiration is usually observed with a defect in the central respiratory control center. the classic pattern of cheyne-stokes respiration is normal or hyperventilation followed by a period of apnea or hypoventilation. in cases of lower cervical cord damage or damage to the central respiratory control center in the cns, the diaphragm alone may assume most of the ventilatory movement. with diaphragmatic fatigue, severe hypoventilation and resultant hypoxemia may require mechanical ventilation. immediate management of any patient in respiratory distress is to minimize stress at all costs. relatively benign procedures such as radiography or intravenous catheter placement can be fatal in patients with severe respiratory compromise. stabilization should always precede further diagnostic evaluation. in some cases, sedation may be required before performing any diagnostics, to prevent further stress. all patients should receive some form of supplemental oxygen, either by mask, cage, or flow-by techniques. in cases in which a severe pneumothorax or pleural effusion is suspected, perform therapeutic and diagnostic thoracocentesis bilaterally to allow lung re-expansion and alleviate respiratory distress, whenever possible. if thoracocentesis alone is not effective at maintaining lung re-expansion, place a thoracostomy tube (particularly in cases of tension pneumothorax). if hypovolemic/ hemorrhagic shock is present, initiate treatment while stabilizing the respiratory system (see section on shock). if an animal is suspected of having an upper airway obstruction, reestablish airflow. in cases of laryngeal paralysis, tracheal collapse, and brachycephalic airway syndrome, sedation is often very useful in alleviating the distress of airway obstruction. in cases of laryngeal collapse, however, sedation may make the condition worse. if laryngeal edema is severe, administer a dose of short-acting glucocorticosteroids (dexamethasone sodium phosphate) to decrease laryngeal inflammation and edema. if a foreign body is lodged in the pharynx, perform the heimlich maneuver by thrusting bluntly several times on the patient's sternum. objects such as balls or bones may be small enough to enter the larynx but too large to be expelled, and will require rapid-acting general anesthesia to facilitate dislodgement and removal. if the obstruction cannot be removed, bypassing the obstruction with an endotracheal tube or temporary tracheostomy should be considered. in an emergency, a temporary transtracheal oxygen catheter can quickly be placed in the following manner. connect a -or -gauge needle to a length of intravenous extension tubing and a -ml syringe. place the male connector of the syringe into the female portion of the extension tubing. cut off the syringe plunger and connect the resulting blunt end to a length of flexible tubing attached to a humidified oxygen source. run the oxygen at l/minute to provide adequate oxygenation until a tracheostomy can be performed. (see sections on oxygen supplementation and tracheostomy). once the animal's condition has been stabilized, specific diagnostic tests, including arterial blood gas analyses, thoracic radiographs, and/or transtracheal wash, can be performed, depending on the patient's condition and needs. specific therapies for management of upper airway obstruction, pleural space disease, and pulmonary disease are discussed next. upper airway obstruction can occur as a result of intraluminal or extraluminal mass lesions or foreign bodies in the oropharynx (abscess, neoplasia), laryngeal paralysis, trauma, and anatomic abnormalities. clinical signs of an upper airway obstruction are associated with an animal's extreme efforts to inhale air past the obstruction. marked negative pressure occurs in the extrathoracic airways and can cause worsening of clinical signs. mucosal edema and inflammation further worsen the obstruction. therapy for upper airway obstruction is aimed at breaking the cycle of anxiety and respiratory distress. administer the anxiolytic tranquilizer acepromazine ( . - . mg/kg iv, im, sq) to decrease patient anxiety. many animals develop hyperthermia from increased respiratory effort and extreme anxiety. implement cooling measures in the form of cool intravenous fluids and wet towels soaked in tepid water placed over the animal (see section on hyperthermia). administer supplemental oxygen in a manner that is least stressful for the animal. short-acting glucocorticosteroids can also be administered (dexamethasone sodium phosphate, . mg/kg iv, sq, im) to decrease edema and inflammation. if the airway obstruction is severe and there is no response to initial measures to alleviate anxiety and decrease inflammation, establish control of ventilation by placement of an endotracheal tube (see section on endotracheal intubation), tracheal oxygen catheter, or temporary tracheostomy. to obtain airway control, administer a rapid-acting anesthetic (propofol, - mg/kg iv to effect), and intubate with a temporary tracheostomy. an intratracheal oxygen catheter can be placed with sedation and/or a local anesthetic (see technique for transtracheal wash). laryngeal paralysis is a congenital or acquired condition that occurs primarily in largebreed dogs secondary to denervation of the arytenoid cartilages by the recurrent laryngeal nerve. congenital laryngeal paralysis occurs in the bouvier des flandres, siberian husky, and bull terrier. acquired laryngeal paralysis occurs in labrador retrievers, saint bernards, and irish setters. acquired laryngeal paralysis can be idiopathic, acquired secondary to trauma to the recurrent laryngeal nerve, or can be a component of systemic neuromuscular disease. although rare, this condition also occurs in cats. with dysfunction of the recurrent laryngeal nerve, the intrinsic laryngeal muscles atrophy and degenerate. as a result, the vocal folds and arytenoid cartilage move in a paramedian position within the airway and fail to abduct during inhalation, causing airway obstruction. laryngeal paralysis can be partial or complete, unilateral or bilateral. in many cases, a change in bark is noted prior to the development of clinical signs of respiratory distress or exercise intolerance. when a patient presents with severe inspiratory stridor (with or without hyperthermia) initiate stabilization with anxiolytic tranquilizers, supplemental oxygen, and cooling measures. once the patient's condition has been stabilized, definitive measures to accurately document and assess the patient's airway should be considered. place the patient under very heavy sedation with short-acting barbiturates or propofol ( - mg/kg iv) and observe the arytenoid cartilages closely in all phases of respiration. administer just enough drug to allow careful examination without getting bitten. if the arytenoid cartilages do not abduct during inhalation, administer dopram (doxapram hydrochloride, - mg/kg iv) to stimulate respiration. absent or paradoxical laryngeal motion (closed during inspiration and open during exhalation) is characteristic of laryngeal paralysis. correction of the defect involves documentation and treatment of any underlying disorder and surgical repair of the area to open the airway. partial laryngectomy, arytenoid lateralization ("tie-back" surgery), or removal of the vocal folds has been used with some success. aspiration pneumonitis is common following these procedures. brachycephalic airway syndrome is associated with a series of anatomic abnormalities that collectively increase resistance to airflow. affected animals typically have stenotic nares, an elongated soft palate, and a hypoplastic trachea. components of the syndrome can occur alone or in combination. in severe cases, laryngeal saccular edema and eversion, and eventual pharyngeal collapse, can occur secondary to the severe increase in intrathoracic airway pressure required to overcome the resistance of the upper airways. specific airway anomalies can be identified with general anesthesia and laryngoscopy. severe respiratory distress should be treated as discussed previously. treatment requires surgical correction of the anatomic abnormalities. in animals with laryngeal collapse, surgical correction may not be possible, and a permanent tracheostomy may be required. because an elongated soft palate and stenotic nares can be identified before the onset of clinical signs, surgical correction to improve airflow when the animal is young may decrease the negative intra-thoracic pressure necessary to move air past these obstructions. the chronic consequences of everted laryngeal saccules and laryngeal collapse potentially can be prevented. tracheal collapse is common in middle-aged and older toy and small-breed dogs. the owner typically reports a chronic cough that is readily induced by excitement or palpation of the trachea. the cough often sounds like a "goose honk." diagnostic confirmation is obtained by lateral radiography or fluoroscopy of the cervical and thoracic trachea during all phases of respiration. acute decompensation is uncommon but does occur, particularly with excitement, exercise, and increased environmental temperatures or ambient humidity. therapy of the patient with acute respiratory distress secondary to tracheal collapse includes sedation, administration of supplemental oxygen, and provision of cooling measures to treat hyperthermia. cough suppressants (hydrocodone bitartrate-homatropine methylbromide, . mg/kg po q - h, or butorphanol, . mg/kg po q - h) are useful. tracheal collapse is a dynamic process that usually involves both the upper and lower airways. because of this, bypassing the obstruction is often difficult. tracheal stents have been emergency care used with limited success in combination with treatment of chronic lower airway disease. crush or bite injuries to the neck can result in fractures or avulsion of the laryngeal or tracheal cartilages. bypassing the obstructed area may be necessary until the patient is stable and can undergo surgical correction of the injury. if there is avulsion of the cranial trachea, it may be difficult to intubate the patient. a long, rigid urinary catheter can be inserted past the area of avulsion into the distal segment, and an endotracheal tube passed over the rigid catheter, to establish a secure airway. neck injury can also result in damage to the recurrent laryngeal nerve and laryngeal paralysis. foreign bodies can lodge in the nasal cavity, pharynx, larynx, and distal trachea. signs of foreign bodies in the nares include acute sneezing and pawing at or rubbing the muzzle on the ground. if the object is not removed, sneezing continues and a chronic nasal discharge develops. respiratory distress is uncommon, but the foreign body is severely irritating. pharyngeal and tracheal foreign bodies can cause severe obstruction to airflow and respiratory distress. diagnosis of a foreign body is based on the patient history, physical examination findings, and thoracic or cervical radiographs. smaller foreign bodies lodged in the distal airways may not be apparent radiographically but can cause pulmonary atelectasis. foreign bodies of the nose or pharynx can often be removed with an alligator forceps with the patient under anesthesia. if removal is not possible with a forceps, flushing the nasal cavity from cranial to caudal (pack the back of the mouth with gauze to prevent aspiration) can sometimes dislodge the foreign material into the gauze packing. rhinoscopy may be necessary. if an endoscope is not available, an otoscope can be used. foreign objects lodged in the trachea can be small and function like a ball valve during inhalation and exhalation, causing episodic hypoxia and collapse. when attempting to remove these objects, suspend the patient with its head down. remove the object with an alligator forceps, using a laryngoscope to aid in visualization. foreign bodies lodged in the trachea or bronchi require removal with endoscopic assistance. nasopharyngeal polyps (in cats, tumors, obstructive laryngitis, granulomas, abscesses, and cysts) can cause upper airway obstruction. clinical signs are usually gradual in onset. the lesions can be identified through careful laryngoscopic examination performed with the patient under general anesthesia. the nasopharynx above the soft palpate should always be included in the examination. pedunculated masses and cysts are excised at the time of evaluation. biopsy of diffusely infiltrative masses is indicated for histologic examination and prognosis. it is impossible to distinguish obstructive laryngitis from neoplasia based on gross appearance alone. whenever possible, material should be collected from abscesses and granulomas for cytologic evaluation and bacterial culture. extraluminal masses impinge on and slowly compress the upper airways, resulting in slow progression of clinical signs. masses are usually identified by palpation of the neck. enlarged mandibular lymph nodes, thyroid tumors, and other neoplasms may be present. diagnosis is usually based on a combination of radiography and ultrasonography. ct and/or mri are helpful in identifying the full extent and invasiveness of the lesion. definitive diagnosis is made with a fine-needle aspirate or biopsy. many thyroid tumors bleed excessively. the inside of each side of the hemithorax is covered in parietal pleura. the lung lobes are covered in visceral pleura. the two surfaces are in close contact with each other, and are contiguous at the hilum under normal circumstances. pneumothorax refers to free air within the pleural space, accumulating in between the parietal and visceral pleura. the term pleural effusion refers to fluid accumulation in that area but does not reflect the amount or type of fluid present. the mediastinal reflections of the pleura typically are thin in dogs and cats, and usually, but not always, connect. bilateral involvement of pneumothorax or pleural effusion is common. both pneumothorax and pleural effusion compromise the lungs' ability to expand and result in hypoxia and respiratory distress. pneumothorax can be classified as open versus closed, simple versus complicated, and tension. an open pneumothorax communicates with the external environment through a rent in the thoracic wall. a closed pneumothorax results from tears in the visceral pleura but does not communicate with the outside. a tension pneumothorax occurs as a result of a tear in the lung or chest wall that creates a flap valve, such that air is allowed to leave the lung and accumulate in the pleural space during inhalation, and closes to seal off exit of air from the pleural space during exhalation. tension pneumothorax can cause rapid decline in cardiopulmonary status and death if not recognized and treated immediately. a simple pneumothorax is one that can be controlled with a simple thoracocentesis. complicated pneumothorax involves repeated accumulation of air, requiring placement of a thoracic drainage catheter. in many cases, pneumothorax develops as a result of trauma. spontaneous pneumothorax occurs with rupture of cavitary lesions of the lung that may be congenital or acquired as a result of prior trauma, heartworm disease, airway disease (emphysema), paragonimiasis, neoplasia, or lung abscess. pneumothorax also rarely occurs as a result of esophageal tears or esophageal foreign bodies. rapid circulatory and respiratory compromise following traumatic pneumothorax can develop as a result of open or tension pneumothorax, rib fractures, airway obstruction, pulmonary contusions, hemothorax, cardiac dysrhythmias, cardiac tamponade, and hypovolemic shock. any patient that is rapidly decompensating after a traumatic episode must be quickly assessed, and emergency therapy initiated (see section on immediate management of trauma, a crash plan). diagnosis of pneumothorax is usually made based on a history of trauma, a rapid, shallow, restrictive respiratory pattern, and muffled heart and lung sounds on thoracic auscultation. the clinical signs and history alone should prompt the clinician to perform a bilateral diagnostic and therapeutic thoracocentesis before taking thoracic radiographs (see section on thoracocentesis). the stress of handling the patient for radiography can be deadly in severe cases of pneumothorax. although the mediastinum on both sides of the thorax connects, it is necessary to perform thoracocentesis on both sides to ensure maximal removal of free air in the pleural space and allow maximal lung expansion. if negative pressure cannot be obtained, or if the patient rapidly reaccumulates air, place a thoracostomy tube connected to continuous suction. (see section on thoracostomy tube placement). treat all penetrating wounds to the thorax as open sucking chest wounds unless proved otherwise. to "close" an open sucking chest wound, clip the fur around the wound as quickly as possible, and place sterile lubricant jelly or antimicrobial ointment circumferentially around the wound. cut a sterile glove to provide a covering. place the covering over the wound, making sure to cover all of the sterile lubricant, thus creating a seal to close the wound temporarily from the external environment. evaluate the patient's thorax via thoracocentesis while placing a thoracostomy tube. once the patient is stable, the open chest wound can be surgically explored, lavaged, and definitively corrected. all animals with open chest wounds should receive antibiotics (first-generation cephalosporin) to prevent infection. following stabilization, radiographs can be taken and evaluated. pneumothorax is confirmed by evidence of elevation of the cardiac silhouette above the sternum, increased density of the pulmonary parenchymal tissue, free air in between the parietal and visceral pleura (making the outline of the lungs visible), and absence of pulmonary vascular structures in the periphery. parenchymal lesions within the lungs are best identified after as much air as possible has been removed from the thorax. obtain left and right lateral and ventrodorsal or dorsoventral views. a standing lateral view may reveal air-or fluid-filled cavitary masses. if underlying pulmonary disease is suspected as a cause of spontaneous pneumothorax, a transtracheal wash, fecal flotation, and heartworm test may be indicated. treatment of pneumothorax includes immediate bilateral thoracocentesis, covering of any open chest wounds, administration of supplemental oxygen, and placement of a thoracostomy tube if negative pressure cannot be obtained or if air rapidly reaccumulates. serial radiography, ct, or mri should be performed in dogs with spontaneous pneumothorax, because the condition can be associated with generalized pulmonary parenchymal disease. strict cage rest is required until air stops accumulating and the thoracostomy tube can be removed. the patient's chest tube should be aspirated every hours after discontinuing continuous suction. if no air reaccumulates after hours, the chest tube can be removed. exercise restriction is indicated for a minimum of week. if bullae or mass lesions are present, exploratory thoracotomy should be considered as a diagnostic and potentially therapeutic option for long-term management in prevention of recurrence. pleural fluid cytologic analysis is indicated for all patients with pleural effusion before administration of antibiotics. the general term pleural effusion means a collection of fluid in the space between the parietal and visceral pleura but does not indicate what kind or how much fluid is present. clinical signs associated with pleural effusion depend on how much fluid is present, and how rapidly the fluid has accumulated. clinical signs associated with pleural effusion include respiratory distress, reluctance to lie down, labored breathing with an abdominal component on exhalation, cough, and lethargy. auscultation of the thorax may reveal muffled heart and lung sounds ventrally and increased lung sounds dorsally, although pockets of fluid may be present, depending on the chronicity of the effusion. percussion of the thorax may reveal decreased resonance. in stable patients, the presence of pleural effusion can be confirmed radiographically. radiographic confirmation of the pleural effusion should include right and left lateral and dorsoventral or ventrodorsal views. a handling or standing lateral view should be obtained if an anterior mediastinal mass is suspected. the standing lateral view will allow the fluid to collect in the costophrenic recess. in patients with respiratory distress, muffled heart and lung sounds, and suspicion of pleural effusion, thoracocentesis should be performed immediately. thoracocentesis can be both therapeutic and diagnostic. radiography is contraindicated because the procedure can cause undue stress and exacerbation of clinical signs in an unstable patient. pleural effusion can cause severe respiratory distress, and can be the result of a number of factors that must be considered when implementing an appropriate treatment plan. pathology of the pleura is almost always a secondary process except for primary bacterial pleuritis and pleural mesotheliomas. causes of pleural effusion in the cat and dog include pyothorax, feline infectious peritonitis, congestive heart failure, chylothorax, heartworm disease, hemothorax, hypoalbuminemia, lung lobe torsions, neoplasia, diaphragmatic hernia, and pancreatitis (box - ). in stable animals, diagnosis of pleural effusion can be made based • imbalance of transpleural or hydrostatic or protein osmotic forces • change in membrane permeability • decrease in rate of fluid reabsorption • combination of foregoing mechanisms on thoracic radiography or ultrasound. thoracic radiographs can show whether the pleural effusion is unilateral or bilateral. effusions in dogs and cats are usually bilateral. the lung parenchyma and the cardiac silhouette cannot be fully evaluated until most of the fluid has been evacuated from the pleural cavity. following thoracocentesis, radiography should be performed with left and right lateral and ventrodorsal or dorsoventral views. in cases of suspected heart failure, echocardiography also is necessary. pleural fluid cytologic analysis is indicated for all patients with pleural effusion. collect specimens before administering antibiotics, whenever possible, because treatment with antibiotics can make a septic condition (pyothorax) appear nonseptic. the remainder of the diagnostic workup and treatment is based on the type of fluid present (table - ). the fluid may be a transudate, nonseptic exudate, septic exudate, chylous, hemorrhagic, or neoplastic. ultrasonographic evaluation of the thorax can be helpful in identifying intrathoracic masses, diaphragmatic hernias, lung lobe torsions, and cardiac abnormalities. unlike radiography, ultrasonography is facilitated by the presence of fluid in the pleural space. pyothorax refers to a septic effusion of the pleural cavity. the infection is generally the result of a combination of aerobic and anaerobic bacteria. rarely, fungal organisms are present. the source of the underlying organisms is rarely identified, particularly in cats, but can be caused by penetrating wounds through the chest wall, esophagus, migrating foreign bodies (especially grass awns), or primary lung infections. the most common organisms associated with pyothorax in the cat are pasteurella, bacteroides, and fusobacterium. fever is often present in addition to clinical signs of pleural effusion. septic shock is ununcommon. diagnosis of pyothorax is made based on cytologic analysis and the demonstration of intracellular and extracellular bacteria, toxin neutrophils and macrophages, and sometimes the presence of sulfur granules. gram stains of the fluid can assist in the initial identification of some organisms. bacterial cultures are indicated for bacteria identification and antibiotic susceptibility testing. administration of antibiotics before cytologic evaluation can cause a septic effusion to appear nonseptic. emergency treatment for pyothorax involves placement of an intravenous catheter, intravenous fluids to treat hypovolemic shock, and broad-spectrum antibiotics (ampicillin, mg/kg iv q h, and enrofloxacin, mg/kg iv q h). chloramphenicol also is an appropriate antibiotic to use for penetration into pockets of fluid. administration of a beta-lactam antibiotic (ampicillin or amoxicillin) with a beta-lactamase inhibitor (amoxicillin clavulanate or ampicillin sulbactam) is helpful in achieving better coverage of bacteroides spp. treatment of pyothorax differs in the cat and dog. in the cat, placement of one or two thoracic drainage catheters is recommended to allow continuous drainage of the intrathoracic abscess. inadequate drainage can result in treatment failure. fluid should be evaluated and the pleural cavity lavaged with ml/kg of warmed . % saline or lactated ringer's solution every hours. approximately % of the infused volume should be recovered after each lavage. in dogs, or in cats with refractory pyothorax, perform an exploratory thoracotomy to remove any nidus of infection. rarely a foreign body is visible that can be removed at the time of surgery, but this finding is rare. antibiotics are indicated for a minimum of to weeks after removal of the thoracostomy tube. early diagnosis and aggressive treatment result in a good prognosis in the majority of patients with pyothorax. in cats, clinical signs of ptyalism and hypothermia at the time of presentation worsen the prognosis. chylothorax refers to the abnormal accumulation of chyle (lymphatic fluid) in the pleural cavity. the cisterna chili is the dilated collection pool of lymphatic ducts in the abdomen that accumulate chyle prior to entry into the thoracic duct located within the thoracic cavity. the thoracic duct enters the thorax at the aortic hiatus. numerous tributaries or collateral ducts exist. the functions of the lymphatic vessels collectively serve to deliver triglycerides and fat-soluble vitamins into the peripheral vascular circulation. damage of the thoracic duct or lymphatic system or obstruction to lymphatic flow can result in the development of chylous effusion in the pleural or peritoneal space. it is difficult to identify chylous effusions based on their milky appearance alone. to identify a chylous effusion versus a pseudochylous effusion, the triglyceride and cholesterol levels of the fluid must be compared with those of peripheral blood. chylous effusions have a higher triglyceride and lower cholesterol levels than peripheral blood. pseudochylous effusions have a higher cholesterol and lower triglyceride levels than peripheral blood. disease processes that can result in chylous effusions are listed in the box - . clinical signs associated with chylous effusion are typical of any pleural effusion and of the disease process that caused the effusion. weight loss may be evident, depending on the chronicity of the process. the diagnosis is made based on thoracocentesis, cytology, and biochemical evaluation of the fluid (i.e., triglyceride and cholesterol levels). the fluid often appears milky or bloodtinged but can be clear if the patient has significant anorexia. typical cytologic characteristics are listed in table - . lymphangiography can be used to confirm trauma to the thoracic duct, but this is usually not necessary unless surgical ligation is going to be attempted. the diagnostic evaluation must also attempt to identify an underlying cause. therapy for chylothorax is difficult and primarily involves documentation and treatment of the underlying cause. if an underlying cause is not found, treatment is largely supportive and consists of intermittent thoracocentesis to drain the fluid as it accumulates and causes respiratory dysfunction, nutritional support, and maintenance of fluid balance. a variety of surgical techniques, including ligation of the thoracic duct, pleural-peritoneal shunts, and pleurodesis, have been attempted but have had limited success. most recently, the combination of thoracic duct ligation with subtotal pericardectomy has been shown to improve surgical success rates in the treatment of chylothorax. rutin, a bioflavinoid, has been used with limited success in the treatment of idiopathic chylothorax in cats. prognosis in many cases of chylothorax is guarded. extensive hemorrhage into the pleural cavity can cause fulminant respiratory distress due to sudden hypovolemia and anemia and interference with lung expansion. hemothorax typically is associated with trauma, systemic coagulopathy, lung lobe torsions, and erosive lesions within the thorax (usually neoplasia). diagnosis of hemothorax involves obtaining a fluid sample via thoracocentesis. hemorrhagic effusion must be differentiated from systemic blood inadvertently collected during the thoracocentesis procedure. unless the hemorrhage is peracute, fluid in cases of hemothorax is rapidly defibrinated and will not clot, has a packed cell volume less than that of venous blood, contains rbcs and macrophages. hemorrhagic effusions also usually contain a disproportionately higher number of white blood cells compared with peripheral blood. hemothorax commonly is the sole clinical sign observed in animals with vitamin k antagonist rodenticide intoxication and systemic coagulopathy. whenever an animal presents with signs of a hemorrhagic pleural effusion, perform coagulation testing immediately to determine whether a coagulopathy exists. the prothrombin time test is fast and can be performed as a cage-side test (see section on coagulopathy). therapy for hemorrhagic pleural effusions should address the blood and fluid loss. administer intravenous crystalloid fluids and rbc products (see section on transfusion therapy). when necessary, administer coagulation factors in the form of fresh whole blood or fresh frozen plasma, along with vitamin k ( mg/kg sq in multiple sites with a -gauge needle). if severe respiratory distress is present, evacuate the blood within the pleural space via thoracocentesis until clinical signs of respiratory distress resolve. fluid that remains aids in the recovery of the patient, because rbcs and proteins eventually will be reabsorbed. autotransfusion can be performed to salvage blood and reinfuse it into the anemic patient. in cases of neoplastic or traumatic uncontrollable hemorrhagic effusions, surgical exploration of the thorax is warranted. diaphragmatic hernia, or a rent in the diaphragm, can result in the protrusion of abdominal organs into the thoracic cavity and impair pulmonary expansion. organs that are commonly herniated into the thorax include the liver, stomach, and small intestines. diaphragmatic hernia usually is secondary to trauma but can occur as a congenital anomaly. in cases of trauma, rib fractures, pulmonary contusions, traumatic myocarditis, hemothorax, and shock are also often present concurrently with diaphragmatic hernia. respiratory distress can be caused by any one or a combination of the above lesions. animals with prior or chronic diaphragmatic hernias may have minimal clinical signs despite the presence of abdominal organs within the thorax. clinical signs of acute or severe diaphragmatic hernia include respiratory distress, cyanosis, and shock. a diagnosis of diaphragmatic hernia is made based on the patient's history (traumatic event), clinical signs, and radiographs. in some cases, ultrasonography or contrast peritoneography is necessary to confirm the diagnosis. contrast radiographs may show the presence of the stomach or intestines within the thorax following oral administration of barium. never administer barium directly into the peritoneal cavity or in cases of suspected gastrointestinal rupture. treatment of a patient with a diaphragmatic hernia includes cardiovascular and respiratory system stabilization before attempting surgical repair of the diaphragm. if the stomach is within the thorax, or if the patient's respiratory distress cannot be alleviated with medical management alone, immediate surgery is necessary. if the respiratory distress is minimal and the stomach is not located within the thorax, surgery can be postponed until the patient is a more stable anesthetic candidate. at the time of surgery, the abdominal organs are replaced into the abdominal cavity, and the rent in the diaphragm is closed. air must be evacuated from the thorax following closure of the diaphragm. if chronic diaphragmatic hernia is repaired, the complication of reexpansion pulmonary edema can occur. cardiac injury is a common complication secondary to blunt thoracic trauma. in most cases, cardiac injury is manifested as arrhythmias, including multiple premature ventricular contractions, ventricular tachycardia, st segment depression or elevation secondary to myocardial hypoxemia, and atrial fibrillation (see section on cardiac emergencies). myocardial infarction and cardiac failure can occur. careful and repeated assessments of the patient's blood pressure and ecg tracing should be a part of any diagnostic work-up for a patient that has sustained blunt thoracic trauma. rib fractures are associated with localized pain and painful respiratory movements. radiographs are helpful to confirm the diagnosis. careful palpation may reveal crepitus and instability of the fractured ribs. common problems associated with rib fractures emergency care include pulmonary contusions, pericardial laceration, traumatic myocarditis, diaphragmatic hernia, and splenic laceration or rupture. a flail segment results from rib fractures of more than three adjacent ribs that produce a "floating segment" of the chest wall. the flail segment moves paradoxically with respiration-that is, it moves inward during inhalation and outward during exhalation. respiratory distress is associated with the pain caused by the fractures and the presence of traumatic underlying pulmonary pathology. therapy for rib fractures and flail chest includes administration of supplemental oxygen, treatment of pneumothorax or diaphragmatic hernia, and administration of systemic and local anesthesia to alleviate the discomfort associated with the fractures. although controversial, positioning the patient with the flail segment up may reduce pain and improve ventilation. avoid the use of chest wraps, which do nothing to stabilize the flail segment and can further impair respiratory excursions. following administration of a systemic analgesic, administer a local anesthetic at the dorsocaudal and ventrocaudal segment of each fractured rib, and in one rib in front of and behind the flail segment. often, pulmonary function will improve once the pain associated with rib fractures has been adequately treated. in rare cases in which the flail segment involves five or more ribs, surgical stabilization may be necessary. single rib fractures or smaller flail segments are allowed to heal on their own. feline bronchitis has a variety of names (bronchial asthma, asthma, acute bronchitis, allergic bronchitis, chronic asthmatic bronchitis, feline lower airway disease) and refers to the acute onset of respiratory distress secondary to narrowing of the bronchi. cats may present with an acute onset of severe restrictive respiratory pattern associated with lower airway obstruction. acute bronchitis in cats typically has an inflammatory component in the lower airways, resulting in acute bronchoconstriction, excessive mucus production, and inflammatory exudates. in cats with chronic bronchitis, there may be damage of the bronchial epithelium and fibrosis of the airways. these patients often have a history if intermittent exacerbation of clinical signs, intermittent cough, and periods of normality throughout the year. because there appears to be an allergic or inflammatory component in feline bronchitis, clinical signs can be acutely exacerbated by stress and the presence of aerosolized particles such as perfume, smoke, and carpet powders. causes of feline bronchitis include heartworm disease, parasitic infestation (lungworms), and (rarely) bacterial infection. on presentation, the patient should be placed in an oxygen cage and allowed to rest while being observed from a distance. postpone performing stressful diagnostic procedures until the patient's respiratory status has been stabilized. after careful thoracic auscultation, administer a short-acting bronchodilator (terbutaline, . mg/kg sq or im) along with a glucocorticosteroid (dexamethasone sodium phosphate mg/kg im, sq, iv) to alleviate immediate bronchospasm and airway inflammation. clinical signs of feline bronchitis are characterized by a short, rapid respiratory pattern with prolonged expiration with an abdominal push. wheezes may be heard on thoracic auscultation. in some cases, no abnormalities are found on auscultation, but become acutely worse when the patient is stimulated to cough by tracheal palpation. radiographs may reveal a hyperinflated lung field with bronchial markings and caudal displacement of the diaphragm. in some cases, consolidation of the right middle lung lobe is present. a complete blood count and serum biochemistry profile can be performed, but results usually are unrewarding. in endemic areas, a heartworm test is warranted. fecal examination by flotation and the baermann technique is helpful in ruling out lungworms and other parasites. bronchoalveolar lavage or transtracheal wash is useful for cytologic and bacterial examination. long-term management of feline bronchitis includes isolation from environmental exposure to potential allergens (litter dust, perfumes, smoke, incense, carpet powders) and treatment of bronchoconstriction and inflammation with a combination of oral and inhaled glucocorticosteroids and bronchodilators (table - ). antibiotic therapy is contraindicated unless a pure culture of a pathogen is documented. oral therapy with steroids and bronchodilators should be used for a minimum of weeks after an acute exacerbation and then gradually decreased to the lowest dose possible to alleviate clinical signs. metered dose inhalers are now available (aerokat.com) for administration of inhaled bronchodilators and steroids. fluticasone (flovent, mcg/puff ) can be administered initially every hours for week and then decreased to once daily, in most cases. inhaled glucocorticosteroids are not absorbed systemically, and therefore patients do not develop the adverse side effects sometimes documented with oral glucocorticosteroid administration. because it takes time for glucocorticosteroids to reach peak effects in the lungs, administration of inhaled glucocorticosteroids should overlap with oral prednisolone administration for to days. treatment of pulmonary contusions is supportive. administer supplemental oxygen in a manner that is least stressful for the animal. arterial blood gas analysis or pulse oximetry can determine the degree of hypoxemia and monitor the response to therapy. intravenous fluids should be administered with caution to avoid exacerbating pulmonary hemorrhage or fluid accumulation in the alveoli. treat other conditions associated with the traumatic event. possible complications of pulmonary contusions are rare but include bacterial infection, abscessation, lung lobe consolidation, and the development of cavitary lesions. the routine use of antibiotics or steroids in cases of pulmonary contusions is contraindicated unless external wounds are present. empiric antibiotic use without evidence of external injury or known infection can potentially increase the risk of a resistant bacterial infection. steroids have been shown to decrease pulmonary alveolar macrophage function and impair wound healing and are contraindicated. aspiration pneumonia can occur in animals as a result of abnormal laryngeal or pharyngeal protective mechanisms or can be secondary to vomiting during states of altered mentation, including anesthesia, recovery from anesthesia, and sleep. megaesophagus, systemic polyneuropathy, myasthenia gravis, and localized oropharyngeal defects such as cleft palate can increase the risk of developing aspiration pneumonitis. iatrogenic causes of aspiration pneumonia include improper placement of nasogastric feeding tubes, overly aggressive force-feeding, and oral administration of drugs. aspiration of contents into the airways can cause mechanical airway obstruction, bronchoconstriction, chemical damage to the alveoli, and infection. severe inflammation and airway edema are common. pulmonary hemorrhage and necrosis can occur. diagnosis of aspiration pneumonia is based on clinical signs of pulmonary parenchymal disease, a history consistent with vomiting or other predisposing causes, and thoracic radiographs demonstrating a bronchointerstitial to alveolar pulmonary infiltrate. the most common site is the right middle lung lobe, although the pneumonia can occur anywhere, depending on the position of the patient at the time of aspiration. a transtracheal wash or bronchoalveolar lavage is useful for bacterial culture and susceptibility testing. treatment of aspiration pneumonia includes antibiotic therapy for the infection, administration of supplemental oxygen, and loosening the debris in the airways. administer intravenous fluids to maintain hydration. nebulization with sterile saline and chest physiotherapy (coupage) should be performed at least every hours. antibiotics to consider in the treatment of aspiration pneumonia include ampicillin/enrofloxacin, amoxicillinclavulanate, ampicillin-sulbactam, trimethoprim sulfa, and chloramphenicol. the use of glucocorticosteroids is absolutely contraindicated. continue antibiotic therapy for a minimum of weeks after the resolution of radiographic signs of pneumonia. pulmonary edema arises from the accumulation of fluid in the pulmonary interstitial alveolar spaces, and airways. ventilation-perfusion abnormalities result in hypoxia. pulmonary edema can be caused by increased pulmonary vasculature hydrostatic pressure, decreased pulmonary oncotic pressure, obstruction of lymphatic drainage, or increased capillary permeability. multiple factors can occur simultaneously. the most common cause of edema is increased pulmonary hydrostatic pressure resulting from left-sided congestive heart failure. decreased plasma oncotic pressure with albumin < . g/dl can also result in accumulation of fluid in the pulmonary parenchyma. overzealous intravenous crystalloid fluid administration can result in dilution of serum oncotic pressure and vascular overload. obstruction of lymphatic drainage is usually caused by neoplasia. other causes of pulmonary edema include pulmonary thromboembolic disease, severe upper airway obstruction (noncardiogenic pulmonary edema), seizures, and head trauma. increased capillary permeability is associated with a variety of diseases that cause severe inflammation (systemic inflammatory response syndrome). the resultant pulmonary edema contains a high amount of protein and is known as acute respiratory distress syndrome (ards). ards can be associated with pulmonary or extrapulmonary causes, including direct lung injury from trauma, aspiration pneumonia, sepsis, pancreatitis, smoke inhalation, oxygen toxicity, electrocution, and immune-mediated hemolytic anemia with disseminated intravascular coagulation. diagnosis of pulmonary edema is made based on clinical signs of respiratory distress and the presence of crackles on thoracic auscultation. in severe cases, cyanosis and fulminant blood-tinged frothy edema fluid may be present in the mouth and nostrils. immediate management includes administration of furosemide ( - mg/kg iv, im) and supplemental oxygen. sedation with low-dose morphine sulfate ( . - . mg/kg iv) is helpful in dilating the splanchnic capacitance vasculature and relieving anxiety for the patient. if fluid overload is suspected secondary to intravenous fluid administration, fluids should be discontinued. severely hypoalbuminemic patients should receive concentrated human albumin ( ml/kg of a % solution) or fresh frozen plasma. furosemide as a constant rate infusion ( . - . mg/kg/hour) also can dilate the pulmonary vasculature and decrease fluid accumulation in cases of ards. following initial stabilization of the patient, thoracic radiographs and an echocardiogram should be assessed to determine cardiac side, pulmonary vascular size, and cardiac contractility. further diagnostic testing may be required to determine other underlying causes of pulmonary edema. heart failure is managed with vasodilators, diuretics, oxygen, and sometimes positive inotropes. treatment ultimately consists of administration of supplemental oxygen, minimal stress and patient handling, and judicious use of diuretics. in cases of cardiogenic pulmonary edema, administer furosemide ( - mg/kg iv, im) every to minutes until the patient loses % of its body weight. positive inotropic and antiarrhythmic therapy may be necessary to improve cardiac contractility and control dysrhythmias. the clinician should determine whether the cause of the pulmonary edema is secondary to congestive heart failure with pulmonary vascular overload, volume overload, hypoalbuminemia, or increased permeability (ards). pulmonary edema secondary to ards typically is refractory to supplemental oxygen and diuretic therapy. in many cases, mechanical ventilation should be considered. a diagnosis of pulmonary thromboembolism (pte) is difficult to make and is based on clinical signs of respiratory distress consistent with pte, lack of other causes of hypoxemia, a high index of suspicion in susceptible animals, the presence of a condition associated with pte, and radiographic findings. virchow's triad consists of vascular endothelial injury, sluggish blood flow with increased vascular stasis, and a hypercoagulable state as predisposing factors for thromboembolic disease. clinical conditions that predispose an animal to pte include hyperadrenocorticism, disseminated intravascular coagulation (dic), catheterization of blood vessels, bacterial endocarditis, protein-losing nephropathy or enteropathy, hyperviscosity syndromes, heat-induced illness, pancreatitis, diabetes mellitus, inflammatory bowel disease, and immune-mediated hemolytic anemia. definitive diagnosis requires angiography or a lung perfusion scan. clinical signs associated with pte include an acute onset of tachypnea, tachycardia, orthopnea, and cyanosis. if the embolism is large, the patient may respond poorly to supplemental oxygen administration. pulmonary hypertension can cause a split second heart sound on cardiac auscultation. in some cases, a normal thoracic radiograph is present in the face of severe respiratory distress. this is a classic finding in cases of pte. potential radiographic abnormalities include dilated, tortuous, or blunted pulmonary arteries; wedge-shaped opacities in the lungs distal to an obstructed artery; and interstitial to alveolar infiltrates. the right heart may be enlarged. echocardiography can show right heart enlargement, tricuspid regurgitation, pulmonary hypertension, and evidence of underlying cardiac disease, possibly with clots in the atria. measurement of antithrombin (at) and d-dimer levels can be useful in the identification of hypercoagulable states, including dic. treatment of any patient with at deficiency or dic includes replenishment of at and clotting factors in the form of fresh frozen plasma. treatment of pte includes therapy for cardiovascular shock, oxygen supplementation, and thrombolytic therapy (see section on thromboembolic therapy). for short-term treatment, administer heparin (heparin sodium, - units/kg sq once, followed by units/kg q h of unfractionated heparin; or fractionated heparin). thrombolytic therapy may include tissue plasminogen activator, streptokinase, or urokinase. long-term therapy with low molecular weight heparin or warfarin may be required to prevent further thromboembolic events. ideally, management should include treatment and elimination of the underlying disease. smoke inhalation commonly occurs when an animal is trapped in a burning building. the most severe respiratory complications of smoke inhalation are seen in animals that are close enough to the flames to also sustain burn injuries (see section on burn injury). at the scene, many animals are unconscious from the effects of hypoxia, hypercapnia, carbon monoxide intoxication, and hydrogen cyanide gases that accumulate in a fire. carbon monoxide produces hypoxia by avidly binding to and displacing oxygen binding to hemoglobin, resulting in severe impairment of oxygen-carrying capacity. the percentage of carboxyhemoglobin in peripheral blood depends on the amount or carbon monoxide in inhaled gases and the length of time of exposure. clinical signs of carbon monoxide intoxication include cyanosis, nausea, vomiting, collapse, respiratory failure, loss of consciousness, and death. smoke inhalation of superheated particles also causes damage to the upper airways and respiratory tree. the larynx can become severely edematous and obstruct inspiration. emergency endotracheal intubation, tracheal oxygen, or tracheostomy tube may be required in the initial resuscitation of the patient, depending on the extent of airway edema. inhalation of noxious gases and particles can cause damage to the terminal respiratory bronchioles. specific noxious gases that can cause alveolar damage include combustible particles from plastic, rubber, and other synthetic products. pulmonary edema, bacterial infection, and ards can result. in any case of smoke inhalation, the first and foremost treatment is to get the animal away from the source of the flames and smoke and administer supplemental oxygen at the scene. at the time of presentation, carefully examine the animal's eyes, mouth, and oropharynx suction soot and debris from the mouth and upper airways. evaluate the patient's respiratory rate, rhythm, and pulmonary sounds. arterial blood gases should be analyzed with co-oximetry to evaluate the pao and carboxyhemoglobin concentrations. evaluation of sao by pulse oximetry is not accurate in cases of smoke inhalation, as the pao may appear normal, even when large quantities of carboxyhemoglobin are present. radiographs are helpful in determining the extent of pulmonary involvement, although radiographic signs may lag behind the appearance of clinical respiratory abnormalities by to hours. bronchoscopy and bronchoalveolar lavage provide a more thorough and accurate evaluation of the respiratory tree; however, these procedures should be performed only in patients whose cardiovascular and respiratory status is stable. management of the patient with smoke inhalation includes maintaining a patent airway, administration of supplemental oxygen, correction of hypoxemia and acid-base abnormalities, preventing infection, and treating thermal burns (see section on burn injury). if severe laryngeal edema is present, a temporary tracheostomy may be necessary to allow adequate oxygenation and ventilation. glucocorticosteroids should not be empirically used in the treatment of smoke inhalation, because of the risk of decreasing pulmonary alveolar macrophage function and increasing the potential for infection. in cases of severe laryngeal edema, however, glucocorticosteroids may be necessary to decrease edema and inflammation. the use of empiric antibiotics is contraindicated unless clinical signs of deterioration and bacterial pneumonia develop. epistaxis can be caused by facial trauma, a foreign body, bacterial or fungal rhinitis, neoplasia, coagulopathies, and systemic hypertension. acute, severe bilateral hemorrhage without wounds have been classified in several ways according their degree of tissue integrity, etiologic force, degree of contamination and duration, and degree of contamination and infection (table - ) . there are also unique causes of wounds such as burns, psychogenic dermatoses, frostbite, decubital ulcers, and snake bite. the animal should be transported to the nearest veterinary facility for definitive care. the wound should be covered or packed with dry gauze or clean linen to protect the wound, and to prevent further hemorrhage and contamination. if an open fracture is present, the limb should be splinted without placing the exposed bone back into the wound. replacing the exposed bone fragment back through the skin wound can cause further damage to underlying soft tissue structures and increase the degree of contamination of deeper tissues. if a spinal fracture is suspected, the patient should be transported on a stable flat surface to prevent further spinal mobilization and neurologic injury. at the time of presentation, first refer to the abcs of trauma, taking care to evaluate and stabilize the patient's cardiovascular and respiratory status. after a complete physical examination and history, ancillary diagnostic techniques can be performed if the patient is hemodynamically stable (see section on triage, assessment, and treatment of emergencies). initially, every patient with superficial wound should receive some degree of analgesia and an injection of a first-generation cephalosporin, preferably within hours of the injury. evaluate the wound after the patient's cardiovascular and respiratory status have been stabilized. always cover an open wound before taking an animal to the hospital to prevent a nosocomial infection. evaluate limb wounds for neural, vascular, and orthopedic abnormalities. carefully examine the structures deep to the superficial wounds. when there has been a delay in assessment of the wound, obtain samples for culture and antimicrobial susceptibility testing. if the wound is older and obviously infected, a gram stain can help guide appropriate antimicrobial therapy pending results of culture and susceptibility testing. place a support bandage saturated with a water-soluble antibiotic ointment or nonirritating antimicrobial solution (e.g., . % chlorhexidine, if bone or joint tissue is not exposed) around the wound. in addition to a first-generation cephalosporin, other appropriate antibiotic choices include amoxicillin-clavulanate, trimethoprim-sulfadiazine, amoxicillin, and ampicillin. if gram-negative flora are present, administer enrofloxacin. administer the antibiotics of choice for a minimum of days unless a change of antibiotic therapy is indicated. at the time of wound cleansing or definitive wound repair, the patient should be placed under general anesthesia with endotracheal intubation, unless the procedure will be brief (i.e., less than minutes). in such cases, a short-acting anesthetic combination open lacerations or skin loss closed crushing injuries and contusions etiologic force abrasion loss of epidermis and portions of dermis, usually caused by shearing between two compressive surfaces avulsion tearing of tissue from its attachment because of forces similar to those causing abrasion but of a greater magnitude incision wound created by a sharp object; wound edges are smooth and there is minimal trauma in the surrounding tissues laceration irregular wound caused by tearing of tissue with variable damage to the superficial and underlying tissue puncture penetrating wound caused by a missile or sharp object; superficial damage may be minimal; damage to deeper structures may be considerable; contamination by fur and bacteria with subsequent infection is common class i - hours with minimal contamination class ii - hours with significant contamination class iii > hours with gross contamination (analgesia + propofol, analgesia + ketamine/diazepam) can be administered to effect. heavy sedation with infiltration of a local anesthetic may also be appropriate for very small wounds, depending on the location of the wound and temperament of the patient. protect the wound by packing it with sterile gauze sponges soaked in sterile saline, or with watersoluble lubricating gel such as k-y jelly. clip the fur surrounding the wound, moving from the inner edge of the wound outward, to help prevent wound contamination with fur or other debris. scrub the wound and surrounding skin with an antimicrobial soap and solution such as dilute chlorhexidine until the area is free of all gross debris. gross debris within the wound itself can be flushed using a -ml syringe filled with sterile saline or lactated ringer's solution and an -gauge needle. pressure-lavage systems are also available for use, if desired. grossly contaminated wounds can be rinsed first with warm tap water to eliminate gross contamination, and then prepared as just described. debride the wound, removing skin and other soft tissue that is not obviously viable. obviously viable and questionable tissue should remain, and the wound left open for frequent reassessment on a daily basis. remove any dark or white segments of skin. questionable skin edges may or not regain viability and should be left in place for hours, so the wound can fully reveal itself. excise grossly contaminated areas of fat and underlying fascia. blood vessels that are actively bleeding should be ligated to control hemorrhage, if collateral circulation is present. if nerve bundles are ligated cleanly in a clean wound, the nerve edges should be reapposed and anastomosed. if gross contamination is present, however, definitive neurologic repair should be delayed until healthy tissue is present. excise contaminated muscle until healthy bleeding tissue is present. anastamoe tendon lacerations if the wound is clean and not grossly contaminated. if gross contamination is present, the tendon can be temporarily anastomosed and a splint placed on the limb until definitive repair of healthy tissue is possible. thoroughly lavage open wounds to a joint with sterile saline or lactated ringer's solution. infusion of chlorhexidine or povidone-iodine solution into the joint can cause a decrease in cartilage repair and is contraindicated. smooth sharp edges and remove any obvious fragments. whenever possible, the joint capsule and ligaments should be partially or completely closed. after removing bullets and metal fragments, the subcutaneous tissue and skin should be left open to heal by second intention, or should be partially closed with a drain. the joint should then be immobilized. injuries and exposed bone should be carefully lavaged, taking care to remove any gross debris without pushing the debris further into the bone and wound. the bone should be covered with a moist dressing and stabilized until definitive fracture repair can be made. this type of injury typically is seen with shearing injuries of the distal extremities caused by interaction with slow-moving vehicles. perform wet-to-dry or enzymatic debridement until a healthy granulation bed is present. if large areas of contamination are present (e.g., necrotizing fasciitis), en bloc debridement may be necessary. en bloc debridement consists of complete excision of badly infected wounds without entering the wound cavity, to prevent systemic infection. this technique should be used only if there is sufficient skin and soft tissue to allow later closure and it can be performed without damaging any major nerves, tendons, or blood vessels. open wounds often are managed by second intention healing, delayed primary closure, or secondary closure. see section on wound management and bandaging for a more complete discussion on the use of various bandaging materials in the treatment of open wounds. if an animal is presented very shortly after a wound has occurred and there is minimal contamination and trauma, the wound can be closed after induction of anesthesia and careful preparation of the wound and surrounding tissues. close any dead space under the skin with absorbable suture material in an interrupted suture pattern. avoid incising major blood vessels or nerves. close the subcutaneous tissues with absorbable suture material in an interrupted or continuous suture pattern. take care that there is not too much tension on the wound, or else surgical dehiscence will occur with patient movement. close the skin with nonabsorbable suture or surgical staples ( - to - ) . if there is any doubt at the time of repair about tissue status or inability to close all dead space, place a passive drain (penrose drain) so that the proximal end of the drain is anchored in the proximal aspect of the wound with a suture(s). leave the ends long so that the suture can be accurately identified at the time of drain removal. pass the suture through the skin, through the drain, and out the other side of the skin. place the rest of the drain into the wound and then secure it at the most ventral portion of the wound or exit hole in the most dependent area of the body, to allow drainage and prevent seroma formation. close the subcutaneous tissue over the drain before skin closure. during wound closure, be sure to not incorporate the subcutaneous or skin sutures into the drain, or it will not be possible to remove the drain without reopening the wound. bandage the area to prevent contamination. the drain can be removed once drainage is minimal (usually to days). active drains can be constructed or purchased; their use is indicated in wounds that are free of material that can plug the drain. to construct a small suction drain, remove the female portion or catheter hub at the end of a butterfly catheter. fenestrate the tubing so that there are multiple side holes, taking care to avoid making the holes larger than % of the circumference of the tubing. place the tubing into the wound via a small stab incision distal to the wound. use a purse-string suture around the tubing to facilitate a tight seal and prevent the tubing from exiting the wound. following wound closure, insert the butterfly needle into a -to -ml evacuated blood collection tube to allow fluid to drain into the tube. incorporate the tube into the bandage, and replace it when it becomes full. alternatively, the butterfly portion of the system can be removed and the tube fenestrated as described previously. place the tube into the wound and suture it in place to create a tight seal. secure the catheter hub to a syringe in which the plunger has been drawn back slightly to create suction. insert a metal pin or -to -gauge needle through the plunger at the top of the barrel to hold it at the desired level. incorporate the suction apparatus into the bandage and replace it when it becomes full. delayed primary closure should be considered when there is heavy contamination, purulent exudate, residual necrotic debris, skin tension, edema and erythema, and lymphangitis. delayed primary closure usually is made to days after the initial wound infliction and open wound management has been performed. once healthy tissue is observed, the skin edges should be debrided and the wound closed as with primary closure. secondary wound closure should be considered when infection and tissue trauma necessitate open wound management for more than days. secondary wound closure is performed after the development of a healthy granulation bed. this technique also is useful when a wound has dehisced and has formed granulation tissue. if the wound edges can be manipulated into apposition and if epithelialization has not begun, the wound can be cleansed and the wound edges apposed and sutured. this is known as early secondary closure. late secondary closure should be performed whenever there is a considerable amount of granulation tissue, the edges of the wound cannot be manipulated into position, and epithelialization has already started. in such cases, the wound should be cleaned, and the skin edges debrided to remove the epithelium. the remaining wound edges are then sutured over the granulation tissue ( shock is defined as a state of inadequate circulating volume and inability to meet cellular oxygen demands. there are three types of shock: hypovolemic, cardiogenic, and septic. early recognition of the type of shock present is crucial in the successful clinical management of shock syndrome. tissue oxygen delivery is based on cardiac output and arterial oxygen concentration. knowledge of the components of normal oxygen delivery is essential to the treatment of shock in the critical patient. improper handling of animal during further tissue and neurologic damage may occur transport (e.g., improper limb or spine immobilization). inadequate assessment of animal's animal's condition may worsen or animal may general condition or wounded tissues succumb; tissue injuries may be overlooked. inadequate wound protection during further wound contamination may occur at assessment, resuscitation, or veterinary facility. stabilization procedures inadequate wound protection while further wound contamination with fur and preparing the surrounding area debris may occur. insufficient wound lavage wound infection may occur. hydrogen peroxide wound lavage lavage offers little bactericidal activity and contributes to irritation of tissues and delayed healing. lavage has short residual activity and absorption with large wound. overly aggressive initial layered debridement may result in the removal of viable debridement tissue. en bloc debridement debridement results in removal of large amounts of tissue and a large defect for closure. use of drains potential exists for bacteria to ascend along the drain, for drain removal by the animal or breakage of the drain, and for possible tissue emphysema with air being sucked under the skin with patient movement. tube-type drains drains may cause postoperative discomfort; fenestrations may become occluded to stop intraluminal drainage. deeply placed sutures in the presence drain may be incorporated into the repair and of a drain prevent drain removal. active drains high negative pressure may cause tissue injury; highly productive wounds may necessitate changing the evacuated blood tubes several times a day with constructed drains. oxygen delivery (do ) = cardiac output (q) × arterial oxygen content (cao ) where q = heart rate × stroke volume. stroke volume is affected by preload, afterload, and cardiac contractility. where hb = hemoglobin concentration, sao = oxygen saturation, and pao = arterial partial pressure of oxygen in mm hg. thus, factors that can adversely affect oxygen delivery include inadequate preload or loss of circulating volume, severe peripheral vasoconstriction and increased afterload, depressed cardiac contractility, tachycardia and decreased diastolic filling, cardiac dysrhythmias, inadequate circulating hemoglobin, and inadequate oxygen saturation of hemoglobin. during septic shock, enzymatic dysfunction and decreased cellular uptake and utilization of oxygen also contribute to anaerobic glycolysis. an inadequate circulating volume may develop secondary to maldistribution of available blood volume (traumatic, septic, and cardiogenic origin) or as a result of absolute hypovolemia (whole blood or loss of extracellular fluid). normally, the animal compensates by ( ) splenic and vascular constriction to translocated blood from venous capacitance vessels to central arterial circulation, ( ) arteriolar constriction to help maintain diastolic blood pressure and tissue perfusion, and ( ) an increase in heart rate to help maintain cardiac output. arteriolar vasoconstrictions support perfusion to the brain and heart at the expense of other visceral organs. if vasoconstriction is severe enough to interfere with delivery of adequate tissue oxygen for a sufficient period of time, the animal may die. hypovolemic shock can result from acute hemorrhage or from severe fluid loss from vomiting, diarrhea, or third spacing of fluids. early in shock, baroreceptors in the carotid body and aortic arch sense a decrease in wall stretch from a decrease in circulating fluid volume. tonic inhibition of sympathetic tone via vagal stimulation is diminished, and heart rate and contractility increase and peripheral vessels constrict to compensate for the decrease in cardiac output. the compensatory mechanisms protect and support blood supply to the brain and heart at the expense of peripheral organ perfusion. this is called early compensatory shock. early compensatory shock is characterized by tachycardia, normal to fast capillary refill time, tachypnea, and normothermia. as shock progresses, the body loses its ability to compensate for ongoing fluid losses. early decompensatory shock is characterized by tachycardia, tachypnea, delayed capillary refill time, normotension to hypotension, and a fall in body temperature. end-stage decompensatory shock is characterized by bradycardia, markedly prolonged capillary refill time, hypothermia, and hypotension. aggressive treatment is necessary for any hope of a favorable outcome. septic shock should be considered in any patient with a known infection, recent instrumentation that could potentially introduce infection (indwelling intravenous or urinary catheter, surgery or penetrating injury), disorders or medical therapy that can compromise immune function (diabetes mellitus, immunodeficiency virus, parvovirus or feline panleukopenia virus infection, stress, malnutrition, glucocorticoids, chemotherapy). the presence of bacteria, viruses or rickettsiae, protozoa, or fungal organisms in the blood constitutes septicemia. septic shock is characterized by the presence of sepsis and refractory hypotension that is unresponsive to standard aggressive fluid therapy and inotropic or pressor support. septic shock and other causes of inflammation can lead to systemic inflammatory response syndrome (sirs). in animals, the presence of two or more of the criteria in table - in the presence of suspected inflammation or sepsis constitutes sirs (table - ). clinical signs associated with sepsis may be vague and nonspecific, including weakness, lethargy, vomiting, and diarrhea. cough and pulmonary crackles may be associated with pneumonia. decreased lung sounds may be associated with pyothorax. abdominal pain and fluid may be associated with septic peritonitis. vaginal discharge may or may not be present in patients with pyometra. diagnostic tests should include a white blood cell count, serum biochemical profile, coagulation tests, thoracic and abdominal radiographs, and urinalysis. the white blood cell count in a septic patient that is appropriately responding to the infection will be elevated with a left-shifted neutrophilia and leukocytosis. a degenerative left shift, in which leukopenia with elevated band neutrophils suggests an overwhelming infection. biochemical analyses may demonstrate hypoglycemia and nonspecific hepatocellular and cholestatic enzyme elevations. in the most severe cases, metabolic (lactic) acidosis, coagulopathies, and end-organ failure, including anuria and ards, may be present. cardiogenic shock occurs as a result of cardiac output inadequate to meet cellular oxygen demands. cardiogenic shock is associated with primary cardiomyopathies, cardiac dysrhythmias, pericardial fluid, and pericardial fibrosis. abnormalities seen on physical examination often are similar to those seen in other categories of shock, but they can also include cardiac murmurs, dysrhythmias, pulmonary rales, bloody frothy pulmonary edema fluid from the nares or mouth, orthopnea, and cyanosis. it is important to distinguish the primary cause of shock before implementing treatment (table - ) , whenever possible, because treatment for a suspected ruptured hemangiosarcoma differs markedly from the treatment for end-stage dilatative cardiomyopathy. the patient's clinical signs may be similar and include a peritoneal fluid wave, but the treatment for hypovolemia can dramatically worsen the congestive heart failure secondary to dilatative cardiomyopathy. when a patient presents with some form of shock, immediate vascular access is of paramount importance. place a large-bore peripheral or central venous catheter for the infusion of crystalloid or colloid fluids, blood component therapy, and drugs. monitor the patient's cardiopulmonary status (by ecg), blood pressure, oxygen saturation (as determined by pulse oximetry or arterial blood gas analyses), hematocrit, bun, and glucose. ancillary diagnostics, including thoracic and abdominal radiography, urinalysis, serum biochemistry profile, coagulation tests, complete blood count, abdominal ultrasound, and echocardiography, should be performed as determined by the individual patient's needs and the type of shock. the following list, called the "rule of twenty," is a guideline for case management of the shock patient. consideration of each aspect of the rule of twenty on a daily basis ensures temperature < °f or > . °f < °f or > . °f heart rate > beats/minute in dogs < or > beats/minute in cats respiratory rate > breaths/minute or paco > breaths/minute or paco < mm hg < mm hg white blood cell > , cells/µl , cells/µl count or < cells/µl o r < cells/ml or > % bands or > % bands that major organ systems are not overlooked. the list also provides a means to integrate and relate changes in different organ systems functions with one another.* the treatment of hypovolemic and septic shock requires the placement of large-bore intravenous catheters in peripheral and central veins. if vascular access cannot be obtained percutaneously or by cutdown methods, intraosseous catheterization should be considered. once vascular access is achieved, rapidly administer large volumes of crystalloid or colloid fluids. as a rule of thumb, administer / of a calculated shock dose of fluids-that is, / × ( ml/kg/hour) in dogs and / × ( ml/kg/hour) in cats) of a balanced crystalloid fluid ( normosol-r, plasmalyte-m, lactated ringer's solution, or . % sterile saline). reassess the patient's perfusion parameters (heart rate, capillary refill time, blood pressure, urine output) on a continual basis to direct further fluid therapy. synthetic colloid fluids (hetastarch, dextran , or oxyglobin) can also be administered in the initial resuscitation from shock. a guideline is to administer to ml/kg of hetastarch or dextran as a bolus over to minutes and then reassess perfusion parameters. hypertonic saline ( . % nacl, ml/kg) can be used in cases of hemorrhagic shock to temporarily restore intravascular fluid volume by drawing fluid from the interstitial space. because this type of fluid resuscitation is short-lived, hypertonic saline should always be used with another crystalloid or colloid fluid, and it should not be used in patients with interstitial dehydration. if hemorrhagic shock is present, the goal should be to return a patient's blood pressure to normal (not supraphysiologic) levels (i.e., systolic pressure - mm hg, diastolic pressure > mm hg, and mean arterial pressure ≥ mm hg) to avoid iatrogenically causing clots to fall off and hemorrhage to re-start. in critically ill patients, fluid loss can be measured in the form of urine, vomit, diarrhea, body cavity effusions, and wound exudates. additionally, insensible losses (those that cannot be readily measured from sweat, panting, and cellular metabolism) constitute ml/kg/ day. measurement of fluid "ins and outs" in conjunction with the patient's central venous pressure, hematocrit, albumin, and colloid oncotic pressure can help guide fluid therapy (see also section on fluid therapy). maintenance of normotension is necessary for adequate oxygen delivery to meet cellular energy demands. blood pressure can be measured using direct arterial catheterization, or through indirect means such as doppler plesthymography or oscillometric methods. the systolic pressure should remain at or greater than - mm hg at all times. the diastolic pressure is very important, too, as it constitutes two thirds of the mean arterial pressure; it must be greater than mm hg for coronary artery perfusion. the mean arterial pressure should be greater than mm hg for adequate tissue perfusion. if fluid resuscitation and pain management are not adequate in restoring blood pressure to normal, vasoactive drugs including positive inotropes and pressors should be considered (table - ). in cases of cardiogenic shock, vasodilator drugs (table - ) can be used to decrease vascular resistance and afterload. low-dose morphine ( . mg/kg, iv, im) dilates splanchnic vessels and helps reduce pulmonary edema. furosemide ( mg/kg/hour) also can dilate pulmonary vasculature and potentially reduce edema fluid formation in cases of ards. cardiac output is a function of both heart rate and stroke volume. stroke volume or (the amount of blood that the ventricle pumps in minute) is affected by preload, afterload, and contractility. during hypovolemic shock, there is a fall in cardiac preload due to a decrease in circulating blood volume. during septic and cardiogenic shock, there is a decrease in contractility secondary to inherent defects of the myocardium or due to the negative inotropic effects of inflammatory cytokines such as tnf-alpha, myocardial depressant factor, il- , and il- released during sepsis and systemic inflammation. afterload also may be increased because of the compensatory mechanisms and neurohumoral activation of the renin-angiotensin-aldosterone axis in hypovolemic or cardiogenic shock. as heart rate increases to compensate for a decline in cardiac output, myocardial oxygen demand increases and diastolic filling time becomes shorter. because the coronary arteries are perfused during diastole, coronary perfusion can be impaired, and myocardial lactic acidosis can develop, causing a further decline in contractility. in addition to lactic acidosis, acid-base and electrolyte abnormalities, inflammatory cytokines, direct bruising of the myocardium from trauma, and areas of ischemia can further predispose the patient to ventricular or atrial dysrhythmias. cardiac dysrhythmias should be controlled whenever possible. treatment of bradycardia should be directed at treating the underlying cause. administer anticholinergic drugs such as atropine ( . mg/kg im) or glycopyrrolate ( . mg/kg im) as necessary. in cases of third-degree or complete atrioventricular (av) block, administer a pure betaagonist such as isoproterenol ( . - . µg/kg/minute iv cri, or . mg in ml of % dextrose in water iv slowly). perform passive rewarming if the patient is hypothermic. receptor activity dosage (iv) dopamine da , da , α +++ , - µg/kg/minute (blood pressure support)* β +++ - µg/kg/minute (renal afferent diuresis) dobutamine α + , β +++ - µg/kg/minute* (blood pressure support, positive inotrope) norepinephrine α +++ , β + . - . mg/kg/minute; . - . mg/kg phenylephrine α +++ , β . - . mg/kg epinephrine α +++ , β +++ . - . mg/kg, . - . mg/kg/minute +++, strong receptor activity; , no receptor activity; +, weak receptor activity. *monitor for tachyarrhythmias at higher doses. correct any underlying electrolyte abnormalities such as hyperkalemia and hypo-and hypermagnesemia. treat ventricular dysrhythmias such as multifocal premature ventricular contractions (pvcs), sustained ventricular tachycardia > beats per minute, and r on t phenomenon (the t wave of the preceding beat occurs superimposed on the qrs complex of the next beat, and there is no return to isoelectric shelf), or if runs of ventricular tachycardia cause a drop in blood pressure. intravenous lidocaine and procainamide are the first drugs of choice for ventricular dysrhythmias. supraventricular tachycardia can impair cardiac output by impairing diastolic filling time. control supraventricular dysrhythmias with calcium channel blockers, beta-adrenergic blockers, or quinidine (table - ) . (disorientation); is minute; minutes) light sensitive and must be covered in foil and not kept for longer than hours albumin can decrease as a result of loss from the gastrointestinal tract, urinary system, and wound exudates, or into body cavity effusions. albumin synthesis can decrease during various forms of shock due to a preferential increase in hepatic acute phase protein synthesis. serum albumin contributes % of the colloid oncotic pressure of blood, in addition to its important roles as a free radical scavenger at sites of inflammation and as a drug and hormone carrier. albumin levels < . g/dl have been associated with an increase in morbidity and mortality in human and veterinary patients. administer fresh frozen plasma ( ml/kg) or concentrated human albumin ( ml/kg of % solution) to maintain serum albumin ≥ . g/dl. additional oncotic support can be in the form of synthetic colloids, as indicated. colloid oncotic pressure within the intravascular and interstitial spaces contributes to fluid flux. oncotic pressure can be measured with a colloid osmometer. normal oncotic pressure is mm hg. in cases of sepsis and sirs, increased vascular permeability increases the tendency for leakage of fluids into the interstitial spaces. colloids that can be administered until the source of albumin loss resolves include the synthetic colloids hetastarch and dextran ( - ml/kg/day), synthetic hemoglobin-based oxygen carriers (oxyglobin, - ml/kg/day), concentrated human albumin ( % albumin, ml/kg), and plasma ( ml/kg). oxygenation and ventilation can be evaluated by arterial blood gas analysis or by the noninvasive means of pulse oximetry and capnometry (see sections on pulse oximetry and capnometry). oxygen delivery can be impaired in cases of hypovolemic shock because of hemorrhage and anemia, and thus a decrease in functional capacity to carry oxygen, and is not to be used for more than weeks due to idiosyncratic blindness. in cases of cardiogenic shock as a result of impaired ability to saturate hemoglobin due to pulmonary edema in the lungs, or decrease in cardiac output. in septic shock, decreases in cardiac output due to inflammatory cytokines and a decrease in cellular oxygen extraction can lead to lactic acidosis. increased cellular metabolism and decreases in respiratory function can lead to respiratory acidosis as co increases. administer supplemental oxygen as flow-by, nasal or nasopharyngeal catheter, oxygen hood, or oxygen cage. supplemental oxygen should be humidified, and delivered at - ml/kg/minute. if oxygenation and ventilation are so impaired that the pao remains < mm hg with the patient on supplemental oxygen, a paco > mm hg, or severe respiratory fatigue, develops, and mechanical ventilation should be considered. glucose is a necessary fuel source for red blood cells and neuronal tissues, and serum glucose should be maintained within normal reference ranges. glucose supplementation can be administered as . - % solutions in crystalloid fluids, or in parenteral and enteral nutrition products. arterial and venous ph can be measured by performing blood gas analyses. decrease in tissue perfusion, impaired oxygen delivery, and decreased oxygen extraction in the various forms of shock can lead to anaerobic metabolism and metabolic acidosis. in most cases, improving tissue perfusion and oxygen delivery with crystalloid and colloid fluids, supplemental oxygen, and inotropic drugs will help normalize metabolic acidosis. serial measurements of serum lactate (normal, < . mmol/l) can be used as a guide to evaluate the tissue response to fluid resuscitative efforts. serum electrolytes often become severely deranged in shock states. serum potassium, magnesium, sodium, chloride, and total and ionized calcium should be maintained within normal reference ranges. if metabolic acidosis is severe, sodium bicarbonate can be administered by calculating the formula base deficit × . × body weight in kg = meq bicarbonate to administer because iatrogenic metabolic alkalosis can occur, a conservative approach is to administer / of the calculated dose and then recheck the patient's ph and bicarbonate levels. if the base excess is unknown, sodium bicarbonate can be administered in incremental doses of meq/kg until the ph is above . . complications associated with bicarbonate therapy include iatrogenic hypocalcemia, metabolic alkalosis, paradoxical cerebrospinal fluid acidosis, hypotension, restlessness, and death. massive trauma, neoplasia, sepsis, and systemic inflammation can all lead to coagulation abnormalities, including disseminated intravascular coagulation (dic). cage-side coagulation monitors are available for daily measurement of prothrombin time (pt), activated partial thromboplastin time (aptt), and platelet counts. fibrin degradation products (fibrin split products) become elevated in dic, trauma, hepatic disease, and surgery. coagulation proteins (clotting factors) and antithrombin often are lost with other proteins in hypoproteinemia or are consumed when microclots are formed and then dissolved. antithrombin levels can be measured by commercial laboratories. antithrombin and clotting factors can be replenished in the form of fresh frozen plasma transfusions. a more sensitive and specific test for dic is the detection of d-dimers, which can be measured by commercial laboratories. treatment for dic involves treatment and resolution of the underlying disease and administration of antithrombin and clotting factors in the form of fresh frozen plasma ( ml/kg) and heparin (unfractionated, - units/kg sq tid; fractionated [lovenox], mg/kg sq bid). monitor the patient for changes in mental status, including stupor, coma, decreased ability to swallow and protect the airway, and seizures. elevation of the patient's head can help to protect the airway and decrease the risk of increased intracranial pressure. serum glucose should be maintained within normal levels to prevent hypoglycemia-induced seizures. one of the major components of oxygen delivery is the binding to hemoglobin. packed cell volume must be kept above - % for adequate cellular oxygen delivery. acid-base status can adversely affect oxygen offloading at the tissue level if metabolic or respiratory alkalosis is present. oxygen-carrying capacity and hemoglobin levels can be increased with administration of rbc component therapy or with hemoglobin-based oxygen carriers. monitoring of renal function includes daily measurement of bun, creatinine, and urine output. normal urine output in a hydrated euvolemic patient is - ml/kg/hour. fluid ins and outs should be measured in cases of suspected oliguria or anuria. in patients with oliguria or anuria, furosemide can be administered as a bolus ( - mg/kg) or by constant rate infusion (cri)( . - mg/kg/hour). mannitol should also be administered ( . - g/kg over to minutes). dopamine ( - µg/kg/minute cri) can be administered to dilate renal afferent vessels and improve urine output. the patient's white blood cell count may be elevated, normal, or decreased, depending on the type of shock. the decision to administer antibiotics should be made on a daily basis. superficial or deep staphylococcus or streptococcus infection usually can be treated with a first-generation cephalosporin (cefazolin, mg/kg iv tid). if a known source of infection is present, administer a broad-spectrum antibiotic (cefoxitin, mg/kg iv tid; ampicillin, mg/kg qid, or enrofloxacin, - mg/kg once daily) pending results of culture and susceptibility testing. if broader anaerobic coverage is required, metronidazole ( mg/kg iv tid) should be considered. gentamicin ( - mg/kg iv once daily) is a good choice for gram-negative sepsis, provided that the patient is well hydrated and has normal renal function. ideally, patients receiving any aminoglycoside antibiotic should have a daily urinalysis to check for renal tubular casts that signify renal damage. in dogs, the gut is the shock organ. impaired gastrointestinal motility and vomiting should aggressively be treated with antiemetics and promotility drugs (dolasetron, . mg/kg iv once daily, and metoclopramide, - mg/kg/day iv cri). metoclopramide is contraindicated in cases of suspected gastrointestinal obstruction. histamine-receptor blockers such as famotidine ( . mg/kg bid iv) and ranitidine ( . to mg/kg iv bid, tid) or proton-pump inhibitors (omeprazole, . - mg/kg po once daily) can be administered for esophagitis. administer sucralfate ( . - g po tid) to treat gastric ulceration. if the gastrointestinal barrier function is diminished due to poor perfusion, infection, or inflammation, administer broad-spectrum antibiotics such as ampicillin ( mg/kg iv qid) to prevent gastrointestinal bacterial translocation. the course of drug therapy should be reviewd daily and the patient should be monitored for potential drug interactions. for example, metoclopramide and dopamine, working at the same receptor, can effectively negate the effects of each other. cimetidine, a cytochrome p enzyme inhibitor, can decrease the metabolism of some drugs. drugs that are avidly protein-bound may have an increase in unbound fraction with concurrent hypoalbuminemia or when hypoalbuminemia is present. decreased renal function may impair the renal clearance of some drugs, requiring increased dosing interval or decreased dose. nutrition is of utmost importance in any critically ill patient. patients with septic shock may become hypermetabolic and require supraphysiologic nutrient caloric requirements, while others may actually become hypometabolic. enteral nutrition is preferred, whenever possible, because enterocytes undergo atrophy without luminal nutrient stimulation. a variety of enteral feeding tubes can be placed, depending on what portion of the gut is functional, to provide enteral nutrition in an inappetent patient. loss of gastrointestinal mucosal barrier function may predispose the patients to the development of bacterial translocation and may contribute to sepsis. if enteral nutrition is impossible because of protracted vomiting or gastrointestinal resection, glucose, lipid, and amino acid products are available that can be administered parenterally to meet nutrient needs until the gastrointestinal tract is functioning and the patient can be transitioned to enteral nutrition. assessment of pain in animals in shock can be challenging. pain can result in the release of catecholamines and glucocounterregulatory hormones that can impair nutrient assimilation and lead to negative nitrogen balance, impaired wound healing, and immunocompromise. in any animal determined to be in pain, analgesic drugs should be administered to control pain and discomfort at all times. opioids are cardiovascularly friendly, and their effects can easily be reversed with naloxone if adverse effects such as hypotension and hypoventilation occur. if the patient is nonambulatory, rotate the animal from side to side every to hours to prevent lung atelectasis. passive range-of-motion exercises and deep muscle massage should be performed to increase tissue perfusion, decrease dependent edema, and prevent disuse atrophy. animals should be kept completely dry on soft, padded bedding to prevent the development of decubital ulcers. all bandages, wound sites, and catheter sites should be checked daily for the presence of swelling, erythema, and pain. soiled bandages should be changed to prevent strike-through and contamination of the underlying catheter or wound. hospitalization can be a stressful experience for patient and client alike. allowing brief visits and walks outside in the fresh air can improve a patient's temperament and decrease stress. the preemptive use of analgesic drugs on a regular schedule (not prn) should be used to prevent pain before it occurs. pain decreases the patient's ability to sleep. lack of sleep can promote further stress and impaired wound healing. the use of glucocorticosteroids and antiprostaglandins in shock therapy remains a topic of wide controversy. although the use of these agents potentially may stabilize membranes, decrease the absorption of endotoxin, and decrease prostaglandin release, the routine use of glucocorticosteroids and antiprostaglandins can decrease renal perfusion and gastrointestinal blood flow, promoting gastrointestinal ulceration and impaired renal function. the administration of supraphysiologic levels of glucocorticosteroids in patients in any type of shock can increase sodium and water retention, depress cellular immune function, and impair wound healing. in clinical studies of small animal patients, the routine use of glucocorticosteroids and antiprostaglandins has not demonstrated definite improved survival. the risks of therapy do outweigh the anecdotal reported benefits, and therefore the empiric use of glucocorticosteroids and antiprostaglandins in any shock patient is urinary tract emergencies azotemia azotemia occurs when % or more of the nephrons are nonfunctional. the magnitude of the azotemia alone cannot be used to determine whether the azotemia is prerenal, renal, or postrenal in origin, or whether the disease process is acute or chronic, reversible or irreversible, progressive or nonprogressive. before beginning treatment for azotemia, the location or cause of the azotemia must be identified. take a thorough history and then perform a physical examination. obtain blood and urine samples before initiating fluid therapy, for accurate assessment of the location of the azotemia. for example, an azotemic animal with a history of vomiting and diarrhea that appears clinically dehydrated on physical examination, normally should have a concentrated urine specific gravity (> . ) reflecting the attempt to conserve fluid. if this level is found, the azotemia is much less likely to be renal in origin, and the azotemia will likely resolve after rehydration. if, however, the urine specific gravity is isosthenuric or hyposthenuric ( . - . ) in the presence of azotemia and dehydration, primary intrinsic renal insufficiency is likely present. if the azotemia resolves with fluid therapy, the patient has prerenal and primary renal disease. if the azotemia does not resolve after rehydration, the patient has prerenal and primary renal failure. dogs with hypoadrenocorticism can have both prerenal and primary renal disease secondary to the lack of mineralocorticoid (aldosterone) influence on the renal collecting duct and renal interstitial medullary gradient. medullary washout can occur, causing isosthenuric urine in the presence of dehydration from vomiting and diarrhea. the patient often has azotemia due to fluid loss (dehydration and urinary loss) and gastric or intestinal hemorrhage (elevated bun). the prerenal component will resolve with treatment with glucocorticoids and crystalloid fluids, but the renal component may take several weeks to resolve, until the medullary concentration gradient is reestablished with the treatment and influence of mineralocorticoids. drugs such as corticosteroids and diuretics can influence renal tubular uptake and excretion of fluid, and cause a prerenal azotemia and isosthenuric urine in the absence of primary renal disease. treatment of azotemia includes calculation of the patient's dehydration estimate and maintenance fluid volumes, and administering that volume over the course of hours. identify and treat underlying causes of prerenal azotemia (shock, vomiting, diarrhea). monitor urine output closely. once a patient is euvolemic, oliguria is defined as urine output < - ml/kg/hour. urine output should return to normal in patients with prerenal azotemia as rehydration occurs. if a patient remains oliguric after rehydration, consider the possibility of oliguric acute intrinsic renal failure, and administer additional fluid therapy based on the patient's urine output, body weight, central venous pressure, and response to other medical therapies. prerenal azotemia is caused by conditions that decrease renal perfusion, including hypovolemic shock, severe dehydration, hypoadrenocorticism, congestive heart failure, cardiac tamponade, cardiac dysrhythmias, and hypotension. once renal perfusion is restored, the kidneys can resume normal function. glomerular filtration rate decreases when the mean arterial blood pressure falls to less than mm hg in a patient with normal renal autoregulation. renal autoregulation can be impaired in some diseases. passive reabsorption of urea from the renal tubules can occur during states of low tubular flow (dehydration, hypotension) even if glomerular filtration is not decreased. if renal hypoperfusion is not quickly restored, the condition can progress from prerenal disease to acute intrinsic renal failure. prerenal and renal azotemia can coexist in animals with primary renal disease, as a result of vomiting and ongoing polyuria in the absence of any oral fluid intake. the treatment of prerenal azotemia consists of rehydration, antiemetic therapy, and treatment of the underlying cause of vomiting, diarrhea, or third spacing of fluids. acute intrinsic renal failure is characterized by an abrupt decline in renal function to the extent that azotemia and an inability to regulate solute and fluid balance. patients with acute intrinsic renal failure may be oliguric or polyuric, depending on the cause and state of renal failure. in small animals, the most common causes of acute intrinsic renal failure are renal ischemia and toxins. there are three phases of acute intrinsic renal failure: induction, maintenance, and recovery. during the induction phase, some insult (ischemia or toxin) to the kidneys occurs, leading to a defective concentrating mechanism, decreased renal clearance of nitrogenous waste (azotemia), and polyuria or oliguria. if treatment is initiated during the induction phase, progression to the maintenance phase potentially can be stopped. as the induction phase progresses, there is worsening of the urine-concentrating ability and azotemia. renal tubular epithelial cells and renal tubular casts can be seen on examination of the urine sediment. glucosuria may be present. the maintenance phase of acute intrinsic renal failure occurs after a critical amount of irreversible nephron injury. correction of the azotemia and removal of the cause of the problem do not result in return to normal function. in patients with oliguria, the extent of nephron damage is greater than that observed in patients with polyuria. the maintenance phase may last for several weeks to months. recovery of renal function may or may not occur, depending on the extent of injury. the most serious complications (overhydration and hyperkalemia) are observed in patients with oliguria. the recovery phase occurs with sufficient healing of damaged nephrons. azotemia may resolve, but concentrating defects may remain. if the patient was oliguric in the maintenance phase, a marked diuresis develops during the recovery phase that may be accompanied by fluid and electrolyte losses. this phase may last for weeks to months. treatment of acute intrinsic renal failure consists of determining the cause and ruling out obstruction or uroabdomen whenever possible. a careful history can sometimes determine whether there has been exposure to nephrotoxic drugs, chemicals, or food items. if ingestion or exposure to a toxic drug, chemical, or food occurred recently (within to hours), induce emesis with apomorphine ( . mg/kg iv). next, administer activated charcoal either orally or via stomach tube, to prevent further absorption of the toxin. obtain blood and urine samples for toxicologic analysis (e.g., ethylene glycol) and to determine whether azotemia or abnormalities in the urine sediment exist. (see section on ethylene glycol, grapes and raisins, and nonsteroidal antiinflammatory drugs). obtain a complete blood count, biochemical profile, and urinalysis to determine the presence of signs of chronic renal failure, including polyuria, polydipsia, and nonregenerative anemia. radiographs and abdominal ultrasound can help in determining the chronicity of renal failure. normal renal size is . - . times the length of l in dogs and . - . times the length of l in cats. monitor the patient's body weight at least twice a day to avoid overhydration. also monitor urine output; normal output is - ml/kg/hour. in cases of polyuric renal failure, massive fluid and electrolyte losses can occur. place a urinary catheter for patient cleanliness and to facilitate urine quantitation. measure fluid ins and outs (see section on fluid therapy). after the patient has been rehydrated, the amount of fluids administered should equal maintenance and insensible needs plus the volume of urine produced each day. if a urinary catheter cannot be placed or maintained, serial body weight measurements and central venous pressure should be used to monitor the patient's fluid balance and prevent overhydration. if the patient is oliguric (urine output < - ml/kg/hour), pharmacologic intervention is necessary to increase urine output. first, administer furosemide ( - mg/kg or . mg/kg/hour iv cri). repeat bolus doses of furosemide if there is no response to initial treatment. if necessary, administer low-dose dopamine ( - µg/kg/minute iv cri) to increase renal afferent dilatation and renal perfusion. dopamine and furosemide may be synergistic if administered together. if dopamine and furosemide therapy is ineffective, administer mannitol ( . - . g/kg iv) once only. if polyuria is present, management is simplified because of the decreased risk of overhydration. if oliguria cannot be reversed, monitor the central venous pessure, body weight, and respiratory rate and effort, auscultate for crackles, and examine the patient carefully for signs of chemosis and the presence of serous nasal discharge. correct hyperkalemia with sodium bicarbonate ( . - . meq/kg iv) or with insulin ( . units/kg) plus dextrose ( g/unit of insulin iv, followed by . % dextrose iv cri). treat severe metabolic acidosis (ph < . or hco − < meq/l) with sodium bicarbonate. if anuria develops or oliguria is irreversible despite this therapy, begin peritoneal dialysis. obtain a renal biopsy to establish a diagnosis and prognosis (see section on renal biopsy). administer gastroprotectant drugs and antiemetics to control nausea and vomiting. if possible, avoid the use of nephrotoxic drugs and general anesthesia. initiate nutritional support in the form of an enteral feeding tube or parenteral nutrition as early as possible. once the patient enters the recovery phase, diuresis may occur that can lead to dehydration and electrolyte imbalances (hyponatremia, hypokalemia). dehydration and electrolyte imbalances can be treated with parenteral fluid and electrolyte supplementation. postrenal azotemia is primarily caused by urethral obstruction or leakage from the urinary tract into the abdomen (uroabdomen). complete urinary tract obstruction and uroabdomen are both ultimately fatal within to days if left untreated. in dogs, the most common causes of urethral obstruction are urinary (urethral) calculi or tumors of the urinary bladder or urethra. in male cats, feline urologic syndrome (fus) is the most common cause of urethral obstruction, although there has been an increased incidence of urethral calculi observed in recent years. a ruptured urinary bladder is the most common cause of uroabdomen and is usually secondary to blunt trauma. clinical signs of urinary tract obstruction include dysuria, hematuria, inability to urinate or initiate an adequate stream of urine, and a distended painful urinary bladder. late in the course of obstructive disease, clinical signs referable to uremia and azotemia (vomiting, oral ulcers, hematemesis, dehydration, lethargy, and anorexia) occur. the initial goal of treatment of urinary tract obstruction is to relieve the obstruction. in male dogs, a lubricated catheter can be inserted past the area of obstruction with the animal under heavy sedation or general anesthesia (see section on urohydropulsion). depending on the chronicity of the obstruction, serum electrolytes should be measured;an ecg should be obtained before administering any anesthetic drugs, because of the cardiotoxic effects of hyperkalemia (see section on atrial standstill). correct fluid, electrolyte, and acid-base abnormalities. if a urinary catheter cannot be placed, perform cystocentesis only as a last resort, because of the risk of urinary bladder rupture. definitive treatment includes identification and treatment of the underlying cause (tumor versus urinary calculi). in most cases, surgical intervention is necessary. if an unresectable tumor is present, a low-profile permanent cystostomy tube can be placed, if the owner desires. administration of piroxicam (feldene, . mg/kg po q - h) with or without chemotherapy may shrink the tumor mass and delay the progression of clinical signs. a complete discussion of this disorder is beyond the scope of this text (see additional reading for other sources of information). feline lower urinary tract disease can cause urethral obstruction, particularly in male cats. clinical signs include stranguria, dribbling of small amounts of urine, lethargy, inappetence, and vomiting. often, owners call with the primary complaint of constipation, because the cat is making frequent trips to the litterbox and straining. cases with a duration of obstruction < hours are considered uncomplicated; those with a duration > hours are complicated. treatment of urethral obstruction includes stabilizing and normalizing the patient's electrolyte status, induction of sedation or general anesthesia, and relieving the obstruction. obtain blood samples for analysis of electrolyte abnormalities. treat hyperkalemia (k + > . meq/l) with sodium bicarbonate ( . - . meq/kg iv), regular insulin ( . unit/ kg iv) plus dextrose ( g//unit of insulin iv), followed by . % dextrose iv cri to prevent hypoglycemia; or calcium gluconate ( . ml/kg % iv slowly). administer non-potassiumcontaining intravenous fluids in . % saline solution. obtain an ecg to detect atrial standstill (see section on atrial standstill). in some cases, a urethral plug is visible at the tip of the penis. the urethral plug can sometimes be manually extracted or massaged from the penis, and the obstruction temporarily relieved. in such cases, it is still necessary to pass a urethral catheter to flush sediment from the urethra and urinary bladder. unless a patient is obtunded, administer an anesthetic such as ketamine, atropine, or propofol ( - mg/kg iv) with diazepam iv for patient comfort and muscle relaxation. once the patient is under anesthesia or heavily sedated, urinary catheterization should be performed. in some cases, it will be difficult to advance the catheter. lubricate a closedended tomcat catheter and pass the tip into the distal urethra. fill a -ml syringe with sterile saline and sterile lubricant and connect the syringe to the hub of the catheter. pulse the fluid into the catheter as you gently move the catheter tip back and forth against the urethral obstruction. when the catheter has been passed into the urinary bladder, obtain a urine sample for urinalysis. drain the bladder and flush with sterile saline solution until the urine efflux appears clear. remove the tomcat catheter and insert a - fr red rubber tube or argyle infant feeding catheter into the urethra for urine collection and quantitation. secure the urinary catheter to prepuce with a butterfly strip of -inch adhesive tape secured around the catheter and then sutured to either side of the prepuce. the catheter should be connected to a closed urinary collection system for cleanliness and to reduce the risk of ascending bacterial infection. an elizabethan collar should be placed at all times to prevent the patient from damaging or removing the catheter. when the urethral obstruction has been relieved and the catheter placed, continue intravenous fluid diuresis to alleviate postrenal azotemia. monitor the urine for bacteria and other sediment. in some cases, postobstructive diuresis can be severe. carefully monitor fluid ins and outs, along with body weight, to maintain adequate hydration and perfusion. remove the urinary catheter can be removed after to hours. palpate the bladder frequently to make sure that the patient is voiding normally and to detect the recurrence of obstruction. in patients with severe penile or urethral trauma or edema, administer a short-acting steroid (dexamethasone sodium phosphate, . mg/kg iv, im, sq). at the time of initial diagnosis and again at the time of discharge, the clients need to be instructed about the long-term management of feline lower urinary tract disease at home, and informed of the risks and consequences of recurrence. uroabdomen can occur from trauma or leakage from the kidneys, ureter, or urinary bladder. clinical signs of uroabdomen (azotemia, uremia, hyperkalemia) can also occur secondary to third spacing of urine and leakage into muscular tissue from a ruptured urethra. in most cases, urinary bladder trauma and rupture are secondary to blunt trauma. abdominocentesis should be performed in any animal with suspected blunt abdominal trauma, and any fluid obtained should be analyzed for creatinine or potassium and compared with the patient's serum levels. an abdominal effusion that has a low packed cell volume and a potassium or creatinine level greater than that of the patient's serum is consistent with the diagnosis of uroabdomen. uroabdomen is not a surgical emergency. however, medical management consists of placement of a temporary abdominal drainage catheter into the abdomen, to facilitate removal of urine from the peritoneal cavity. to place the catheter, position the patient in dorsal or lateral recumbency, shave the ventral abdomen, as for any exploratory laparotomy. aseptically scrub the clipped area, and instill a local anesthestic (lidocaine, - mg/kg) caudal and to the right of the umbilicus, through the skin, subcutaneous tissues, and rectus emergency care clinical differentiation of acute necrotizing from chronic nonsuppurative pancreatitis in cats: cases acute pancreatitis in dogs mesenteric volvulus in the dog: a retrospective study of cases incidence and prognostic value of low plasma ionized calcium concentration in cats with pancreatitis: cases ( - ) review of feline pancreatitis. part : clinical signs, diagnosis and treatment gastric dilatation-volvulus syndrome in dogs diagnostic approach to acute pancreatitis pathophysiology of organ failure in severe acute pancreatitis in dogs washabau rj: gastrointestinal motility disorders and gastrointestinal prokinetic therapy watson pt: exocrine pancreatic insufficiency as an end-stage of pancreatitis in dogs clinical signs, underlying cause, and outcome in cats with seizures: cases fibrocartilaginous embolism in dogs: clinical findings and factors influencing the recovery rate kirk's current veterinary therapy xiii intervertebral disc extrusion in six cats medical management of acute spinal cord disease risk factors for recurrence of clinical signs associated with thoracolumbar intervertebral disk herniation in dogs: cases intervertebral disk disease in cats long-term functional outcome of dogs with severe injuries of the thoracolumbar spinal cord: cases canine status epilepticus: a retrospective study of cases risk factors for development of status epilepticus in dogs with idiopathic epilepsy and effects of status epilepticus on outcome and survival time: cases ( - ) skills laboratory part i: performing a neurologic examination skills laboratory part ii: interpreting the results of the neurologic examination accuracy of localization of cervical intervertebral disk extrusion or protrusion using survey radiography in dogs medical and surgical management of the glaucoma patient the feline glaucomas: cases ( - ) the canine glaucomas traumatic ocular protrusion in dogs and cats: cases traumatic glaucoma in a dog ocular and orbital porcupine quills in the dog: a review and case series hyphema: pathophysiologic considerations. comp cont educ pract vet van der woerdt a: the treatment of acute glaucoma in dogs and cats administer crystalloid intravenous fluids at maintenance rates using a balanced electrolyte solution perform urinary catheterization and collection to monitor urine output monitor serum urea nitrogen and creatinine every hours treat oliguria, defined as a drop in urine output to less than ml/kg/hour ml/kg) bolus start dopamine at to µg/kg/minute if no response to crystalloid/colloid bolus occurs within minutes consider mannitol ( . to g/kg iv) administration if no response to dopamine occurs within minutes consider furosemide ( to mg/kg iv, or . to mg/kg/hour iv cri) if no response to dopamine or mannitol occurs in to minutes if no response to furosemide, peritoneal dialysis or hemodialysis is indicated immediately, particularly if anuria is present administered with caution, because of the risk of exacerbating increased capillary permeability and causing pulmonary edema. animal patients. chlorphenoxy derivatives exert their toxic effects by an unknown mechanism, and cause clinical signs of gastroenteritis and muscle rigidity severe anemia should be treated with packed rbcs or hemoglobin-based oxygen carriers handbook of small animal toxicology and poisonings macadamia nut toxicosis in dogs the recognition and treatment of the intermediate syndrome of organophosphate poisoning in a dog acute renal failure in four dogs after raisin or grape ingestion pleural effusion in cats pulmonary function, ventilator management, and outcome of dogs with thoracic trauma and pulmonary contusions: cases ( - ) acute lung injury and acute respiratory distress syndrome smoke exposure in cats: cases ( - ) smoke exposure in dogs: cases ( - ) thoracic duct ligation and pericardectomy for treatment of idiopathic chylothorax use of intraluminal nitinol stents in the treatment of tracheal collapse in a dog clinical approach to epistaxis the veterinary icu book. teton newmedia radiographic diagnosis of diaphragmatic hernia: review of cases in dogs and cats tracheal collapse: diagnosis and medical and surgical management acute respiratory distress syndrome brachycephalic syndrome in dogs outcome and postoperative complications in dogs undergoing surgical treatment of laryngeal paralysis: cases ( - ) full recovery following delayed neurologic signs after smoke inhalation in a dog aspiration pneumonitis the veterinary icu book. teton newmedia allergic airway disease canine pleural and mediastinal effusion, a retrospective study of cases suggested strategies for ventilatory management in veterinary patients with acute respiratory distress syndrome laryngeal and tracheal disorders the veterinary icu book. teton newmedia medical and surgical treatment of pyothorax in dogs: cases traumatic diaphragmatic hernia in cats: cases canine pyothorax: clinical presentation, diagnosis, and treatment canine pyothorax: pleural anatomy and pathophysiology treatment of chronic pleural effusion with pleuroperitoneal shunt in dogs: cases ( - ) effects of doxapram hydrochloride on laryngeal function of normal dogs and dogs with naturally occurring laryngeal paralysis an overview of positive pressure ventilation risk factors, prognostic indicators, and outcome of pyothorax in cats: cases ( - ) use of percutaneous arterial embolization for the treatment of intractable epistaxis in dogs systemic inflammatory response syndrome, sepsis, and multiple organ dysfunction cardiogenic shock and cardiac arrest hemostatic changes in dogs with naturally occurring sepsis multiple organ dysfunction syndrome in humans and dogs increased lactate concentrations in ill and injured dogs the role of albumin in health and disease pathophysiologic characteristics of hypovolemic shock usefulness of systemic inflammatory response syndrome criteria as an index for prognosis judgement current principles and application of d-dimer analysis in small animal practice choosing fluids in traumatic hypovolemic shock: the role of crystalloids, colloids and hypertonic saline colloid and crystalloid resuscitation thromboembolic disease: predispositions and management marks sl: systemic arterial thromboembolism retrospective study of streptokinase administration in cats with arterial thromboembolism feline arterial thromboembolism: an update arterial thromboembolism in cats: acute crises in cases ( - ) and long-term management with low-dose aspirin in cases cut multiple holes in the side of a - fr red rubber tube or thoracic drainage catheter, using care not to make the cut wider than % of the circumference of the tube. insert the catheter into the abdominal cavity in a dorsal caudal direction. make sure that all incisions within the abdomen. secure the tube by placing a pursestring suture around the tube entrance site in the abdominal musculature with absorbable suture material. close the dead space in the subcutaneous tissues with absorbable suture. close the skin around the tube with another purse-string suture secured using a finger-trap technique. connect the tube to a closed urinary collection system and bandage the catheter to the abdomen. the tube can remain in place until the patient retrospective evaluation of acute renal failure in dogs uroabdomen in dogs and cats drug-induced nephrotoxicity: recognition and prevention peritoneal dialysis in emergency and critical care acute renal failure caused by lily ingestion in six cats early diagnosis of renal disease and renal failure acute renal failure in four dogs after raisin or grape ingestion disorders of the feline lower urinary tract the use of a low-profile cystostomy tube to relieve urethral obstruction in a dog renal biopsy: methods and interpretation feline idiopathic cystitis: current understanding of pathophysiology and management today's problem when did you first notice that something was wrong with your pet? when was the last time you noticed your pet act normally? what was the first abnormal sign noticed? what other conditions have developed and what are they? how soon did other signs develop? have the signs become better or worse since you first saw them? what is the name of the product? do you have the container with you today? is it a liquid concentrate, dilute spray, or solid? how long ago do you think that your pet was exposed to the poison? where do you think it happened? do you have any over-the-counter or prescription medications that your animal may have had access to? did you give any medications to your animal? is there any possibility of recreational drug exposure?your pet's recent activity did your pet eat this morning or last night? what is he/she normally fed? is there a chance that your pet may have gotten into the garbage? have you fed table scraps or anything new recently? if so, what? has your pet been off your property in the last - hours? does your pet run loose unattended? has your pet had any antiflea/tick medication within the last week?your pet's environment is your animal kept inside or outside of the house? is your pet kept in a fenced-in yard or allowed to run loose unattended? does your pet have access to neighboring properties (even for a short time)? where has your pet been in the last hours? has your pet traveled outside of your immediate geographic location? if so, when? has your pet been to rural areas in the last week? has there been any gardening work recently? does your pet have access to a compost pile? any fertilizers or weed killer used in the last week? any construction work or renovation recently? any mouse or rat poison in your house, yard, or garage? any cleaning products used inside or outside the house within the last hours? if so, which? have you changed your radiator fluid or does a car leak antifreeze? induce and maintain a patent airway and stabilize the patient's cardiovascular and respiratory status. control cns excitation with diazepam, if necessary, and control the patient's body temperature (both hypo-and hyperthermia) . induce vomiting if the patient is alert and can protect its airway; otherwise, perform orogastric lavage with the patient under general anesthesia with a cuffed endotracheal tube in place. alcohols do not bind well with activated charcoal. treat dermal exposure by bathing the area with warm water. introduction: if ingested, sodium or potassium hydroxide can cause severe contact dermatitis or irritation of the gastrointestinal tract. esophageal burns and full-thickness coagulative necrosis can occur. if an animal ingests a caustic alkali substance, feed the animal four egg whites mixed with quart of warmed water. perform endoscopy within hours to evaluate the extent of injury and to place a feeding tube, in severe cases. do not induce emesis , and do not perform orogastric lavage, because of the risk of worsening esophageal irritation. in cases of contact exposure to the skin or eyes, rinse the exposed area with warm water baths for at least minutes. administer gastroprotectant, antiemetic, and analgesic drugs as necessary. avoid neutralization, which can cause a hyperthermic reaction and worsen injury to the skin and gastrointestinal tract. amitraz is the active ingredient in ascaricides and anti-tick and anti-mite products such as mitaban and taktic. the toxic dose is to mg/kg. amitraz exerts its toxic effects by causing α-adrenergic stimulation, and causes clinical signs similar to those observed with administration of xylazine: bradycardia, cns depression, ataxia, hypotension, hyperglycemia, hypothermia, cyanotic mucous membranes, polyuria, mydriasis, and emesis. a coma can develop. treatment of amitraz intoxication includes cardiovascular support with intravenous crystalloid fluids and induction of emesis in asymptomatic animals. if clinical signs are present, orogastric lavage may be required. many toxic compounds are impregnated in a collar form. if the patient has ingested a collar and does not vomit it, it should be removed using endoscopy or gastrotomy. administer activated charcoal to prevent or delay absorption of the toxic compound. yohimbine or atepamizole, both α-adrenergic antagonists, are the treatment(s) of choice to reverse the clinical signs of toxicity. avoid the use of atropine, because it can potentially increase the viscosity of respiratory secretions and cause gastrointestinal ileus, thus promoting increased absorption of the toxic compound. ammonium hydroxide, or cleaning ammonia, can be caustic at high concentrations (see alkalis/caustics) and cause severe injury to the respiratory system if inhaled. pulmonary edema or pneumonia can occur, resulting in respiratory distress. ingestion of ammonia can cause severe irritation to the gastrointestinal tract and cause vomiting and esophageal injury. if ammonia is ingested, administer a dilute solution of egg white.administer gastroprotectant, antiemetic, and analgesic drugs as necessary. if pneumonia or pulmonary edema occurs secondary to aspiration of ammonia into the airways and alveolar spaces, treatment is largely supportive with supplemental oxygen administration, antibiotics, fluid therapy, and mechanical ventilation as necessary. diuretics may or may not be useful in the treatment of pulmonary edema secondary to ammonia inhalation. amphetamines cause cns excitation due to neurosynaptic stimulation, resulting in hypersensitivity to noise and motion, agitation, tremors, vomiting, diarrhea, and seizures. clinical signs of amphetamine toxicity include muscle tremors, tachyarrhythmias, mydriasis, ptyalism, and hyperthermia. amphetamines are rapidly absorbed from the gastrointestinal tract. treatment includes administration of intravenous fluids to maintain hydration and renal perfusion and correction of hyperthermia. administer sedative drugs such as chlorpromazine to control agitation and tremors, and diazepam to control seizures. urinary acidification can promote excretion and prevent reabsorption from the urinary bladder. in severe cases, treat cerebral edema with a combination of mannitol followed by furosemide to control increased intracranial pressure.antifreeze: see ethylene glycol antihistamines introduction antihistamines (loratadine, diphenhydramine, doxylamine, clemastine, meclizine, dimenhydrinate, chlorpheniramine, cyclizine, terfenadine, hydroxyzine) are available as over-thecounter and prescription allergy and anti-motion sickness products. clinical signs of antihistamine toxicity include restlessness, nausea, vomiting, agitation, seizures, hyperthermia, and tachyarrhythmias. treatment of antihistamine intoxication is largely symptomatic and supportive, as there is no known antidote. if ingestion is recent (within to hours) and the patient is not actively seizing and can protect its airway, induce emesis or perform orogastric lavage, followed by administration of activated charcoal and a cathartic. monitor the patient's heart rate, rhythm, and blood pressure. treat cardiac arrhythmias, if present, with appropriate therapies (see section on cardiac dysrhythmias). administer cooling measures and intravenous fluids to treat hyperthermia. a constant rate infusion of guaifenasin can be used to control muscle tremors. introduction α-naphthylthiourea (antu) is manufactured as a white or blue-gray powder. the toxic dose in dogs is - mg/kg, and in cats is - mg/kg. younger dogs appear to be more resistant to its toxic effects. antu usually causes profound emesis and increased capillary permeability that eventually leads to pulmonary edema. treatment of antu toxicity includes respiratory support. mechanical ventilation may be required in severe cases of pulmonary edema. if an animal does not vomit, orogastric lavage should be performed. administer gastrointestinal protectant, antiemetic, and analgesic drugs. cardiovascular support in the form of intravenous crystalloids should be arsenic introduction inorganic arsenic (arsenic trioxide, sodium arsenite, sodium arsenate) is the active ingredient in many herbicides, defoliants, and insecticides, including ant killers. the toxic dose of sodium arsenate is - mg/kg; that of sodium arsenite is - mg/kg. sodium arsenite is less toxic, although cats are very susceptible. arsenic compounds interfere with cellular respiration by combining with sulfhydryl enzymes. clinical signs of toxicity include severe gastroenteritis, muscle weakness, capillary damage, hypotension, renal failure, seizures, and death. in many cases, clinical signs are acute in onset. treatment of arsenic toxicity involves procuring and maintaining a patent airway. administer intravenous crystalloid fluids to correct hypotension and hypovolemia, and normalize acidbase and electrolyte balance. if no clinical signs are present and if the compound was ingested within hours, induce emesis. if clinical signs are present, perform orogastric lavage followed by administration of activated charcoal. if dermal exposure has occurred, throughly bathe the animal to prevent further absorption. dimercaprol (bal, - mg/kg im q h) can be administered as a chelating agent. n-acetylcysteine (mucomyst) (for cats, - mg/kg po iv, then mg/kg po iv q h for days; for dogs, mg/kg po or iv, then mg/kg po iv q h for days) has been shown to decrease arsenic toxicity in rats. aspirin causes inhibition of the production of prostaglandins, a high anion gap metabolic acidosis, gastrointestinal ulceration, hypophosphatemia, and decreased platelet aggregation when ingested in high quantities (> mg/kg/ hours in dogs; > mg/kg/ hours in cats). clinical signs of aspirin toxicity include tachypnea, vomiting, anorexia, lethargy, hematemesis, and melena. treatment of aspirin toxicity is largely supportive. if the ingestion was recent (within the last hour), induce emesis or perform orogastric lavage followed by administration of activated charcoal. administer intravenous crystalloid fluids to maintain hydration and correct acid-base abnormalities. administer synthetic prostaglandin analogues (misoprostol), gastroprotectant drugs, and antiemetics. alkalinization of the urine can enhance excretion. introduction baclofen is a gaba agonist centrally acting muscle relaxant. clinical signs of toxicity include vomiting, ataxia, vocalization, disorientation, seizures, hypoventilation, coma, and apnea. clinical signs can occur at doses as low as . mg/kg. treatment of baclofen ingestion includes induction of emesis if the animal is asymptomatic. otherwise, perform orogastric lavage. emesis or orogastric lavage should be followed by administration of activated charcoal. perform intravenous crystalloid fluid diuresis to promote elimination of the toxin, maintain renal perfusion, and normalize body temperature. supplemental oxygen or mechanical ventilation may be required for hypoventilation or apnea. if seizures occur, avoid the use of diazepam, which is a gaba agonist and can potentially worsen clinical signs. control seizures with intravenous introduction β-adrenergic agonists, including terbutaline, albuterol (salbutamol), and metaproterenol, are commonly used in inhaled form for the treatment of asthma. animals commonly are exposed to the compounds after chewing on their owners' inhalers. clinical signs of β-adrenergic stimulation include tachycardia, muscle tremors, and agitation. severe hypokalemia can occur. treatment of β-adrenergic agonist intoxication includes treatment with beta-blockers (propranolol, esmolol, atenolol), intravenous fluids, and intravenous potassium supplementation. diazepam or acepromazine may be administered for sedation and muscle relaxation. introduction barbiturates such as phenobarbital are gaba agonists and induce cns depression. clinical signs of barbiturate overdose or toxicity include weakness, lethargy, hypotension, hypoventilation, stupor, coma, and death. treatment of barbiturate toxicity includes maintenance and support of the cardiovascular and respiratory systems. if clinical signs are absent and the patient can protect its airway, induce emesis followed by repeated doses of activated charcoal. perform orogastric lavage if emesis is contraindicated. administer supplemental oxygen if hypoventilation occurs. some animals may require mechanical ventilation. administer intravenous fluids to control perfusion and blood pressure. positive inotropic drugs may be required if dosedependent decrease in cardiac output and blood pressure occurs. alkalinization of the urine and peritoneal dialysis can be performed to enhance excretion and elimination. hemodialysis should be considered in severe cases, if available. automotive and dry cell batteries contain sulfuric acid that can be irritating on contact with the eyes, skin, and gastrointestinal tract. button batteries, which contain sodium or potassium hydroxide, cause contact irritation if chewed. to treat exposure, rinse the eyes and skin with copious amounts of warm tap water or sterile saline solution for a minimum of minutes. if ingestion occurred, administer gastroprotectant and antiemetic drugs. induction of emesis and orogastric lavage is absolutely contraindicated because of the risk of aspiration pneumonia and worsening esophageal irritation. no attempt should be made at performing neutralization because of the risk of causing an exothermic reaction and worsening tissue damage. administer analgesics to control discomfort. benzoyl peroxide is the active ingredient in many over-the-counter acne preparations. ingestion can result in production of hydrogen peroxide, gastroenteritis, and gastric dilatation. topical exposure can cause dermal irritation and blistering. if an animal has ingested benzoyl peroxide, do not induce emesis, because of the risk of worsening esophageal irritation. instead, perform orogastric lavage. administer gastroprotectant and antiemetic medications and closely observe the patient observed for signs of gastric dilatation.bismuth subsalicylate (pepto-bismol): see aspirin bleach, chlorine (sodium hypochlorite) introduction sodium hypochlorite is available in dilute ( %- %) or concentrated ( % industrial strength or swimming pool) solutions for a variety of purposes. sodium hypochlorite can cause severe contact irritation and tissue destruction, depending on the concentration. affected animals may have a bleached haircoat. treatment of exposure includes dilution with copious amounts of warm water or saline baths and ocular lavage. induction of emesis and orogastric lavage is absolutely contraindicated because of the risk of causing further esophageal irritation. to treat ingestion, give the animal milk or large amounts of water, in combination with gastroprotectant and antiemetic drugs, to dilute the contents in the stomach. administration of sodium bicarbonate or milk of magnesia is no longer recommended. nonchlorine bleaches (sodium peroxide or sodium perborate) have a moderate toxic potential if ingested. sodium peroxide can cause gastric distention. sodium perborate can cause severe gastric irritation, with vomiting and diarrhea; renal damage and cns excitation followed by depression can occur, depending on the amount ingested. to treat dermal or ocular exposure, rinse the skin or eyes with copious amounts of warm tap water or sterile saline for a minimum of minutes; treat ocular injuries as necessary, if corneal burns have occurred. if the bleach has been ingested, do induce emesis and perform orogastric lavage. administer milk of magnesia ( - ml/kg). boric acid is the active ingredient in many ant and roach killers. the toxic ingredient (in amounts of - g/kg) can cause clinical signs in dogs by an unknown mechanism. clinical signs include vomiting (blue-green vomitus), blue-green stools, renal damage, and cns excitation and depression. treatment of boric acid or borate ingestion includes gastric decontamination with induction of emesis or orogastric lavage, followed by administration of a cathartic to hasten elimination. activated charcoal is not useful to treat ingestion of this toxin. administer intravenous fluid therapy to maintain renal perfusion. administer gastroprotectant and antiemetic drugs, as necessary. clostridium botulinum endospores can be found in carrion, food, garbage, and the environment. ingestion of endospores and c. botulinum endotoxin rarely can cause generalized neuromuscular blockade of spinal and cranial nerves, resulting in miosis, anisocoria, lower motor neuron weakness, and paralysis. respiratory paralysis, megaesophagus, and aspiration pneumonia can occur. clinical signs usually develop within days of ingestion. differential diagnosis includes acute polyradiculoneuritis (coonhound paralysis), bromethalin intoxication, and tick paralysis. treatment of botulism is largely supportive; although an antitoxin exists, it often is of no benefit. treatment may include administration of intravenous fluids, frequent turning of the patient and passive range-of-motion exercises to prevent disuse muscle atrophy, and supplemental oxygen administration or mechanical ventilation. administer amoxicillin, ampicillin, or metronidazole. recovery may be prolonged, up to to weeks in some cases. bromethalin is the active ingredient in some brands of mouse and rat poisons. it usually is packaged as . % bromethalin in green or tan pellets, and packaged in - . g place packs. the toxic dose for dogs is . g/kg, and for cats g/kg. bromethalin causes toxicity by uncoupling of oxidative phosphorylation. an acute syndrome of vomiting, tremors, extensor rigidity, and seizures occurs within hours of ingestion of high doses. delayed clinical signs occur within to days of ingestion of a lower dose and include posterior paresis progressing to ascending paralysis, cns depression, and coma. treatment of known bromethalin ingestion includes induction of emesis or orogastric lavage, and repeated doses of activated charcoal every to hours for days, because bromethalin undergoes enterohepatic recirculation. supportive care includes intravenous fluids, anticonvulsants, muscle relaxants (methocarbamol up to mg/kg/day iv to effect), frequent turning of the patient, and passive range-of-motion exercises. supplemental oxygen and /or mechanical ventilation may be required in patients with coma and severe hypoventilation. administer mannitol ( . - g/kg) in conjunction with furosemide ( mg/kg iv) if cerebral edema is suspected. the majority of caffeine toxicities occur in dogs that ingest coffee beans. caffeine causes phosphodiesterase inhibition, and can cause cardiac tachyarrhythmias, cns stimulation (hyperexcitability and seizures), diuresis, gastric ulcers, vomiting, and diarrhea. muscle tremors and seizures can occur, resulting in severe hyperthermia. treatment of caffeine toxicity is largely symptomatic and supportive, as there is no known antidote. if clinical signs are not apparent and the patient is able to protect its airway, induce emesis. alternatively, orogastric lavage can be performed, followed by administration of activated charcoal. administer diazepam to control seizures. administer betaadrenergic blockers (e.g., esmolol, propranolol, atenolol) to control tachyarrhythmias. give intravenous fluids to maintain hydration and correct hyperthermia. the patient should be walked frequently or have a urinary catheter placed to prevent reabsorption of the toxin from the urinary bladder. carbamate compounds are found in agricultural and home insecticide products. examples of carbamates include carbofuran, aldicarb, propoxur, carbaryl, and methiocarb. the toxic dose of each compound varies. carbamate compounds function by causing acetylcholinesterase inhibition. toxic amounts cause cns excitation, muscarinic acetylcholine overload, and slud (salivation, lacrimation, urination, and defecation). miosis, vomiting, treatment of carbamate intoxication includes maintaining an airway and, if necessary, artificial ventilation. administer intravenous crystalloid fluids to control the patient's hydration, blood pressure, and temperature. cooling measures may be warranted. induce emesis if the substance was ingested within minutes and the animal is asymptomatic. give repeated doses of activated charcoal if the animal can swallow and protect its airway. control seizures with diazepam ( . mg/kg iv). bathe the patient thoroughly. atropine ( . mg/kg iv) is useful in controlling some of the muscarinic signs associated with the toxicity. pralidoxime hydrochloride ( -pam) is not useful in cases of carbamate intoxication. control muscle tremors with methocarbamol (up to mg/kg iv) or guaifenesin. in humans, ingestion or inhalation of - ml of carbon tetrachloride can be fatal. clinical signs of carbon tetrachloride toxicity include vomiting and diarrhea, then progressive respiratory and central nervous system depression. ventricular dysrhythmias and hepatorenal damage ensue. the prognosis is grave. treatment of carbon tetrachloride inhalation includes procurement and maintenance of a patent airway with supplemental oxygen, and cardiovascular support. to treat ingestion, administer activated charcoal, and give intravenous fluids to maintain hydration and support renal function. chlorinated hydrocarbons include ddt, methoxychlor, lindane, dieldrin, aldrin, chlordane, chlordecone, perthane, toxaphene, heptachlor, mirex, and endosulfan. the toxic dose of each compound varies. chlorinated hydrocarbons exert their toxic effects by an unknown mechanism, and can be absorbed through the skin and the gastrointestinal tract. clinical signs are similar to those observed in organophosphate toxicity: cns excitation, seizures, slud, (salivation, lacrimation, urination, defecation), excessive bronchial secretions, vomiting, diarrhea, muscle tremors, and respiratory paralysis. secondary toxicity from toxic metabolites can cause renal and hepatic failure. chronic exposure may cause anorexia, vomiting, weight loss, tremors, seizures, and hepatic failure. the clinical course can be prolonged in small animal patients. treatment of chlorinated hydrocarbon toxicity is largely supportive in nature, as there is no known antidote. procure and maintain the patient's airway. normalize the body temperature to prevent hyperthermia. if the substance was just ingested and the patient is not demonstrating any clinical signs, induce emesis. if the patient is symptomatic, perform orogastric lavage followed by activated charcoal administration. bathe the patient thoroughly in cases of topical exposure. administer intravenous crystalloid fluids to maintain hydration. these compounds do not appear to be amenable to fluid diuresis. introduction: chlorphenoxy derivatives are found in , -d, , , -t, mcpa, mcpp, and silvex. the ld of , -d is mg/kg; however, the toxic dose appears to be much lower in small treatment treatment of chlorphenoxy derivative toxicity is largely supportive in nature, as there is no known antidote. secure the patient's airway and administer supplemental oxygen, as necessary. control cns excitation with diazepam ( . mg/kg iv). intravenous crystalloid fluid diuresis and urinary alkalinization can promote elimination. administer gastroprotectant and antiemetic drugs, as needed. the toxic effects of chocolate are related to theobromine. various types of chocolate have different concentrations of theobromine and thus can cause clinical signs of toxicity with ingestion of varying amounts of chocolate, depending on the type. the toxic dose of theobromine is - mg/kg in dogs. milk chocolate contains mg/oz ( mg/ g) of chocolate, and has a low toxic potential. semisweet chocolate contains mg/oz ( mg/ g), and baking chocolate contains mg/oz ( mg/ g). semisweet and baking chocolate, being the most concentrated, have a moderate to severe toxic potential, even in large dogs.clinical signs of theobromine intoxication are associated with phosphodiesterase inhibition and include cns stimulation (tremors, anxiety, seizures), myocardial stimulation (tachycardia and tachyarrhythmias), diuresis, and (at very high doses) gastrointestinal ulceration. with treatment, the condition of most dogs returns to normal within to hours (t / = . hours in dogs). potential side effects include gastroenteritis and pancreatitis due to the fat content of the chocolate. treatment of chocolate toxicity includes obtaining and maintaining a protected airway (if necessary), intravenous fluid diuresis, induction of emesis or orogastric lavage followed by administration of repeated doses of activated charcoal, and placement of a urinary catheter to prevent reabsorption of the toxin from the urinary bladder. cholecalciferol rodenticide ingestion can lead to increased intestinal and renal reabsorption of calcium, causing an increase in serum calcium and dystrophic mineralization of the kidneys and liver at - mg/kg. clinical signs include lethargy, anorexia, vomiting, constipation, and renal pain within to days of ingestion. seizures, muscle twitching, and central nervous system depression may be observed at very high doses. as renal failure progresses, polyuria, polydipsia, vomiting/hematemesis, uremic oral ulcers, and melena may be observed. if the compound was ingested recently (within to hours) induce emesis or perform orogastric lavage, followed by administration of activated charcoal. check the patient's serum calcium once daily for three days following ingestion. if clinical signs of toxicity or hypercalcemia are present, decrease serum calcium with loop diuretics (furosemide, - mg/kg po or iv q h) and glucocorticosteroids (prednisone or prednisolone, - mg/kg po bid) to promote renal calcium excretion. in severe cases, salmon calcitonin ( - iu/kg sc q - h in dogs) or bisphosphonate compounds may be required. correct acid-base abnormalities with intravenous crystalloid fluid diuresis and sodium bicarbonate, if necessary. (see section on hypercalcemia.) denture cleaners contain sodium perborate as the active compound. sodium perborate can cause severe direct irritation of the mucous membranes and may also act as a cns depressant. clinical signs are similar to those seen if bleach or boric acid compound is ingested, namely vomiting, diarrhea, cns excitation then depression, and renal failure. treatment for ingestion of denture cleaner includes gastric decontamination along with induction of emesis or orogastric lavage and administration of a cathartic to hasten elimination. activated charcoal is not useful for treatment of ingestion of this toxin. administer intravenous fluid therapy to maintain renal perfusion. administer gastroprotectant and antiemetic drugs, as necessary. deodorants are usually composed of aluminum chloride and aluminum chlorohydrate. both have a moderate potential for toxicity. ingestion of deodorant compounds can cause oral irritation or necrosis, gastroenteritis, and nephrosis. treatment of deodorant ingestion includes orogastric lavage, and administration of antiemetic and gastroprotectant drugs. introduction anionic detergents include sulfonated or phosphorylated forms of benzene. dishwashing liquid is an example of an anionic detergent that can be toxic at doses of - g/kg. anionic detergents cause significant mucosal damage and edema, gastrointestinal irritation, cns depression, seizures, and possible hemolysis. ocular exposure can cause corneal ulcers and edema. treatment of anionic detergent exposure is largely symptomatic, as there is no known antidote. to treat topical toxicity, flush the patient's eyes and skin with warmed tap water or . % saline solution for a minimum of minutes, taking care to avoid hypothermia. to treat ingestion, feed the patient milk and large amounts of water to dilute the toxin. do not induce emesis, because of the risk of worsening esophageal irritation. to dilute the toxin, perform orogastric lavage, followed by administration of activated charcoal. closely monitor the patient's respiratory status, because oropharyngeal edema can be severe. if necessary, perform endotracheal intubation in cases of airway obstruction. monitor the patient for signs of intravascular hemolysis. administer intravenous crystalloid fluids to maintain hydration until the patient is able to tolerate oral fluids. cationic detergents and disinfectants include quaternary ammonia compounds, isopropyl alcohol, and isopropanol. quaternary ammonia compounds have a serious toxic potential treatment treatment of cationic detergent exposure includes careful bathing and ocular rinsing of the patient for a minimum of minutes, taking care to avoid hypotension. secure the patient's airway and monitor the patient's respiratory status. administer supplemental oxygen, if necessary. place an intravenous catheter and administer intravenous crystalloid fluids to maintain hydration. do not induce emesis, because of the risk of causing further esophageal irritation. give milk or large amounts of water orally, as tolerated by the patient, to dilute the toxin. nonionic detergents include alkyl and aryl polyether sulfates, alcohols, and sulfonates; alkyl phenol; polyethylene glycol; and phenol compounds. phenols are particularly toxic in cats and puppies. clinical signs of exposure include severe gastroenteritis and topical irritation. some compounds can be metabolized to glycolic and oxalic acid, causing renal damage similar to that observed with ethylene glycol toxicity. topical and ocular exposure should be treated with careful bathing or ocular irrigation for at least minutes. administer activated charcoal to prevent absorption of the compound. as tolerated, give dilute milk or straight tap water orally to dilute the compound. administer antiemetic and gastroprotectant drugs to control vomiting and decrease gastrointestinal irritation. administer intravenous crystalloid fluids to maintain hydration and decrease the potential for renal tubular damage. monitor the patient's acid-base and electrolyte status and correct any abnormalities with appropriate intravenous fluid therapy. introduction diclone (phigone) is a dipyridyl compound that is a cns depressant. the ld in rats is - mg/kg. dichlone reacts with thiol enzymes to cause methemoglobinemia and hepatorenal damage. to treat dichlone ingestion, induce emesis or perform orogastric lavage, followed by administration of activated charcoal and a cathartic. procure and maintain a patent airway. perform intravenous fluid diuresis to maintain renal perfusion. n-acetylcysteine may be useful in the treatment of methemoglobinemia. diethyltoluamide (deet) is the active ingredient in many insect repellants (e.g., off, cutters, hartz blockade). the mechanism of action of deet is not fully understood, but it acts as a lipophilic neurotoxin within to minutes of exposure. cats appear to be particularly sensitive to deet. a lethal dermal dose is . g/kg; if ingested, the lethal dose is much less. the toxic dose of dermal exposure in dogs is g/kg. clinical signs of toxicity include aimless gazing, hypersalivation, chewing motions, and muscle tremors that progress to seizures. recumbency and death can occur within minutes of exposure at high doses. treatment of deet toxicity is largely supportive, as there are no known antidotes. procure and maintain a patent airway and perform mechanical ventilation, if necessary. place an intravenous catheter and administer intravenous crystalloid fluids to control hydration and treat hypotension, as necessary. treat seizures with diazepam ( . mg/kg iv) or phenobarbital. because of the rapid onset of clinical signs, induction of emesis is contraindicated. perform orogastric lavage if the compound was ingested within the last hours. administer multiple repeated doses of activated charcoal. cooling measures should be implemented to control hyperthermia. if dermal exposure has occurred, bathe the patient thoroughly to avoid further exposure and absorption. diquat is a dipyridyl compound that is the active ingredient in some herbicide compounds. the ld of diquat is - mg/kg. like paraquat, diquat induces its toxic effects by causing the production of oxygen-derived free radical species. clinical signs of diquat intoxication include anorexia, vomiting, diarrhea, and acute renal failure. massive dehydration and electrolyte imbalances can occur as a result of fluid loss into the gastrointestinal tract. treatment of diquat intoxication is similar to that for paraquat ingestion. if the animal had ingested diquat within hour of presentation, induce emesis. in clinical cases, orogastric lavage may be required. both emesis and orogastric lavage should be followed by administration of kaolin or bentonite as an adsorbent, rather than activated charcoal. place an intravenous catheter and administer crystalloid fluids to restore volume status and maintain renal perfusion. monitor urine output. if oliguria or anuria occurs, treatment with mannitol, furosemide, and dopamine may be considered. ecstasy ( , -methylenedioxymethylamphetamine; mdma) is a recreational drug used by humans. ecstasy causes release of serotonin. clinical signs of intoxication are related to the serotonin syndrome (excitation, hyperthermia, tremors, and hypertension), and seizures may be observed. a urine drug screening test can be used to detect the presence of mdma. treatment of ecstasy intoxication is largely supportive, as there is no known antidote. administer intravenous fluids to maintain hydration, correct acid-base status, and treat hyperthermia. serotonin antagonist drugs (cyproheptadine) can be dissolved and administered per rectum to alleviate clinical signs. intravenous propranolol has additional antiserotonin effects. administer diazepam ( . - mg/kg iv) to control seizures. if cerebral edema is suspected, administer mannitol, followed by furosemide. ethylene glycol is most commonly found in antifreeze solutions but is also in some paints, photography developer solutions, and windshield wiper fluid. ethylene glycol in itself is only minimally toxic. however, when it is metabolized to glycolate, glyoxal, glyoxylate, and oxalate, the metabolites cause an increased anion gap metabolic acidosis and precipitation of calcium oxalate crystals in the renal tubules, renal failure, and (ultimately) death.the toxic dose in dogs is . ml/kg, and in cats is . ml/kg. the toxin is absorbed quite readily from the gastrointestinal tract and can be detected in the patient's serum within an hour of ingestion. colorimetric tests that can be performed in most veterinary hospitals can detect larger quantities of ethylene glycol in the patient's serum. in a dog with clinical treatment begin treatment of known ethylene glycol ingestion immediately. induce emesis or perform orogastric lavage and adminiser repeated doses of activated charcoal. place an intravenous catheter and perform crystalloid fluid diuresis with a known antidote. the treatment of choice for dogs is administration of -methylpyrrazole ( -mp), which directly inhibits alcohol dehydrogenase, thus preventing the conversion of ethylene glycol to its toxic metabolites. the dose for dogs is mg/kg initially, followed by mg/kg at and hours and mg/kg at hours. -mp has been used experimentally at . times the recommended dose for dogs. in cats, treatment with -mp is effective if it is administered within the first hours of ingestion.cats will demonstrate signs of sedation and hypothermia with this treatment. if -mp is not available, administer ethanol ( mg/kg iv loading dose, followed by mg/ kg/hour), or as a % solution (for dogs, . ml/kg iv q h for five treatments, then q h for five more treatments; for cats, ml/kg q h for four treatments). grain alcohol ( proof) contains approximately mg/ml of ethanol. antiemetics and gastroprotective agents should be considered. urinary alkalinization and peritoneal dialysis may enhance the elimination of ethylene glycol and its metabolites. many fertilizers are on the market, and may be composed of urea or ammonium salts, phosphates, nitrates, potash, and metal salts. fertilizers have a moderate toxic potential, depending on the type and amount ingested. clinical signs of fertilizer ingestion include vomiting, diarrhea, metabolic acidosis, and diuresis. nitrates or nitrites can cause formation of methemoglobin and chocolate-brown blood. electrolyte disturbances include hyperkalemia, hyperphosphatemia, hyperammonemia, and hyperosmolality. treatment of fertilizer ingestion includes cardiovascular support, and administration of milk or a mixture of egg whites and water, followed by induction of emesis or orogastric lavage. correct electrolyte abnormalities as they occur (see section on hyperkalemia). administer antiemetic and gastroprotectant drugs, as necessary. administer intravenous fluids to control hydration and maintain blood pressure. n-acetylcysteine may be useful if methemoglobinemia is present. fipronil is the active ingredient in frontline, a flea control product. fipronil exerts its effects by gaba antagonism and can cause cns excitation. treatment of fiprinol toxicity includes treatment of cns excitation, treatment of hyperthermia by cooling measures, and administration of activated charcoal. fire extinguisher fluid contains chlorobromomethane or methyl bromide, both of which have a serious toxic potential. dermal or ocular irritation can occur. if ingested, the compounds can be converted to methanol, and cause high anion gap metabolic acidosis, cns excitation and depression, aspiration pneumonitis, and hepatorenal damage. to treat ocular or dermal exposure to fire extinguisher fluids, flush the eyes or skin with warmed tap water or . % saline solution for a minimum of minutes. do not induce emesis or perform orogastric lavage to treat ingestion, because of the risk of causing severe aspiration pneumonitis. gastroprotectant and antiemetic drugs may be used, if indicated. administer intravenous fluids to maintain hydration and renal perfusion. supplemental oxygen or mechanical ventilation may be required in severe cases of aspiration pneumonitis. fireplace colors contain salts of heavy metals-namely, copper rubidium, cesium, lead, arsenic, antimony, barium, selenium, and zinc, all of which have moderate toxic potential, depending on the amount ingested and the size of the patient. clinical signs are largely associated with gastrointestinal irritation (vomiting, diarrhea, anorexia). zinc toxicity can cause intravascular hemolysis and hepatorenal damage. to treat ingestion of fireplace colors, administer cathartics and activated charcoal and gastroprotectant and antiemetic drugs. place an intravenous catheter for intravenous crystalloid fluid administration to maintain hydration and renal perfusion. specific chelating agents may be useful in hastening elimination of the heavy metals. fireworks contain oxidizing agents (nitrates and chlorates) and metals (mercury, copper, strontium, barium, and phosphorus). ingestion of fireworks can cause hemorrhagic gastroenteritis and methemoglobinemia. to treat firework ingestion, induce emesis or perform orogastric lavage and administer activated charcoal. administer specific chelating drugs if the amount and type of metal are known, and administer gastroprotectant and antiemetic drugs. if methemoglobinemia occurs, administer n-acetylcysteine; a blood transfusion may be necessary. introduction fuels such as barbecue lighter fluid, gasoline, kerosene, and oils (mineral, fuel, lubricating) are petroleum distillate products that have a low toxic potential if ingested but can cause severe aspiration pneumonitis if as little as ml is inhaled into the tracheobronchial tree. cns depression, mucosal damage, hepatorenal insufficiency, seizures, and corneal irritation can occur. if fuels are ingested, administer gastroprotectant and antiemetics drugs. do not induce emesis or perform orogastric lavage, because of the risk of aspiration pneumonia. to treat topical exposure, rinse the skin and eyes copiously with warm tap water or . % saline solution. administer antiemetic and gastroprotectant drugs, as necessary. administer intravenous fluids to maintain hydration and treat acid-base and electrolyte abnormalities. children's glue contains polyvinyl acetate, which has a very low toxic potential. if inhaled, the compound can cause pneumonitis. treatment of polyvinyl acetate should be performed as clinical signs of pneumonitis (increased respiratory effort, cough, lethargy, respiratory distress) occur. introduction superglue contains methyl- -cyanoacrylate, a compound that can cause severe dermal irritation on contact. do not induce emesis. do not bathe the animal, and do not apply other compounds (acetone, turpentine) in an attempt to remove the glue from the skin. the fur can be shaved, using care to avoid damaging the underlying skin. the affected area should be allowed to exfoliate naturally. glyophosate is a herbicide found in roundup and kleenup. if applied properly, the product has a very low toxic potential. clinical signs of toxicity include dermal and gastric irritation, including dermal erythema, anorexia, and vomiting. cns depression can occur. treatment includes thorough bathing in cases of dermal exposure, and induction of emesis or orogastric lavage followed by administration of activated charcoal. administer antiemetic and gastroprotectant drugs as necessary. administer intravenous crystalloid fluids to prevent dehydration secondary to vomiting. even small amounts of grapes and raisins can be toxic to dogs. the mechanism of toxicity remains unknown. clinical signs occur within hours of ingestion of raisins or grapes, and include vomiting, anorexia, lethargy, and diarrhea (often with visible raisins or grapes in the fecal matter). within hours, dogs demonstrate signs of acute renal failure (polyuria, polydipsia, vomiting) that can progress to anuria. to treat known ingestion of raisins or grapes, induce emesis or perform orogastric lavage, followed by repeated doses of activated charcoal. if clinical signs of vomiting and diarrhea are present, administer intravenous fluids and monitor urine output. aggressive intravenous fluid therapy, in conjunction with maintenance of renal perfusion, is necessary. in cases of anuric renal failure, dopamine, furosemide, and mannitol can be useful in increasing urine output. peritoneal or hemodialysis may be necessary in cases of severe oliguric or anuric renal failure. calcium channel blockers such as amlodipine and diltiazem can be used to treat systemic hypertension. supportive care includes treatment of hyperkalemia, and administration of gastroprotectant and antiemetic drugs and (if the animal is eating) phosphate binders. aromatic hydrocarbons include phenols, cresols, toluene, and naphthalene. all have a moderate toxic potential if ingested. toxicities associated with ingestion of aromatic hydrocarbons include cns depression, hepatorenal damage, muscle tremors, pneumonia, methemoglobinemia, and intravascular hemolysis. if an aromatic hydrocarbon is ingested, do not induce emesis, because of the risk of aspiration pneumonia. a dilute milk solution or water can be administered to dilute the compound. perform orogastric lavage. carefully monitor the patient's respiratory and cardiovascular status. administer supplemental oxygen if aspiration pneumonia is present. to treat topical exposure, thoroughly rinse the eyes and skin with copious amounts of warm tap water or . % saline solution. imidacloprid is the compound used in the flea product advantage. clinical signs of toxicity are related to nicotinic cholinergic stimulation, causing neuromuscular excitation followed by collapse. the compound may induce respiratory paralysis. to treat imidacloprid toxicity, procure and maintain a patent airway with supplemental oxygen administration. control cns excitation with diazepam, phenobarbital, or propofol. administer enemas to hasten gastrointestinal elimination, and administer activated charcoal. bathe the animal thoroughly to prevent further dermal absorption. closely monitor the patient's oxygenation and ventilation status. if severe hypoventilation or respiratory paralysis occurs, initiate mechanical ventilation. iron and iron salts can cause severe gastroenteritis, myocardial toxicity, and hepatic damage if high enough doses are ingested. lawn fertilizers are a common source of iron salts. treatment of ingestion of iron and iron salts includes cardiovascular support in the form of intravenous fluids and antiarrhythmic drugs, as needed. induce emesis or perform orogastric lavage for gastric decontamination. a cathartic can be administered to promote elimination from the gastrointestinal tract. antiemetic and gastroprotectant drugs should be administered to prevent nausea and vomiting. in some cases, radiographs can aid in making a diagnosis of whether the compound was actually ingested. iron toxicity can be treated with the chelating agent deferoxamine. ivermectin is a gaba agonist that is used in commercial heartworm prevention and antihelminthic compounds and can be toxic in predisposed breeds, including collies, collie loperamide is an opioid derivative that is used to treat diarrhea. clinical signs of loperamide intoxication include constipation, ataxia, nausea, and sedation. induce emesis or perform orogastric lavage, followed by administration of activated charcoal and a cathartic. naloxone may be beneficial in the temporary reversal of ataxia and sedation. ingestion of macadamia nuts can cause clinical signs of vomiting, ataxia, and ascending paralysis in dogs. the toxic principle in macadamia nuts is unknown. there is no known antidote. treatment consists of supportive care, including administration of intravenous fluids and antiemetics and placement of a urinary catheter for patient cleanliness. clinical signs resolve in most cases within hours. marijuana is a hallucinogen that can cause cns depression, ataxia, mydriasis, increased sensitivity to motion or sound, salivation, and tremors. along with these findings, a classic clinical sign is the sudden onset of dribbling urine. urine can be tested with drug test kits for tetrahydrocannabinoid (thc), the toxic compound in marijuana. there is no known antidote for marijuana toxicity; therefore, treatment is largely symptomatic. place an intravenous catheter and administer intravenous fluids to support hydration. administer atropine if severe bradycardia exists. induction of emesis can be attempted but because of the antiemetic effects of thc, is usually unsuccessful. orogastric lavage can be performed, followed by repeated doses of activated charcoal. clinical signs usually resolve within to hours. introduction "strike anywhere" matches, safety matches, and the striking surface of matchbook covers contain iron phosphorus or potassium chlorate. both compounds have a low toxic potential but can cause clinical signs of gastroenteritis and methemoglobinemia if large quantities are ingested. treatment of match and matchbook ingestion includes gastric decontamination with induction of emesis or orogastric lavage and administration of activated charcoal and a cathartic. if methemoglobinemia occurs, administer n-acetylcysteine, intravenous fluids, and supplemental oxygen. metaldehyde is the active ingredient in most brands of snail bait. the exact mechanism of toxicity is unknown but may involve inhibition of gaba channels. clinical signs associated with metaldehyde toxicity include severe muscle tremors, cns excitation, and treatment treatment of mushroom toxicity is largely supportive. if the mushroom was ingested within the last hours, induce emesis or perform orogastric lavage and then administer activated charcoal. symptomatic treatment includes intravenous fluids to promote diuresis and treat hyperthermia and skeletal muscle relaxants to control tremors and seizures (methocarbamol, diazepam). if amanita ingestion is suspected, administer hepatoprotectant agents including milk thistle. mycotoxins from penicillium spp. are found in moldy foods, cream cheese, and nuts. clinical signs of intoxication include tremors, agitation, hyperesthesia, and seizures. if tremorigenic mycotoxin toxicity is suspected, a sample of the patient's serum and gastric contents or vomitus can be submitted to the michigan state university veterinary toxicology laboratory for tremorigen assay. there is no known antidote. perform orogastric lavage, followed by administration of activated charcoal. control tremors and seizures with methocarbamol, diazepam, phenobarbital, or pentobarbital. administer intravenous fluids to control hyperthermia and maintain hydration. in cases in which cerebral edema is suspected secondary to severe refractory seizures, administer intravenous mannitol and furosemide. naphthalene is the active ingredient in mothballs and has a high toxic potential. clinical signs associated with naphthalene toxicity include vomiting, methemoglobinemia, cns stimulation, seizures, and hepatic toxicity. a complete blood count often reveals heinz bodies and anemia. do not induce emesis if naphthalene ingestion is suspected. if the ingestion was within hour of presentation, perform orogastric lavage. control seizures with diazepam or phenobarbital. administer intravenous fluids to control hyperthermia and maintain hydration. n-acetylcysteine can play a role in the treatment of methemoglobinemia. a packed rbc transfusion may be necessary if anemia is severe. observe the patient for clinical signs associated with hepatitis. nicotine toxicity occurs in animals as the result of ingestion of cigarettes, nicotine-containing gum, and some insecticides. nicotine stimulates autonomic ganglia at low doses, and blocks autonomic ganglia and the neuromuscular junction at high doses. absorption after ingestion is rapid. clinical signs include hyperexcitability and slud (salivation, lacrimation, urination, and defecation). muscle tremors, respiratory muscle fatigue or hypoventilation, tachyarrhythmias, seizures, coma, and death can occur. if the patient presents within hour of ingestion and has no clinical signs, induce emesis, followed by administration of repeated doses of activated charcoal. in patients with clinical signs of toxicity, perform orogastric lavage. administer intravenous fluids to maintain hydration and promote diuresis, and treat hyperthermia. administer atropine to treat cholinergic symptoms. urinary acidification can promote nicotine excretion. nonsteroidal antiinflammatory drugs (nsaids) include ibuprofen, ketoprofen, carprofen, diclofenac, naproxen, celecoxib, valdecoxib, rofecoxib, and deracoxib. nsaids cause inhibition of prostaglandin synthesis, leading to gastrointestinal ulceration, renal failure and hepatotoxicity. ibuprofen toxicity has been associated with seizures in dogs, cats, and ferrets. the toxic dose varies with the specific compound ingested. to treat nsaid toxicity, induce emesis or perform orogastric lavage, followed by administration of multiple repeated doses of activated charcoal. place an intravenous catheter for crystalloid fluid diuresis to maintain renal perfusion. administer the synthetic prostaglandin analogue misoprostol to help maintain gastric and renal perfusion. control seizures, if present, with intravenous diazepam. administer gastroprotectant and antiemetic drugs to control vomiting and gastrointestinal hemorrhage. continue intravenous fluid diuresis for a minimum of hours, with frequent monitoring of the patient's bun and creatinine. when the bun and creatinine levels are normal or have plateaued for hours, slowly decrease fluid diuresis % per day until maintenance levels are restored. onions, garlic, and chives contain sulfoxide compounds that can cause oxidative damage of rbcs, leading to heinz body anemia, methemoglobinemia, and intravascular hemolysis. clinical signs of toxicity include weakness, lethargy, tachypnea, tachycardia, and pale mucous membranes. vomiting and diarrhea can occur. intravascular hemolysis can cause treatment treatment of onion, chive, and garlic toxicity includes administration of intravenous fluid diuresis, and induction of emesis or orogastric lavage, followed by administration of activated charcoal and a cathartic. in cases of severe anemia, packed rbc transfusion or administration of a hemoglobin-based oxygen carrier should be considered. opiate drugs include heroin, morphine, oxymorphone, fentanyl, meperidine, and codeine. opiate compounds bind to specific opioid receptors throughout the body and produce clinical signs of miosis or mydriasis (cats), and cns excitation, followed by ataxia and cns depression, leading to stupor and coma. hypoventilation, bradycardia, hypoxia, and cyanosis can occur. to treat known overdose or ingestion of an opiate compound, induce emesis (in asymptomatic animals) or perform orogastric lavage, followed by administration of activated charcoal. administer intravenous fluids and supplemental oxygen to support the cardiovascular and respiratory systems. mechanical ventilation may be necessary until hypoventilation resolves. administer repeated doses of naloxone as a specific antidote to reverse clinical signs of narcosis and hypoventilation. if seizures are present (meperidine toxicity), administer diazepam. organophosphate compounds traditionally are used in flea control products and insecticides. common examples of organophosphates include chlorpyrifos, coumaphos, diazinon, dichlorvos, and malathion. the toxic dose varies, depending on the particular compound and individual animal sensitivity. organophosphate toxicity causes acetylcholinesterase inhibition, resulting in clinical signs of cns stimulation, including tremors and seizures. muscarinic acetylcholine overload causes the classic slud signs of salivation, lacrimation, urination, and defecation. miosis, excessive bronchial secretions, muscle tremors, and respiratory paralysis can occur. an intermediate syndrome of generalized weakness, hypoventilation, and eventual paralysis with ventral cervical ventroflexion that may require mechanical ventilation has been described. if organophosphate toxicity is suspected, whole-blood acetylcholinesterase activity can be measured and will be low. treatment of toxicity includes careful and thorough bathing in cases of dermal exposure and, if the substance was ingested, gastric decontamination with induction of emesis or orogastric lavage, followed by administration of activated charcoal, and administration of the antidote pralidoxime hydrochloride . atropine can help control the muscarinic clinical signs. supportive care in the form of cooling measures, intravenous crystalloid fluids, and supplemental oxygen or mechanical ventilation may be required, depending on the severity of clinical signs. introduction ingestion of large amounts of paintballs can cause neurologic signs, electrolyte abnormalities, and occasionally death. paintballs are gelatin capsules that contain multiple colors of if ingestion was recent and if no clinical signs of toxicity are present, induce emesis or perform orogastric lavage, followed by administration of a cathartic and activated charcoal. there is no known antidote. treatment includes supportive care in the form of intravenous fluids and administration of phenobarbital or methocarbamol to control seizures and tremors. diazepam, a gaba agonist, is contraindicated, because it can potentially worsen clinical signs. urine acidification may hasten elimination. clinical signs can last from to days. pyrethrin and pyrethroid compounds are extracted from chrysanthemums, and include allethrin, decamethrin, tralomethrin, fenpropanthrin, pallethrin, sumethrin, permethrin, tetramethrin, cyfluthrin, and resemethrin. the oral toxicity is fairly low; however, the compounds can be significantly harmful if inhaled or applied to the skin. pyrethrin and pyrethroid compounds cause depolarization and blockade of nerve membrane potentials, causing clinical signs of tremors, seizures, respiratory distress, and paralysis. contact dermatitis can occur. to distinguish between pyrethrin/pyrethroid toxicity and organophosphate toxicity, acetylcholinesterase levels should be obtained; they will be normal if pyrethrins are the cause of the animal's clinical signs. treatment of toxicity is supportive, as there is no known antidote. carefully bathe the animal in lukewarm water to prevent further oral and dermal exposure. both hyperthermia and hypothermia can worsen clinical signs. administer activated charcoal to decrease enterohepatic recirculation. atropine may control clinical signs of excessive salivation. to control muscle tremors, administer methocarbamol to effect. administer diazepam or phenobarbital to control seizures, as necessary. rotenone is used as a common garden and delousing insecticide. fish and birds are very susceptible to rotenone toxicity. rotenone inhibits mitochondrial electron transport. clinical signs of tissue irritation and hypoglycemia can occur after topical or oral exposure. if the compound is inhaled, cns depression and seizures can occur. to treat toxicity, perform orogastric lavage, followed by administration of a cathartic and activated charcoal. bathe the animal carefully to prevent further dermal exposure and further ingestion. administer diazepam or phenobarbital to control seizures. the prognosis generally is guarded. treatment of ingestion includes dilution with milk, water, or egg whites. perform orogastric lavage, followed by administration of activated charcoal. administer intravenous crystalloid fluids to maintain hydration. administer antiemetic and gastroprotectant drugs to treat gastroenteritis and vomiting.shampoos, nonmedicated: see detergents, nonionic shampoos, selenium sulfide introduction selenium sulfide shampoos (e.g., selsun blue) have a low toxic potential, and primarily cause gastroenteritis. treatment of ingestion includes dilution with water, milk, or egg whites and administration of activated charcoal. carefully and thoroughly rinse the skin and eyes to prevent further exposure. administer antiemetic and gastroprotectant drugs in cases of severe gastroenteritis. zinc-based (zinc pyridinethione) anti-dandruff shampoos have a serious toxic potential if ingested or if ocular exposure occurs. gastrointestinal irritation, retinal detachment, progressive blindness, and exudative chorioretinitis can occur. treatment of ingestion includes gastric decontamination. induce emesis or perform orogastric lavage, followed by administration of a cathartic and activated charcoal.to treat ocular exposure, thoroughly rinse the patient's eyes for a minimum of minutes. carefully monitor the animal for clinical signs of blindness. implement intravenous fluid to maintain hydration and renal perfusion in cases of severe gastroenteritis. silver polish contains the alkali substance sodium carbonate and cyanide salts, and has a serious toxic potential. ingestion results in rapid onset of vomiting and possibly cyanide toxicity. to treat ingestion, monitor and maintain the patient's respiration and cardiovascular status and administer intravenous crystalloid fluids. induce emesis, followed by administration of activated charcoal. administer sodium nitrite or sodium thiosulfate iv for cyanide toxicity. bath soap (bar soap) usually has low toxic potential and causes mild gastroenteritis with vomiting if ingested. to treat ingestion, include dilution with water, administration of intravenous fluids to maintain hydration, and administration of antiemetic and gastroprotectant drugs to treat gastroenteritis. sodium fluoroacetate is a colorless, odorless, tasteless compound that causes uncoupling of oxidative phosphorylation. the toxic dose in dogs and cats is . - . mg/kg. clinical signs of toxicity include cns excitation, seizures, and coma secondary to cerebral edema. the prognosis is guarded. to treat toxicity, procure and maintain a patent airway, monitor and stabilize the cardiovascular status, and control hyperthermia. perform orogastric lavage, followed by administration of activated charcoal. if clinical signs are not present at the time of presentation, induce emesis. administer intravenous fluids and supplemental oxygen, as necessary. strattera (atomoxetine hydrochloride) is a selective norepinephrine reuptake inhibitor used in the treatment of attention deficit hyperactivity disorder (adhd) in humans. peak serum concentrations occur in dogs within to hours of ingestion, with a peak half-life at to hours following ingestion. clinical signs of toxicity include cardiac tachyarrhythmias, hypertension, disorientation, agitation, trembling, tremors, and hyperthermia. treatment of intoxication is largely symptomatic and supportive in nature. first, induce emesis if the patient is conscious and has an intact gag reflex. orogastric lavage can also be performed. administer one dose of activated charcoal to prevent further absorption of the compound from the gastrointestinal tract. identify cardiac dysrhythmias and treat accordingly. control hypertension with sodium nitroprusside or diltiazem as a constant rate infusion. administer acepromazine or chlorpromazine to control agitation. do not use diazepam, because it can potentially worsen clinical signs. administer intravenous fluids to maintain hydration and promote diuresis. strychnine is the active ingredient in pesticides used to control rodents and other vermin. the toxic dose in dogs is . mg/kg, and in cats is mg/kg. strychnine antagonizes spinal inhibitory neurotransmitters and causes severe muscle tremors, muscle rigidity, and seizures. clinical signs are stimulated or exacerbated by noise, touch, light, and sound. mydriasis, hyperthermia, and respiratory paralysis can occur. if strychnine toxicity is suspected, gastric contents should be collected and saved for analysis. if the animal is asymptomatic at the time of presentation, induce emesis. if clinical signs are present, perform orogastric lavage. both emesis and orogastric lavage should be followed by the administration of activated charcoal. administer intravenous crystalloid fluids to support the cardiovascular system, aid in cooling measures, and improve renal diuresis. treat cns stimulation with methocarbamol, diazepam, or phenobarbital. the animal should have cotton packed in its ears to prevent noise stimulation, and should be placed in a quiet, dark room. treatment of ingestion includes dilution with milk of magnesia or water, administration of antiemetic and gastroprotectant drugs, and administration of intravenous crystalloid fluids to maintain hydration. do not induce emesis, because of the risk of causing further esophageal irritation.sunscreen: see zinc and zinc oxide suntan lotion: see shampoos, zinc-based, and alcohols tar: see fuels tea tree oil (melaleuca oil) introduction tea tree (melaleuca) oil is an herbal-origin flea-control product. the toxic principles in tea tree oil are monoterpenes, which produce clinical signs of neuromuscular weakness, and ataxia. treatment of tea tree oil toxicity includes administration of cathartics and activated charcoal to prevent further absorption. carefully bathe the animal to prevent further dermal exposure. tetanus spores from clostridium tetani organisms are ubiquitous in the soil and feces, particularly in barnyards. cases have been reported in dogs after tooth eruption and after abdominal surgeries performed with cold sterilization packs. anaerobic wound infections can contain tetanus spores. the neurotoxin from c. tetani inhibits spinal inhibitory neurons, causing motor neuron excitation. extensor muscle rigidity ("sawhorse stance"), erect ears, and risus sardonicus (a sardonic grin) are characteristic features of tetanus. administer tetanus antitoxin if toxin has not already been bound in the cns. to eliminate the source of the toxin (e.g., abscess), open and debride all wounds. intravenous administration of ampicillin or penicillin g is the treatment of choice for tetanus. supportive care in the form of skeletal muscle relaxants, intravenous fluids and parenteral nutrition, and nursing care to prevent decubitus ulcer formation is required. in extreme cases, mechanical ventilation may be necessary. triazene compounds include atrazine, prometone, and monuron (telvar). the toxic mechanism of triazene compounds is unknown. clinical signs of toxicity include salivation, ataxia, hyporeflexia, contact dermatitis, hepatorenal damage, muscle spasms, respiratory difficulty, and death. treatment of triazene exposure includes cardiovascular and renal support in the form of intravenous crystalloid fluids, inotropic drugs, and antiarrhythmic agents, as necessary. if the exposure is recent, induce emesis. perform orogastric lavage in animals that cannot protect the airway. emesis and orogastric lavage should be followed by the administration of activated charcoal and a cathartic. carefully bathe the patient to prevent further dermal absorption. a variety of tricyclic antidepressants are available for use in both humans and animals, including amitriptyline, amoxapine, desipramine, doxepine, fluoxetine (prozac), fluvoxamine (luvox), imipramine, nortriptyline, paroxetine (paxil), protriptyline, sertraline (zoloft), and trimipramine. selective serotonin reuptake inhibitors (ssris) are rapidly absorbed from the digestive tract, with peak serum concentrations occurring to hours after ingestion. the elimination half-life for each drug differs in dogs, but typically last to hours. ssris inhibit the reuptake of serotonin, causing serotonin to accumulate in the brain. this can cause "serotonin syndrome," characterized by trembling, seizures, hyperthermia, ptyalism or hypersalivation, cramping or abdominal pain, vomiting, and diarrhea. other clinical signs of ssri intoxication include depression, tremors, bradycardia, tachyarrhythmias, and anorexia. any animal that has ingested an ssri should be promptly treated and carefully observed for at least hours for side effects. the treatment of suspected ssri intoxication involves gastric decontamination if the patient is not depressed and has an intact gag reflex. perform orogastric lavage and administer activated charcoal to prevent further toxin absorption and hasten elimination from the gastrointestinal tract. treat other clinical signs symptomatically. administer intravenous diazepam to control seizures. treat tachyarrhythmias according to type. administer methocarbamol to control muscle tremors. cyproheptadine ( mg/kg), a serotonin antagonist, can be dissolved in water and administered per rectum. vitamin k antagonist rodenticides, which are commonly found in pelleted or block form, inhibit the activation of the vitamin k-dependent coagulation factors ii, vii, ix, and x. clinical signs of hemorrhage occur within to days of exposure. hemorrhage can occur anywhere in the body, and can be manifested as petechiation of the skin or mucous membranes, hemorrhagic sclera, epistaxis, pulmonary parenchymal or pleural hemorrhage, gastrointestinal hemorrhage, pericardial hemorrhage, hematuria, retroperitoneal hemorrhage, hemarthrosis, and central nervous system hemorrhage. clinical signs include respiratory distress, cough, bleeding from the gums or into the eyes, ataxia, paresis, paralysis, seizures, hematuria, joint swelling, lameness, lethargy, weakness, inappetence, and collapse.diagnosis is made based on clinical signs and a prolonged activated clotting time, or prothrombin time. the pivka (proteins induced by vitamin k antagonism) test may be helpful but usually cannot be performed in-house. slight thrombocytopenia may be present secondary to hemorrhage; however, blood levels usually do not reach the critical level of < , platelets/µl to cause clinical signs of hemorrhage. in some cases, severe stressinduced hyperglycemia and glucosuria may be present but resolves within hours. if the rodenticide was ingested within the last hours, induce emesis. alternatively, orogastric lavage can be performed in an uncooperative patient. both emesis and orogastric lavage should be followed by administration of activated charcoal. the stomach contents can be submitted for analysis. following successful treatment, administer oral vitamin k for days after the exposure; or a check prothrombin time days after gastric decontamination. if the prothrombin time is prolonged, administer fresh frozen plasma and vitamin k.if the prothrombin time is normal, gastric decontamination was successful, and no further treatment is necessary.if an animal presents with clinical signs of intoxication, administer activated clotting factors in the form of fresh frozen plasma ( ml/kg), and vitamin k ( mg/kg sq in multiple sites with a -gauge needle). packed rbcs or fresh whole blood may be required if the patient is also anemic. supportive care in the form of supplemental oxygen may be necessary in cases of pulmonary or pleural hemorrhage. following initial therapy and discharge, the patient should receive vitamin k ( . mg/kg po q - h for days), and prothrombin time should be checked days after the last vitamin k capsule is administered. in some cases, depending on the type of anticoagulant ingested, an additional weeks of vitamin k therapy may be required. xylitol is a sugar alcohol that, when ingested by humans, does not cause a significant increase in blood glucose, and therefore does not stimulate insulin release from the human pancreas. in dogs, however, xylitol causes a massive rapid and dose-dependent release of insulin from pancreatic beta-cells. following insulin release, clinically significant hypoglycemia can develop, followed by signs of vomiting, weakness, ataxia, mental depression, hypokalemia, hypoglycemic seizures, and coma. clinical signs associated with xylitol ingestion can be seen within minutes of ingestion and can last for more than hours, even with aggressive treatment. known xylitol ingestion should be treated as for other toxin ingestion. if no neurologic abnormalities exist at the time the patient is seen, induce emesis, followed by administration of activated charcoal. it remains unknown at this time whether activated charcoal actually delays or prevents the absorption of xylitol from the canine gastrointestinal tract. if clinical signs have already developed, perform orogastric lavage and gastric decontamination. blood glucose concentrations should be analyzed and maintained with supplemental dextrose as a constant rate infusion ( . %- %) until normoglycemia can be maintained with multiple frequent small meals. hypokalemia may develop because it is driven intracellularly by the actions of insulin. treat hypokalemia with supplemental potassium chloride by infusion, not to exceed . meq/kg/hour. pennies minted in the u.s. after contain large amounts of zinc rather than copper. other sources of zinc include zinc oxide ointment and hardware such as that found in metal bird cages. zinc toxicity causes intravascular hemolysis, anemia, gastroenteritis, and renal failure. if zinc toxicity is suspected, take an abdominal radiograph to document the presence of the metal in the stomach or intestines. (if zinc-containing ointment was ingested, this will not be visible on radiographs.) induce emesis or perform orogastric lavage, depending on the size of the object ingested. often, small objects such as pennies can be retrieved using endoscopy or surgical gastrotomy/enterotomy. always take an additional radiograph after the removal procedure to ensure that all objects have been successfully removed. administer intravenous fluids to maintain renal perfusion and promote fluid diuresis. administer gastroprotectant and antiemetic drugs. chelation therapy with succimer, calcium edta, dimercaprol, or penicillamine may be necessary. do not administer pulmonary contusions are a common sequela of blunt traumatic injury. a contusion basically is a bruise characterized by edema, hemorrhage, and vascular injury. contusions may be present at the time of presentation or can develop over the first hours after injury. a diagnosis of pulmonary contusion can be made based on auscultation of pulmonary crackles, presence of respiratory distress, and the presence of patchy interstitial to alveolar infiltrates on thoracic radiographs. radiographic signs can lag behind the development of clinical signs of respiratory distress and hypoxemia by hours. in most cases, cage rest is sufficient to temporarily diminish blood loss. sedation (acepromazine, . - . mg/kg iv, im, sq) may be helpful in alleviating anxiety and decreasing blood pressure. the hypotensive effects of acepromazine are potentially harmful if severe blood loss has occurred. if evidence of hypovolemia is present (see section on hypovolemic shock), intravenous fluid resuscitation should be administered. rapid assessment of clotting ability, with a platelet count estimate and clotting profile (act or aptt and pt), should be performed. if epistaxis secondary to vitamin k antagonist rodenticide intoxication is suspected, administer vitamin k and fresh frozen plasma or fresh whole blood.persistent hemorrhage from a nasal disorder can be treated with dilute epinephrine ( : , ) into the nasal cavity with the nose pointed toward the ceiling to promote vasoconstriction. if this fails, the animal can be anesthetized, and the nasal cavity packed with gauze, and the caudal oropharynx and external nares covered with umbilical tape to control hemorrhage. a rhinoscopy should be performed to determine the cause of ongoing hemorrhage. continued excessive hemorrhage can be controlled with ligation of the carotid artery on the side of the hemorrhage, or with percutaneous arterial embolization. systemic thromboembolism is most commonly recognized in cats with cardiomyopathies (hypertrophic, restrictive, unclassified, and dilatative) but can also occur in dogs with hyperadrenocorticism, disseminated intravascular coagulation (dic), systemic inflammatory response syndrome (sirs), protein-losing enteropathy and nephropathy, and tumors affecting the aorta and vena cava. thrombosis occurs through a complex series of mechanisms when the components of virchow's triad (hypercoaguable state, sluggish blood flow, and vascular endothelial injury or damage) are present. in cats, blood flow through a severely stretched left atrium is a predisposing factor to the development of clots and thromboembolism.the most common site of embolism is the aortic bifurcation, or "saddle thrombus." other, less common locations of thromboembolism include the forelimbs, kidneys, gastrointestinal tract, and cerebrum. diagnosis usually is made based on clinical signs of cool extremities, the presence of a cardiac murmur or gallop rhythm, auscultation of pulmonary crackles resulting from pulmonary edema, acute pain or paralysis of one or more peripheral extremities, respiratory distress, and pain and lack of a palpable pulse in affected limbs. the affected nailbeds and paw pads are cyanotic, and nails do not bleed when cut with a nail clipper.client education is one of the most important aspects of emergency management of the patient with thromboembolic disease. concurrent congestive heart failure (chf) occurs in % to % of cats with arterial thromboembolism. more than % of cats are euthanized during the initial thromboembolic event because of the poor long-term prognosis and the high risk of recurrence within days to months after the initial event, even with aggressive therapy. although the long-term prognosis varies from months to years after initial diagnosis and treatment, in the majority of cats thromboembolic disease recurs within months. rectal temperature hypothermia and bradycardia on presentation are negative prognostic indicators.immediate treatment of a patient with chf and thromboembolic disease involves management of the chf with furosemide, oxygen, and vasodilators (nitroglycerine paste, morphine, nitroprusside). additional management includes analgesia (butorphanol, . - . mg/kg iv, im) and prevention of further clot formation. aspirin ( mg/kg po q h) is beneficial bcause of its antiplatelet effects. heparin works in conjunction with antithrombin to prevent further clot formation ( - units/kg iv, followed by - units/kg sq q h in cats, and - units/kg sq q h in dogs). acepromazine can cause peripheral vasodilation and decreased afterload but also can promote hypotension in a patient with concurrent chf. acepromazine ( . - . mg/kg sq) should be used with extreme caution, if at all.thrombolytic therapy can also be attempted, but in most cases is not without risk, and may be cost-prohibitive for many clients. streptokinase ( , units iv over minutes and then , units/hour iv cri for hours) was administered with some success in cats; however, many died of hyperkalemia or other complications during the infusion. tissue plasminogen activator ( . - mg /kg/hour iv cri, up to mg/kg total dose, to effect) has been used with some success but is cost-prohibitive for most clients. side effects of thrombolytic therapy include hyperkalemia with reperfusion and hemorrhage.in cats, the primary cause of arterial thromboembolism is cardiomyopathy. once an animal is determined to be stable enough for diagnostic procedures, lateral and dv thoracic radiographs and an echocardiogram should be performed. ultrasound of the distal aorta and renal arteries should also be performed to determine the location of the clot and help establish the prognosis.other diagnostic procedures to evaluate the presence and cause of thromboembolism include a complete blood count, serum biochemistry profile, urinalysis (to rule out proteinlosing nephropathy), urine protein:creatinine ratio, antithrombin levels, acth stimulation test (to rule out hyperadrenocorticism), heartworm antigen test (in dogs), thyroid profile (to rule out hyperthyroidism in cats, and hypothyroidism in dogs), thoracic radiographs, arterial blood gas analyses, coagulation tests, and coombs' test. selective and nonselective angiography can also be performed to determine the exact location of the thrombus.long-term management of thromboembolism involves management of the underlying disease process and preventing further clot formation. begin therapy with heparin until the aptt becomes prolonged . times; then administer warfarin ( . - . mg/kg/day). monitoring therapy based on prothrombin time and the international normalized ratio (inr, . - . ) is recommended. low-dose aspirin ( - mg/kg q h) also has been recommended. physical therapy with warm water bathing, deep muscle massage, and passive range-of-motion exercises should be performed until the patient regains motor function. future therapy may involve the use of platelet receptor antagonists to prevent platelet activation and adhesion. key: cord- -pb k da authors: dulek, daniel e; fuhlbrigge, robert c; tribble, alison c; connelly, james a; loi, michele m; el chebib, hassan; chandrakasan, shanmuganathan; otto, william r; diorio, caroline; keim, garrett; walkovich, kelly; jaggi, preeti; girotto, jennifer e; yarbrough, april; behrens, edward m; cron, randy q; bassiri, hamid title: multidisciplinary guidance regarding the use of immunomodulatory therapies for acute covid- in pediatric patients date: - - journal: j pediatric infect dis soc doi: . /jpids/piaa sha: doc_id: cord_uid: pb k da background: immune-mediated lung injury and systemic hyperinflammation are characteristic of severe and critical coronavirus disease (covid- ) in adults. although the majority of sars-cov- infections in pediatric populations result in minimal or mild covid- in the acute phase of infection, a small subset of children develop severe and even critical disease in this phase with concomitant inflammation that may benefit from immunomodulation. therefore, guidance is needed regarding immunomodulatory therapies in the setting of acute pediatric covid- . this document does not provide guidance regarding the recently emergent multisystem inflammatory syndrome in children (mis-c). methods: a multidisciplinary panel of pediatric subspecialty physicians and pharmacists with expertise in infectious diseases, rheumatology, hematology/oncology, and critical care medicine was convened. guidance statements were developed based on best available evidence and expert opinion. results: the panel devised a framework for considering the use of immunomodulatory therapy based on an assessment of clinical disease severity and degree of multi-organ involvement combined with evidence of hyperinflammation. additionally, the known rationale for consideration of each immunomodulatory approach and the associated risks and benefits was summarized. conclusions: immunomodulatory therapy is not recommended for the majority of pediatric patients, who typically develop mild or moderate covid- . for children with severe or critical illness, the use of immunomodulatory agents may be beneficial. the risks and benefits of such therapies are variable and should be evaluated on a case-by-case basis with input from appropriate specialty services. when available, the panel strongly favors immunomodulatory agent use within the context of clinical trials. the framework presented herein offers an approach to decision-making regarding immunomodulatory therapy for severe or critical pediatric covid- and is informed by currently available data, while awaiting results of placebo-controlled randomized clinical trials. severe acute respiratory syndrome coronavirus (sars-cov- ) is a recently emergent human pathogen that causes a variety of disease manifestations termed coronavirus disease . the spectrum of covid- ranges from asymptomatic infections to severe and critical illness with multi-organ involvement that can prove fatal [ ] . in adults, the most common disease presentation involves respiratory disease that either resolves or evolves into progressive pulmonary involvement and acute respiratory distress syndrome (ards); in adult patients with progressive disease, extrapulmonary manifestations and evidence of multi-organ involvement is common. while severe and critical covid- is substantially more prevalent in adults, a small proportion of children also develop progressive respiratory disease and concomitant multi-organ dysfunction with high morbidity, but fatalities are rarely reported [ ] [ ] [ ] [ ] . comorbid conditions for this presentation of severe and critical covid- in adults include obesity, diabetes, and underlying cardiac disease [ , ] however, such risk factors are currently not well defined in children [ , , ] . initial descriptions of covid- presentations and outcomes indicate a substantial inflammatory component to severe disease [ ] [ ] [ ] . inflammatory phenotypes include significant pulmonary inflammation accompanied by prolonged fevers [ ] and/or a biphasic illness course characterized by initial improvement followed by rapid occurrence of respiratory failure and pulmonary inflammation [ , ] . in addition, covid- -associated cardiac injury is an independent risk factor for mortality, suggesting that inflammation beyond lung parenchyma contributes to poor outcomes [ , ] . further, severe covid- is associated with more significant lymphopenia, systemically elevated pro-inflammatory cytokine levels, and impaired cd + t cell ifn-γ expression compared to moderate covid- [ ] . these early reports of hyperinflammation are reminiscent of the cytokine storm features described in the setting of prior emergent respiratory virus infections including sars ( ), middle east respiratory syndrome (mers), h n avian influenza, and h n pandemic influenza [ , ] . in each of these viral infections, significant pulmonary and systemic inflammation was identified and, in some cases, linked to poor outcomes and mortality (sars [ ] [ ] [ ] [ ] ; avian influenza [ , ] ; h n [ ] [ ] [ ] ; mers [ ] [ ] [ ] ). since the initial sars outbreak in , several developments have changed the paradigm for treatment of hyperinflammation. first, detailed mechanistic knowledge of the genetics and immunopathology of macrophage activation syndrome (mas) and hemophagocytic lymphohistiocytosis (hlh) have led to targeted treatments for cytokine storm syndrome (css) [ ] . in addition, new immunomodulators targeting specific cytokines, cytokine receptors, and immune pathways have been developed and studied in a wide variety of other a c c e p t e d m a n u s c r i p t inflammatory and autoimmune diseases [ , ] . finally, the advent of chimeric antigen receptor (car) t cell therapy for leukemia/lymphoma and resultant cytokine release syndrome (crs) has provided a specific example of cytokine-targeted therapy leading to rapid clinical improvement in the setting of marked and hyperacute inflammation [ ] . symptoms of severe covid- in children as currently reported fall into two categories. in a small subset of pediatric patients, severe lung disease occurs and appears to mimic severe adult covid- with respiratory failure, ards, and associated multi-organ failure [ ] . in other pediatric patients, an emerging inflammatory disease has recently been described [ ] [ ] [ ] this latter presentation has been variably called pims-ts [ ] , pmis [ ] , and mis-c [ ] and manifests as acute onset of fever with multisystem involvement, frequently including hypotension and cardiac dysfunction in the absence of respiratory symptoms. some reported cases mimic severe kawasaki disease or toxic shock syndrome phenotypes. mis-c appears to be associated with prior exposure to sars-cov- . although immune modulation with corticosteroids and intravenous immunoglobulin (ivig) is used in severe cases of mis-c [ ] , given the very limited information about the mechanisms of this disease process, in this document we do not provide specific guidance for treatment of this syndrome. anecdotal reports of immunomodulator use in the setting of covid- have been widespread [ ] [ ] [ ] [ ] [ ] . while numerous trials of immunomodulatory therapies for covid- in adults are being launched, few clinical trials for immunomodulatory therapy in pediatric patients with covid- are currently enrolling or planned. therefore, we undertook a comprehensive review of the current state of literature regarding immunomodulatory therapy in covid- . the goal for this document is to provide pediatric practitioners a framework for interpreting currently available data and a rationale for considering immunomodulatory therapy in the care of pediatric acute covid - patients. in the sections below, cytokine storm, css, and hyperinflammation are used interchangeably to refer to the pulmonary and systemic inflammation that accompanies severe and critical covid- . this review does not represent a final or definitive guideline for diagnosis or treatment, but a review of current knowledge, and we emphasize the importance of enrollment in clinical trials for covid- immunomodulatory therapy when available. a c c e p t e d m a n u s c r i p t a multidisciplinary panel of pediatric subspecialty physicians and pharmacists with expertise in infectious diseases, rheumatology, hematology/oncology, and critical care medicine was convened. we relied on the guidance approach recently published addressing antiviral use in children with covid- [ ] . guidance statements were developed based on best available evidence and expert opinion. given the lack of currently available randomized controlled trials for the therapies considered in this document and the overall limited nature of the data, a systematic review was not performed, nor was evidence formally evaluated using grading of recommendations, assessment, development, and evaluation (grade) or other methodology. as previously described [ ] , we used the following definitions and similarly assert the importance of "first do no harm" in the consideration of proposed immunomodulatory therapies with yet unknown efficacy in the setting of covid- . statements using the term "suggest" indicate the panel's view that currently available evidence is weighted towards risk or benefit from a proposed therapy. guidance statements of "consider" reflect uncertainty by the panel with regard to risk or benefit of a proposed therapy. the panel considered three major questions related to immunomodulatory therapy for children with covid- : . are immunomodulatory agents indicated in children with covid- ? . what criteria define the pediatric population in whom immunomodulatory therapy may be considered? . what agents, if any, are preferred if immunomodulatory therapy is considered for children with covid- ? in addressing these questions, we utilize the definitions of severe and critical covid- previously published (table ) [ ] . we provide background information for several categories of immunomodulatory therapies that have been proposed for potential use in caring for severely or critically ill covid- patients. these categories include il- inhibitors, il- inhibitors, glucocorticoids, convalescent plasma, janus kinase inhibitors, intravenous immunoglobulin (ivig), and interferons. within each section we provide a guidance statement followed by rationale and an a c c e p t e d m a n u s c r i p t evidence summary. in vitro and animal model data is reviewed in selected sections, though we do not significantly address sars-cov- animal models given a current lack of such published data. in addition, we provide information regarding potential adverse events and practical guidance regarding dosing within each section. a summary of key guidance statements is provided in table . guidance statement: we emphasize that the vast majority of children with covid- the aforementioned antiviral guidance document provided key rationale for consideration of therapies beyond supportive care in children with covid- [ ] . multiple studies and experience in a variety of settings have demonstrated that the majority of children with covid- recover without immunomodulatory interventions and do not develop severe manifestations. given the lack of available results from randomized-controlled trials of immunomodulatory therapy in children with covid- , the risk-benefit ratio for most pediatric patients points toward supportive care as the key management strategy. however, a subset of pediatric patients develops severe or critical illness with acute covid- [ ] . it is therefore possible that immunomodulation is a key part of treatment strategy for such patients. guidance statement: we suggest that immunomodulatory therapy only be used for pediatric patients in the setting of confirmed critical covid- (sars-cov- rt-pcr positive) with evidence of hyperinflammation (table ). in addition, pediatric covid- patients with hyperinflammation whose pace of illness progression suggests imminent progression to critical covid- based on currently available data, very few pediatric patients with covid- will become severely or critically ill [ ] [ ] [ ] ] . therefore, for the majority of pediatric covid- patients the risks of immunomodulatory therapy outweigh potential benefits. however, in pediatric patients with critical covid- (defined previously; table , [ ] ), or who are rapidly progressing towards this category, the potential benefits of immunomodulatory therapy may offset the potential risks. in addition, given the potential risks of immunomodulatory therapy, consideration of this therapy in the setting of severe or critical acute covid- should be reserved for patients with rt-pcr-confirmed infection. we are aware that there may be a subset of patients for whom there is a high suspicion of acute covid- despite negative rt-pcr testing. given the difficulties in determining disease etiology and defining benefits of immunomodulatory therapy, we do not provide specific guidance for this scenario. in the evidence summary below, we review the current data on immunopathology in severe and critical covid- and discuss potential clinical and laboratory criteria on which to base the decision to use immunotherapy. adult patients severely affected by covid- demonstrate a variety of overlapping phenotypes of severity including features of ards, hypercoagulability, hyperinflammation/css, and multi-organ failure [ ] [ ] [ ] . currently published cohorts indicate that a very large percentage of covid- affected children do well after infection with sars-cov- . despite these overall reassuring findings in children, reports have emerged that a small subset of pediatric patients are severely affected by covid- with a presentation/severity similar to that seen in adult patients [ , , , ] . the clinical and laboratory presentation of patients with severe acute sars-cov- infection has revealed similarities and differences with css. css is associated with dysregulated, inappropriate, and unbalanced immune responses that include enhanced production of proinflammatory cytokines (table ). css is driven by excessive activation of both innate (monocytes, macrophages, neutrophils, nk cells) and adaptive immune cells (t-cells) and overproduction of pro-inflammatory cytokines that produce a recognizable clinical and laboratory pattern of tissue pathology. in general, css is associated with evidence of aberrant systemic inflammation including elevated ferritin, low fibrinogen, cytopenias, hemophagocytosis, and variable occurrence of coagulopathy, nk cell dysfunction, and pulmonary, liver, spleen and/or cns involvement [ ] . current reports of severely and critically ill covid- patients indicate that aberrant and dysregulated inflammation contributes to morbidity and mortality in these patients. a c c e p t e d m a n u s c r i p t however, parsing out contributions from ards, cytokine storm, secondary infections, and coagulopathy/hypercoagulability in the laboratory findings associated with severe/critical covid- remains difficult. in addition, there is likely to be distinct pathophysiology between severe/critical covid- with css compared to other categories of css. therefore, we emphasize caution in the application of diagnostic criteria used in other css to the assessment of css in the setting of covid- current information regarding immunopathogenesis of css in acute covid- derives primarily from adult data. we have summarized much of this data below within each specific immunotherapeutic section. briefly, a subset of patients progress to severe lung injury and death. autopsy studies demonstrate exudative diffuse alveolar damage with significant capillary congestion and microthrombi [ ] and an association with venous thromboembolism in non-pulmonary sites [ ] . laboratory evidence of markedly elevated inflammation including elevated ferritin, crp, and esr values is also noted in severe/critical covid- cases [ ] . further, many patients critically ill with covid- demonstrate evidence of coagulopathy including significantly elevated d-dimer [ ] . cytokine profiling of patient samples has been performed in adult and pediatric patients. elevation in other cytokines/chemokines associated with hyperinflammation states, including cxcl- (mig), cxcl- (ip- ), ccl- (mcp- ), and il- ra have also been shown [ ] . single cell immune profiling demonstrates an inflammatory signature that includes significant inflammatory gene expression with classical monocytes [ ] . furthermore, several early studies have suggested a link between impaired innate interferon expression and development of excessive inflammation in covid- patients [ ] [ ] [ ] [ ] . based on this data, several cohorts reporting use of immunomodulation in covid- have been recently published [ , [ ] [ ] [ ] [ ] . these manuscripts and related reports are reviewed in specific sections below. the panel recommends that use of immunomodulatory therapy for the treatment covid- related hyperinflammation/ cytokine storm should be conducted in the context of a clinical trial, if available. in the absence of such opportunity, and recognizing that definitive evidence is lacking, consideration for use of immunomodulatory agents in cases of sars-cov- infection with clinical and biochemical evidence of cytokine storm physiology (e.g., features of secondary hlh) should be limited to patients with clear evidence of critical covid- disease and risk for multi-organ failure. in this restricted scenario, an experimental approach using immunotherapy has theoretical potential for benefit and is supported by increasing evidence, as detailed below. we propose several key categories of clinical information to consider regarding the use of immunomodulatory therapy for pediatric covid- patients (table ). these include clinical and laboratory illness features and a c c e p t e d m a n u s c r i p t take into account the pace of illness progression, evidence of organ injury/impending organ failure, and evidence of hyperinflammation. importantly, given the current state of knowledge and lack of available clinical trial results, we are not able to provide specific cutoffs or laboratory results that indicate a definite need for immunomodulatory therapy. although current literature has identified proposed laboratory findings demonstrating hyperinflammation as indicators of risk for severe/critical covid- , these are not validated and individual values should be assessed in the context of the patient's overall status. no single feature or laboratory value is known to be sufficient to recommend immunotherapy. however, it is anticipated that timely recognition and intervention could improve outcomes, such that trends toward worsening disease and the cadence of change should be considered. despite the overlap with css and noted elevations in inflammatory markers and pro-inflammatory cytokines, diagnostic criteria for familial hlh (fhlh), mas, and crs should not be necessarily be used to identify covid- patients who may benefit from immune suppression or immune modulation, as these definitions are likely to differ for sars-cov- infections, just as they do between the different css categories. consideration for use of experimental therapies should entail discussion between the patient's primary team and appropriate consulting teams with experience in the use of immunomodulatory drug treatment in the setting of infection including infectious diseases, rheumatology, and/or hematology/oncology, critical care, and with involvement of pharmacists. further, use of these therapies should be performed only with appropriate counseling and consenting of patients and families for off-label use of immune modulatory medications according to each individual institution's policies. guidance statement: there are no immunomodulators with proven efficacy for the treatment of covid- in pediatric patients as of july . therefore, no guidance can be provided to support the use of one immunomodulatory therapy over another. as outlined in topic-specific sections below, there are no randomized, controlled trials evaluating the use of immunomodulatory therapies in pediatric covid- patients. numerous cohort studies have recently been published though many of these are limited by absence or inadequacy of a rigorous comparator group. a few randomized controlled trials have recently been published. however, none of these provides comparison between immunomodulatory agents. therefore, the current state of evidence does not allow for selection of one immunomodulatory therapy over another. as with the decision to use or not use an immunomodulator in a pediatric covid- patient, the choice of which immunomodulatory therapy to use should be driven by an individualized weighing of potential risks and potential benefits. in addition, though superseded by evidence of efficacy and adverse effects, relative drug availability and cost may play a role in immunomodulatory therapy choice. for each immunomodulatory therapy addressed below, we provide rationale, evidence summary, potential risks, and practical considerations to assist in these individualized patient care decisions. the order of discussion of immunomodulatory therapies below does not reflect any preference for one category over another. guidance statement: given to clinical trial enrollment if available. the effects of il- can be inhibited by blocking binding to the il- receptor using monoclonal antibodies such as tocilizumab, siltuximab, and sarilumab. notably, these are each being tested for use in covid- in over a dozen clinical trials that are currently recruiting patients. given that the pediatric experience with il- inhibition has mostly been with use of tocilizumab, the panel would favor use of this agent, should this therapeutic modality be considered, although other agents may also be considered in select situations (e.g. anaphylaxis with tocilizumab or drug shortages). a c c e p t e d m a n u s c r i p t tocilizumab is fda approved (albeit not for this indication), and dosing data are extrapolated from that used in css following car t cell or blinatumomab therapy. mechanism and current uses: il- is a pleiotropic cytokine produced by a number of nonhematopoietic cells and cells of myeloid origin during infections and in response to tissue injury. il- binds its receptor (il- r) and initiates a jak/stat-mediated signaling pathway, which results in transcription of numerous genes [ ] . increased il- levels are observed in a number of viral infections, and in animal models elevated il- levels favor the persistence of some of these viruses [ ] [ ] [ ] [ ] . tocilizumab is currently fda approved for the treatment of car t cell-induced crs in both children and adults, rheumatoid arthritis and giant cell arteritis in adults, as well as polyarticular and systemic juvenile idiopathic arthritis in children [ ] . in vitro and animal data: a number of prior in vitro studies demonstrated that infection of airway epithelial cells or macrophages [ , ] with either sars or mers coronaviruses could elicit production of il- and tnf-; similar results were also observed with purified coronavirus spike (s) or nucleocapsid (n) proteins [ , ] . in an animal model, primary infection with sars is associated with an il- gene signature and an associated self-sustaining acute phase response [ ] . human data: several studies have examined the cytokine response to sars-cov- ; these have demonstrated the presence of mild to moderately elevated il- in serum or plasma of adult and pediatric covid- patients [ , , ] . moreover, the levels of il- transcript and protein appear to correlate with the severity of covid- and mortality in adults [ , , ] . a proportion of pediatric covid- patients were also shown to possess similarly elevated il- levels [ ] . while blinded and randomized clinical trial data are still needed, several recently published cohort studies have indicated mixed evidence for benefit for tocilizumab in adult covid- patients. in a single center cohort study of adult covid- patients requiring mechanical ventilation, treatment with tocilizumab was associated with decreased mortality with a hazard ratio (hr) for death of . ( % ci . - . ) after adjusting for disease severity at tocilizumab initiation [ ] . similarly, in an italian multicenter retrospective cohort study of severe covid- adult patients, tocilizumab treatment was associated with decreased risk of mechanical ventilation or death (adjusted hr . ; % ci . - . ) [ ] . these results are supported by several smaller cohorts [ , , [ ] [ ] [ ] [ ] [ ] a c c e p t e d m a n u s c r i p t potential risks: while it is generally recommended that tocilizumab not be initiated in patients with neutropenia or thrombocytopenia or in those with elevations of alanine aminotransferase or aspartate aminotransferase, in practice many patients with severe or life-threatening systemic inflammatory response syndrome (sirs) experience cytopenias or elevated transaminases due to multi-system organ dysfunction or the use of concurrent medications. therefore, it may be reasonable to cautiously initiate tocilizumab therapy in such patients with close monitoring. additionally, tocilizumab therapy may increase risk of bacterial and mycobacterial infections (especially the reactivation of m. tuberculosis), viral reactivation (especially hepatitis b), and invasive fungal disease [ ] . notably most of these reports are from adults (e.g. those with rheumatoid arthritis) who have received chronic il- inhibition and, therefore, risk for children on shorter courses of tocilizumab may not be as significant. interestingly, the above cited studies have shown an increase in superinfection in patients treated with tocilizumab though without clear impact on outcomes [ , ] . other adverse events such as pneumatosis intestinalis and intestinal perforation [ ] , hepatic injury and liver failure [ ] , hypertriglyceridemia and pancreatitis [ ] have also been reported. note that dosing for tocilizumab in pediatric populations is largely based on use for rheumatologic indications and in car t cell-associated crs [ , ] . the suggested intravenous (iv) dosing is mg/kg for patients with a total body weight less than kg and mg/kg for those kg and above, with a maximum dose of mg. monitoring for anaphylaxis should be performed and, if noted, should be treated using standard local protocols. additionally, periodic laboratory (e.g. complete blood counts (cbcs), hepatic and pancreatic function testing, etc.) and clinical exam monitoring is suggested. of note, while tocilizumab does not directly affect the p cytochrome system, elevated levels of il- can inhibit these enzymes, and thus drugs that are metabolized by this system may also require monitoring. the half-life of tocilizumab is concentration dependent and is estimated to be up to weeks in high-dose therapy (as suggested for use in covid- hyperinflammation) of adult patients receiving chronic therapy at mg/kg every weeks. most adult studies of covid- -associated hyperinflammation use a single dose with some studies providing two doses separated by - hours if there is lack of response to the first dose. finally, since tocilizumab blocks the il- receptor, following il- levels is not useful for monitoring and is not recommended. a c c e p t e d m a n u s c r i p t guidance statement: given to clinical trial enrollment if available. if il- inhibition is used as a treatment modality for pediatric covid- patients, we suggest the use of anakinra based on its safety profile and favorable pharmacokinetics. available data from the prior sars and mers outbreaks and early data from the current sars-cov- pandemic suggest that il- β may play a role in sars-cov- -related immunopathology. evidence that inhibition of il- -signaling may safely improve outcomes in covid- [ ] [ ] [ ] ] . mechanism and current uses: il- α and il- β are two of the members (including il- and il- ) of the il- cytokine family and play a critical role in a wide variety of pro-inflammatory states. il- β is the major biologically active and secreted form in the setting of infection and other inflammation, but il- α is likely released by dying endothelial cells. il- β exerts its effects through binding to its specific receptor subunit il- r followed by co-receptor recruitment. biological effects of il- β include recruitment of endothelial adhesion molecule expression and inflammatory cell recruitment, upregulation of prostaglandins and nitric oxide, and metalloproteinase production. systemically, il- β contributes to hypotension, fever, neutrophilia, and other acute phase responses. importantly, il- β contributes to cd + th differentiation by contributing to key aspects of transcriptional activation [ ] . il- can also act upstream to increase il- expression [ ] . several targeted therapies inhibit il- β signaling including anakinra, a recombinant form of il- receptor antagonist (il -ra) which mimics native il- ra and prevents binding of il- α and il- β a c c e p t e d m a n u s c r i p t to il- r and thereby prevent il- -mediated immune effects [ ] . another il- targeting drug, rilonacept, binds and neutralizes both il- α and il- β. in addition, a monoclonal antibody, canakinumab, binds il- β and prevent its interaction with il- r [ ] . anakinra is fda approved for adults with rheumatoid arthritis who have failed one or more disease modifying antirheumatic drug (dmard) and for the treatment of neonatal onset multisystem inflammatory disease (nomid). randomized controlled trials of each of the medications have demonstrated their safety with infections occurring infrequently when used in these settings [ ] [ ] [ ] [ ] [ ] [ ] [ ] . anakinra has been best studied in terms of treating other css, including macrophage activation syndrome and secondary hemophagocytic lymphohistiocytosis. it is a recombinant human protein with over , patient-years of favorable safety data. anakinra has a short half-life of - hours, can be given intravenously or subcutaneously, and works quickly. anakinra also has a wide therapeutic window (effective with minimal adverse effects reported from - mg/kg/day, including use in patients with sepsis). retrospective analysis of a large randomized clinical trial demonstrated that anakinra improved survival in sepsis patients with features of css (hepatobiliary dysfunction and disseminated coagulopathy) from % (placebo) to % [ ] . an early case series of pediatric rheumatic disease patients with refractory css reported resolution in out of patients treated with anakinra [ ] . more recently, a retrospective report of children with rheumatologic, oncologic, and infectious etiologies of css reported % survival rate for those who received anakinra at any point during their hospitalization [ ] . further, anakinra has also been touted to effectively treat css in children requiring intensive care [ ] . importantly, for interpreting results and studies summarized below, measurement of il- β in peripheral blood compartments such as plasma and serum may not accurately reflect its biological activity in a variety of disease states. this is likely due in part to its short half-life and tight regulation of il- β's activity in human immune responses via soluble-il- receptors and il- ra, among other regulators [ ] . in vitro and animal data: several sars proteins (e protein, protein a, and orf b) are shown in in vitro studies to activate the nlrp inflammasome. protein a and e protein each activate the nlrp inflammasome in lps-primed mouse bone marrow derived macrophages and african green monkey kidney-derived vero cells, respectively [ ] [ ] [ ] [ ] . a c c e p t e d m a n u s c r i p t human data: an association between il- β and sars-associated immunopathology was made predominantly through measurement of serum/plasma il- β levels in sars-cov- infected patients. in general, these studies are limited by a lack of control groups and/or measurement of il- β at few or inconsistent time points post-infection [ , [ ] [ ] [ ] . immunohistochemistry of lung sections from autopsy specimens of four sars patients did demonstrate elevated il- β expression in sarsinfected cells [ ] . following several small case series [ , ] , two retrospective cohort studies have shown potential benefit for anakinra in the context of severe covid- . adult covid- patients with moderate to severe ards and evidence of hyperinflammation (crp ≥ mg/l or ferritin ≥ ng/ml) were treated with either iv or subcutaneous anakinra [ ] . comparison between the subjects in the high dose iv anakinra treatment group and historical controls indicated improved survival in the treatment group at days post treatment initiation ( %, anakinra; % standard, p= . ) though mechanical ventilation free survival differences were not statistically significant. no differences in bacteremia rates or frequency of hepatic transaminase elevation were noted [ ] . in a separate study in france, adult patients with severe covid- treated with subcutaneous anakinra were compared to historical controls [ ] . anakinra-treated subjects had significant decreased frequency of the primary composite outcome of death or intensive care unit admission for mechanical ventilation ( % versus %) with difference remaining significant on multivariate analysis (hr . ; % ci . - . ). thus, anakinra appears to be safe and may provide mortality benefit in the treatment of adult covid- patients. randomized clinical trials are needed to validate these findings. adverse effects of anakinra are difficult to distinguish versus effects of the disease processes being treated but include hematologic suppression, infections, hypersensitivity reactions, and malignancies. those most commonly reported in pediatric studies include increased liver enzymes, which are usually self-limiting, but a few cases of acute liver failure have been reported. in addition, severe injection site reactions and cytopenias have been reported [ , ] . importantly for the discussion of high versus standard dose anakinra below, high dose anakinra was associated with increased risk of serious bacterial infection in a pooled analysis of studies enrolling adults with ra [ ] . though the majority of studies of anakinra have been performed using subcutaneous (sc) dosing route, iv administration is evolving as an option for the treatment of critically ill patients [ , , ] . the potential utility of iv anakinra in covid- patients is highlighted by the above cited study in which low dose sc anakinra ( mg twice daily in adult patients) did not produce either significant clinical or laboratory changes in a small subset of covid- patients while the mg/kg iv over hour administered every hours was associated with sustained clinical benefit [ ] . despite these encouraging results, stability data for iv anakinra are incomplete and caution is warranted in the use of iv anakinra. as noted above, measurement of il- levels in peripheral blood is difficult and therefore not recommended during anakinra administration. for toxicity, close monitoring of ast and alt should be performed as these can become elevated on anakinra therapy. in general, hepatic transaminase elevation resolves with discontinuation or lowering of dose. cbc with differential should be monitored to evaluate for anakinra-induced leukopenia and/or thrombocytopenia. though increased infections are a reported risk with the use of anakinra, anakinra has a low rate of secondary infection risk and a long record of safety in clinical practice [ , , ] . in the use of anakinra in covid- , tuberculosis screening is not needed to initiate therapy because of clinical urgency for therapy initiation. guidance statement: given their pleiotropic effects, glucocorticoids are used in a variety of inflammatory conditions and settings. as such, it is difficult to definitively support or discourage the use of glucocorticoids in all situations. therefore, we have provided guidance for specific situations in the following section. given the concerns over immune dysregulation associated with covid- , glucocorticoids have been proposed and used as a potential treatment modality [ ] [ ] [ ] . glucocorticoids regulate the immune system in a broad and multimodal manner and block multiple signaling pathways that propagate inflammatory signals [ , ] . during the early phase of the immune response, glucocorticoid-glucocorticoid receptor complexes attenuate the signaling of toll-like receptors, inhibit the production of numerous pro-inflammatory cytokines [ ] , and dampen cytokine signaling [ ] . glucocorticoids also have a potent effect on cellular immunity, especially t cell a c c e p t e d m a n u s c r i p t signaling and activation [ ] . overall, glucocorticoids are one potential therapeutic option for the treatment of covid- , but benefits offered by glucocorticoids in attenuating immune dysregulation must be balanced with their inhibitory effect on the immune response needed to control viral replication as well as the risk of opportunistic infections and associated side-effects. glucocorticoids, however, are available readily and at low cost. given the numerous clinical uses for glucocorticoids and mechanisms by which they impact the immune system, we have limited this evidence summary to address prior data from sars, and currently available data from covid- . data regarding the efficacy of glucocorticoids in the treatment of sars are difficult to interpret. glucocorticoids were frequently used as part of many treatment protocols, each with different dosing regimens and varied times of treatment initiation; thus most studies did not contain placebo arms [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . a systematic review of the treatment effects of multiple therapeutic modalities for sars, including glucocorticoids, revealed that / studies examining the effect of steroids were inconclusive [ ] . no studies showed clear benefit, and studies showed possible harm. ultimately, the authors concluded that "it is difficult to make a clear recommendation about whether [glucocorticoids] should be used to treat sars-associated lung injury in any stage of illness, particularly as the drug is immunosuppressive and may delay viral clearance if given before viral replication is controlled" [ ] . peer-reviewed data evaluating the impact of steroids on covid- treatment outcomes is limited, and comparative data is lacking. glucocorticoids have been used frequently in critically ill patients [ , , ] . one retrospective cohort study of adult patients with covid- identified risk factors for the development of ards and death from ards [ ] . sixty-two patients received methylprednisolone, but there were limited data on the dose and timing of initiation of therapy. for those patients with ards, treatment with methylprednisolone reduced the risk of death (hazard ratio, . ; % confidence interval, . - . ). however, these data should be interpreted with caution due to small sample size and risk of bias. an additional retrospective cohort study performed in a multicenter health system in michigan showed potential benefit of early methylprednisolone initiation (median time to initiation of days post admission, iqr - days) compared to a standard of care cohort with later corticosteroid initiation (median time to initiation of days with iqr of - days). in this study, early a c c e p t e d m a n u s c r i p t corticosteroid initiation was associated with decreased occurrence of escalation of care to icu, mechanical ventilation, or death (adjusted or . ; % ci . - . ) [ ] . recently, data from the recovery trial, a randomized clinical trial evaluating the impact of treatment with dexamethasone in adults infected with sars-cov- , were published [ ] . a total of , adults were enrolled, and , patients received mg of dexamethasone daily for up to days. for the primary outcome of -day mortality, receipt of dexamethasone was associated with a significant decrease in mortality for those patients receiving invasive mechanical ventilation ( % versus %, rate ratio . , % confidence interval . to . ) or for those patients who received supplemental oxygen but not mechanical ventilation ( % versus %, rate ratio . , % confidence interval . to . ). receipt of dexamethasone was also associated with lower risk of progression to invasive mechanical ventilation, and patients in the dexamethasone group had a shorter duration of hospitalization. there was no difference in -day mortality, however, for patients who were not receiving any respiratory support at the time of randomization. conversely, there was a slight trend towards worse outcomes in those receiving no respiratory support (mortality % vs %). at time of writing, the pediatric portion of the recovery trial is still enrolling patients. similarly, results from a large ( received glucocorticoids vs. , who did not) noncontrolled retrospective cohort comparison reported decreased mortality or mechanical ventilation (odds ratio . ; % ci, . - . ) in those with high crp values ( mg/dl or higher) receiving glucocorticoids within hours of hospital admission [ ] . however, mortality or mechanical ventilation was increased in those receiving glucocorticoids if the crp was less than mg/dl (odds ratio, . ; % ci, . - . ). finally, it is worth noting that some centers are reporting their experience with a combined tocilizumab plus corticosteroid strategy for severe/critical covid- patients [ , ] . current evidence is too limited to determine whether this strategy is truly safe, effective, and/or applicable to pediatric covid- patients. thus, there is evolving evidence to support benefit of corticosteroid treatment for critically ill adult patients with covid- . the degree to which findings from these studies are applicable to children with severe or critical covid- is not clear at this point. despite this uncertainty, glucocorticoid therapy could be considered in select clinical scenarios based on individualized riskbenefit assessment. scenario-specific guidance follows. we have not provided specific dosing a c c e p t e d m a n u s c r i p t recommendations beyond the below guidance statements due to the lack of covid- specific evidence for dosing at present. guidance statement: glucocorticoid therapy is not currently indicated for outpatients or hospitalized patients with mild or moderate covid- . based on currently available data, most sars-cov- infected children, even those with mild or moderate disease, will recover with supportive care. based on sars and mers studies in which glucocorticoid recipients had delayed viral clearance [ , ] , administration of glucocorticoids may attenuate the immune response needed to clear viral infection. in addition, no evidence exists that glucocorticoid therapy prevents progression from mild/moderate to severe covid- . therefore, the panel recommends against the use of glucocorticoids in children without symptoms or who have only mild or moderate disease. glucocorticoid therapy may be considered for pediatric patients with critical covid- with preference for use in the setting of clinical trials, if available. as described above with regard to il- and il- inhibition, current evidence suggests that excessive diagnosis with covid- does not preclude use of steroids when they are otherwise indicated (for example, in asthma or catecholamine-refractory shock). glucocorticoid therapy offers benefit in the treatment of many pediatric conditions such as asthma exacerbation or flares in inflammatory bowel disease [ , ] . in the critical care setting, there is limited pediatric data surrounding the efficacy of glucocorticoids in septic shock, with published studies containing small numbers of children [ ] [ ] [ ] . a recent meta-analysis evaluating the efficacy of glucocorticoids in sepsis included published rcts of which enrolled only children and enrolled both adults and children [ ] . pooled analysis showed that use of glucocorticoids may decrease both short-term and long-term mortality, though any effect is likely small. a recent cochrane review included rcts, though only trials included children [ ] . the authors found a small reduction in -day mortality in the pooled analysis, though there was significant heterogeneity across trials. there is no high-quality pediatric data to support the routine use of glucocorticoid therapy for the treatment of sepsis without shock or shock that is responsive to fluid resuscitation or vasopressors. however, if a child with covid- develops circulatory shock and remains hypotensive despite fluid resuscitation and titration of vasoactive drugs, use of glucocorticoids can be considered for treatment of critical-illness related corticosteroid insufficiency. this condition is characterized by dysregulated systemic inflammation resulting from inadequate glucocorticoid-mediated antiinflammatory activity relative to the severity of the patient's critical illness [ ] . hydrocortisone is the synthetic form of cortisol, and there is significant experience with its use in the treatment of circulatory shock. published dosing regimens in this setting are based dosing ranges for the use of hydrocortisone in pediatric adrenal insufficiency [ , ] . prior guidelines for critically ill adults with covid- have also addressed this topic [ , ] . short-term use of glucocorticoids is associated with significant adverse effects, including hypertension and fluid retention, hyperglycemia, adrenal suppression, gastritis and gastrointestinal bleeding, posterior reversible encephalopathy syndrome [ ] , and psychosis [ , ] . we recommend routine screening for electrolyte abnormalities, hyperglycemia and hypertension in hospitalized children receiving steroids. these studies are often part of routine care. after the sars pandemic, there were reports of avascular necrosis after treatment with glucocorticoids [ - a c c e p t e d m a n u s c r i p t ]. in symptomatic patients, or those at increased risk of avascular necrosis, screening with plain radiographs or mri should be considered. glucocorticoid therapy is also associated with immunosuppression and potentially increases the risk of secondary infection. in one recent meta-analysis of the use of glucocorticoids as adjunctive treatment for influenza, adults treated with glucocorticoids had increased odds of hospital-acquired infection, though the overall quality of evidence was low [ ] . in addition, several reports of adult covid- patients indicate risk for invasive pulmonary aspergillosis [ , ] . whether this risk is similarly present in children with covid- remains unclear. regardless, monitoring for secondary infection should occur for all patients receiving glucocorticoids. monitoring for drug-drug interactions is essential with glucocorticoid use. glucocorticoids are metabolized through cytochrome p , a , one of the more common hepatic isoenzymes. if this isoenzyme is inhibited by another agent (e.g. macrolide, protease inhibitor), significantly increased glucocorticoid exposure can result. alternatively, glucocorticoid exposure can also be significantly reduced by an inducer of cyp a such as rifampin requiring increased dosing to obtain the same effect [ ] . guidance statement: while there exists a theoretical rationale for the use of jak inhibitors in severe covid- , the exact clinical impact of these drugs is difficult to predict, and it is unclear whether treatment with these agents would prove beneficial or harmful. additionally, there are very little data supporting safety or efficacy in the use of jak inhibitors for management of covid- . moreover, data from use in other settings suggests the potential for impaired viral clearance as evidenced by herpesvirus reactivations on jak inhibitor therapy [ ] . therefore, currently, we recommend against the use of these drugs for children with covid- outside of clinical trials. mechanism and current uses: the janus kinases (jaks) are a family of four tyrosine kinases (jak , jak , jak , and tyk ) that serve as intracellular signal transducers [ ] . following the binding of cytokines to types and cytokine receptors on the cell surface, jaks initiate a cascade of intracellular signaling that leads to activation or suppression of gene transcription. more than cytokines signal via the jak/stat pathway to coordinate hematopoiesis, induce inflammation and control the immune response [ ] . cytokine receptor subunits bind specific jaks, although some bind more than one. complete receptors are thus associated with a pair of jaks, and individual cytokines signal through a variety of jak combinations. given the key role played by jaks in hematopoiesis and immune signaling, these enzymes are an important target for pharmacologic inhibition. several oral small molecule jak inhibitors have been recently developed, and these drugs have efficacy in treating a number of neoplastic and inflammatory conditions [ ] . emerging pediatric uses for these drugs include treatment of juvenile idiopathic arthritis [ ] , psoriasis [ ] , interferonopathies [ ] , and graft versus host disease [ , ] . in addition to the above conditions, jak inhibitors are also increasingly studied in the setting of css such as hlh and mas [ ] [ ] [ ] [ ] . in a small open-label series of five adults with secondary hlh, ruxolitinib initiation was temporally associated with improvement in cytopenias and declines in both ferritin and soluble-il- receptor (sil r) [ ] . in vitro and animal data: data summarizing key cytokines and pathways relevant for jak inhibitors are discussed in other sections. human data: a few small studies have evaluated the utility of jak inhibitor therapy for covid- in adults have recently been published. a single-blinded, randomized controlled trial of adults with severe covid- showed that ruxolitinib did not significantly enhance clinical improvement compared to placebo [ ] . ruxolitinib-treated patients in this study did show more rapid radiographic and more rapid recovery from lymphopenia as well as decrease in peripheral blood cytokine expression. in a small cohort study, fifteen adult patients hospitalized with covid- (with requiring icu care and requiring mechanical ventilation) received baracitinib treatment [ ] . twelve of the fifteen patients survived though no comparator group was provided in this analysis. finally, in a multicenter study from italy, baracitinib-treated patients with moderate covid- were compared to historical controls that did not receive baracitinib [ ] . icu admission and mortality a c c e p t e d m a n u s c r i p t at weeks post-enrollment was decreased in the baracitinib versus control groups (icu: . % vs. . %, p= . ; mortality: % vs. . %, p= . ). thus, clinical evidence is lacking to strongly support the use of jak inhibitors for covid- patients at this point. however, large prospective studies assessing both efficacy and adverse effects are needed prior to consideration for use of these medications in children. when studied for treatment of rheumatoid arthritis, jak inhibitors have been shown to have a similar increased risk of infection compared to placebo as with biologic agents (e.g. adalimumab). however, increased risk of primary or reactivation of herpes simplex and varicella zoster virus infections has been specifically noted for jak inhibitors [ , ] . these agents have also been associated with anemia, lymphopenia or neutropenia (and thrombocytopenia in patients with myelofibrosis treated with ruxolitinib), and with elevated cholesterol and abnormal liver function tests [ ] . these effects are generally mild but occasionally require drug dosage decrease, and in cases of moderate to severe lymphopenia or neutropenia, discontinuation of the drug. indeed, a two-patient case series recently reported demonstrated adverse effects of ruxolitinib therapy for covid- , including anemia, thrombocytopenia, soft tissue infection, and herpes labialis [ ] . of note, baracitinib has a black box warning for potential increased risk of thrombosis which merits close attention in light of high reported incidence of thromboembolism in hospitalized covid- patients [ , ] . finally, several jak inhibitor agents (fedratinib, ruxolitinib and tofacitinib, specifically) are considered strong substrates of cyp a , and therefore put patients at risk for multiple clinically relevant drug interactions including with -azole antifungals [ ] . of particular interest, care providers should use extreme caution in patient receiving potent cyp a inhibitors (-azole antifungals) and potent inducers of a (phenobarbital and phenytoin) as this combination will likely cause subtherapeutic response or increase risks for adverse event with even single dose therapy. [ ] . while the passive transfer of polyclonal antibodies via convalescent plasma may be of benefit, no sars-cov- -specific monoclonal antibody therapies have yet been clinically tested. with respect to coronaviruses, use of cp was trialed in mers and more extensively in sars [ ] [ ] [ ] . while the sars case series suggested possible benefit from cp for patients (especially those treated before day of illness), the true efficacy of this approach remains difficult to ascertain due to lack of control groups, study biases, and use of concomitant therapeutics, all of which confound the interpretations; nonetheless, two meta-analyses of the published case series did not reveal significant harm [ , ] . with respect to sars-cov- , plasma collected from patients in the convalescent phase of infection has been used as an empirical treatment in small numbers of patients with severe covid- disease, with some laboratory improvements and disease mitigation observed [ ] [ ] [ ] [ ] [ ] . a small randomized controlled trial has also been conducted, although, was stopped early due to challenges with enrollment [ ] . human data: two studies demonstrate that while the patterns of development of igm and igg can differ among different individuals, nearly all covid- patients eventually do develop appreciable antibody titers [ , ] . these and other studies collectively show that the titers of sars-cov- specific antibodies appear to be stable over the few ensuing weeks and that the majority of the neutralizing antibodies are directed against the spike protein, and specifically the portions of this protein that are responsible for the binding of virus to the ace receptor. as such, these studies a c c e p t e d m a n u s c r i p t suggest that the cp of many covid- patients may contain sufficient quantities of neutralizing antibodies for therapeutic utility. there have been several case reports and series examining passive transfer of cp to covidafflicted adult patients with moderate to critical disease [ - , , ] . not all of these case series reported the titers of sars-cov- antibodies in the transferred plasma. most of these case series reported that subsequent to cp, most patients displayed stabilization or improvement of disease, as evidenced by resolution of fever, decreases in c-reactive protein and inflammatory cytokine levels, and improved radiographic findings; in some cases these improvements resulted in the extubation of mechanically ventilated patients and weaning from ecmo support. however, in one case series, in which patients were treated with cp at a median time of . days from detection of sars-cov- , of patients eventually died [ ] , even though the virus could no longer be detected. while the numbers of patients reported to have been treated with cp to date are too small to draw any definitive conclusions, similarly to the use of cp in sars-cov- [ ] , these reports do suggest that if cp is to be effective, it may need to be used earlier in the course of disease. the largest case series to date, including hospitalized adults at several centers across the united states, demonstrated only severe adverse events, with only of these severe adverse events judged to be definitively related to the infusion of convalescent plasma (joyner et al jci). this study was not designed to evaluate the efficacy of convalescent plasma. as of july , a single randomized controlled trial has evaluated convalescent plasma for sars-cov- [ ] . this trial enrolled patients with severe or life-threatening covid- but was discontinued early due to poor accrual and thus was not powered sufficiently to answer questions of efficacy. there was no clinically significant difference found in the primary measure of time to clinical improvement. similarly, no significant difference was identified in -day mortality or time to discharge. however, convalescent plasma infusion was associated with a statistically significant negative conversion rate of viral rt-pcr at hours post infusion. there is a single case report of the use of convalescent plasma for a pediatric patient [ ] . potential risks: there are a number of potential risks associated with the use of convalescent plasma therapy, including those that may result from the transfer of immunoglobulins, and those that may ensue from transfusion of human blood products. as noted above, current reports indicate that cp-infusion is generally safe. therefore, we focus below on immunoglobulin-dependent risks including ade and exacerbation of deleterious immune responses, as well as the potential for inhibiting the development of effective humoral immunity. ade is a phenomenon in which complement pre-existing (or transferred) virus-specific antibodies could increase the entry of virus into cells expressing fc receptors; ade has been demonstrated to occur in a variety of viruses in vitro and in animal studies, and postulated to occur with coronaviruses [ ] . ade is believed to occur with sub-neutralizing or non-neutralizing antibodies and higher viral titers. notably, ade has not been demonstrated in the passive transfer of convalescent plasma in patients infected with sars-cov- and mers. yet, the risk of ade still remains a concern since some patients who have recovered from mild covid- may have delayed development of significant titers of neutralizing antibody titers against sars-cov- , and since not all current convalescent plasma protocols mandate the use of cutoffs for neutralizing antibody titers [ , ] . in addition to ade, transfer of virus-specific antibodies may also exacerbate the inflammatory response and lead to further lung injury; this was demonstrated in vitro and in a sars-cov- macaque infection model [ ] . another theoretical concern is that use of exogenous immunoglobulins may inhibit endogenous development of adequate titers of high affinity antibodies that could protect the patient from re-challenge with the same virus, as has been demonstrated with respiratory syncytial virus [ , ] . in addition to the antibody-mediated concerns, there are risks associated with transfer of human blood products. these include transmission of infectious pathogens (sars-cov- and other viruses, as well as parasites such as babesia spp. or trypanosoma cruzi), and transfusion-related reactions including anaphylaxis, hemolysis, or transfusion-related acute lung injury (trali) or transfusion related acute cardiac overload (taco) [ ] . while passive immunotherapy protocols all include measures to minimize the possibility of such harm, some cannot be predicted or prevented, thus necessitating close monitoring in the days to weeks following therapy. we do not currently recommend use of ivig for treatment of acute covid- in pediatric patients with the exception of specific clinical scenarios in which ivig is typically used. importantly, our recommendations do not apply to the use of ivig in the treatment of mis-c. studies to suggest a benefit for ivig treatment in covid- are not available. therefore, ivig is not indicated in the majority of pediatric cases. a recent study, however, has shown that lots available from the usa and other parts of the world may variably contain antibodies to sars-cov- . whether these are neutralizing antibodies or are clinically insignificant is difficult to discern at this time and future clinical trials are needed to prove any potential benefit for their use in patients with covid- [ ] . mechanism and current uses: immunoglobulin (ig) for intravenous (iv) administration, commonly referred to as ivig, although licensed in the us as igiv, contains pooled immunoglobulin g (igg) from the plasma of thousands of blood donors and contains more than % unmodified igg, and various amounts of iga depending on the formulation. ivig is an immunomodulating agent that produces effects on many components of the innate and adaptive immune system including the following: inhibition of complement activation, saturation of fc receptors on macrophages [ ] and suppression of inflammatory mediators [ ] . specific humoral effects include inhibition of b-cell differentiation, induction of b-cell apoptosis, down-regulation of specific auto-reactive b-cells, and overall inhibition of antibody production. ivig also induces the expansion of regulatory t cells (tregs) and downregulates the expansion of th cells [ ] . currently, the fda has approved the use of ivig in different clinical conditions: replacement of igg in primary and secondary immunodeficiency disorders; prevention of coronary aneurysms in kawasaki disease; chronic inflammatory demyelinating polyneuropathies and multifocal motor neuropathy to improve neuromuscular disability; and to improve platelet counts in immunemediated thrombocytopenia [ ] .ivig has also been used as an adjunct treatment in hlh along with other specific anti-cytokine therapy [ , ] . in vitro and animal data: assessment of the role of ivig in sars, mers, or covid- has not been evaluated through in vitro or animal model studies. human data: in the sars-cov- outbreak, thrombocytopenia was reported to occur in up to % of patients and was identified as a significant risk factor for mortality. the etiology of this thrombocytopenia was likely multifactorial, but may have been immune complex mediated [ ] . while no data is available regarding ivig use in the setting of sars-cov- or mers infection, several other viral infections (e.g., hiv, hepatitis c, parvovirus b and zika virus) may be associated with secondary itp and ivig has been used as therapy in these settings. among patients with severe thrombocytopenia with zika virus who received ivig, the median platelet count increase was × /l, in contrast to the median increase of . × /l in the patients who received platelet transfusions [ ] . since severe thrombocytopenia may be associated with increased mortality in covid- disease [ , ] , there may be a role for ivig in treatment regimens for pediatric covid- patients with thrombocytopenia. however, a causal relationship between thrombocytopenia and covid- -a c c e p t e d m a n u s c r i p t associated mortality is not established and the mechanisms for covid-associated thrombocytopenia are not known. therefore, benefit of ivig in the care of pediatric covid- patients is not defined. given the nature of the product and its source, immediate hypersensitivity and infusion related reactions such as headaches, flushing of the face, malaise, chest tightness, fever, chills, myalgia, dyspnea, nausea, vomiting, diarrhea, change in blood pressure, and tachycardia may occur and may be more likely in iga-deficient patients who are receiving products that contain iga [ ] . most of these reactions can be managed or resolved by appropriate premedication regimens and slowing rates of infusion. renal injury may be more likely in those with preexisting renal disease, volume depletion, sepsis and concomitant nephrotoxic drug usage, and may also be specific to the ivig product used. other specific considerations should include thromboembolism and volume considerations with administration of ivig, especially in patients with underlying cardiac disease. dosing recommendations for ivig vary by indication. therefore, we do not provide extensive dosing guidance here. in pediatric patients with macrophage activation syndrome-related css an ivig dose of g/kg has been used [ ] . a recent publication reported ivig dosing of . - . mg/kg x days in a case series of covid- patients [ ] . there are three types of interferons: type i, type ii and type iii. type i ifn includes ifn- and -, type ii interferon is ifn- and type iii includes ifn-. while type i interferon is produced by all cell types, plasmacytoid dendritic cells, fibroblasts, and monocytes are among the main sources [ ] . type i ifn secretion is induced by any cell type upon encountering viral signatures. in addition, type i ifns enhances the clearance of virally infected cells by cytotoxic cd t cells by increasing the expression of mhc class i. other effects that further enhance antiviral responses include the activation of dendritic cells, macrophages and nk cells, and the induction of the production of chemokines such as cxcl , - , and - [ ] . unlike the ubiquitously expressed ifn- receptor, the ifn- receptor is expressed in epithelial cells and a subset of immune cells, including neutrophils [ , ] . based on its receptor expression, it is thought that type iii ifn responses have critical roles in epithelial and mucosal antiviral immunity [ , ] . when used in patients, ifn- due to limited expression of its receptor in the immune compartment results in less systemic side effects than type i ifn therapy [ ] . type i ifn has been extensively used in the management of hepatitis b and c infections. due to side effects, modest efficacy, and the advent of effective antivirals, the use of ifn- has decreased in the management of hepatitis b [ , ] . additionally, ifn- has also shown clinical efficacy similar to type ifn in the management of hepatitis c [ ] . in vitro and animal data: in vitro studies have shown ifn- and - have antiviral activity. however, ifn- has a more potent coronavirus suppression activity than ifn-α * - ] . studies in murine models have shown that ifn- in combination with antivirals improves pulmonary function but does not reduce viral replication or severe lung pathology [ ] . similar reduction in mortality was also noted in non-human primates treated with ifn-β b following mers-cov infection [ ] . though there are no corresponding coronavirus murine model studies with ifn-, murine models of influenza a virus (iav) infection showed promising results. ifn-λ treatment resulted in enhanced epithelial barrier function and suppressed initial viral spread without activating the systemic effects seen with ifn-α therapy * , +. importantly, while ifn-λ treatment of iav-infected mice lowered the viral load and protected from disease, treatment with ifn-α decreased the viral load but exacerbated disease [ ] . human data: it was recently suggested that one component of sars-cov- -related immunopathology is insufficient robust type i, ii or iii ifn response [ ] [ ] [ ] [ ] ] . this finding along with the known antiviral properties of both type i and type iii ifn form the basis of its use in sars-a c c e p t e d m a n u s c r i p t cov- [ ] . currently, there is only one published clinical trial of ifn therapy in covid- [ ] . in this multicenter, open-label randomized study in hong kong, control subjects received lopinavirritonavir and adult subjects received intervention therapy with lopinavir-ritonavir as well as oral ribavirin and subcutaneous ifn-β b. subjects were comparable with regard to clinical and laboratory characteristics and underwent treatment initiation a median of days after symptom onset. subjects in the group receiving the addition of ribavirin and ifn-β b to lopinavir-ritonavir had shorter time to complete symptom resolution and shorter time to negative nasopharyngeal swab compared to the control group. further studies of interferon-β are ongoing, including with administration via inhalation which may decrease systemic side effects (nct ). therapy with type i or type iii ifn is associated with a variety of common adverse effects including fatigue, anorexia, nausea, diarrhea, alopecia, fever, rigors, headache, and myalgia [ , ] . additionally, neuropsychiatric events including depression are reported [ ] . finally, neutropenia and anemia are also reported. additionally, there is theoretical risk for worsening disease with the use of interferon therapy. as morbidity due to covid- appears to be due in large part to hyperinflammation, potential exists for interferon therapy to result in enhanced inflammation and thereby lead to worsening of the css [ ] . further, there is a concern that type i and iii ifns may impair successful antibacterial immune responses and thereby contribute to increased susceptibility to secondary bacterial infections [ , ] . as we do not suggest use of interferon therapy in pediatric covid- patients, potential dosing and other considerations are not provided. in addition to the above described immunomodulatory therapies, a number of other agents and immune pathways are being evaluated for treatment of covid- . these include immunosuppressive medications traditionally used in the setting of solid organ or hematopoietic cell transplantation, such as tacrolimus (nct ), sirolimus (nct , nct ), and ctla -fc fusion molecules. the anti-ifn-γ antibody, emapalumab, has recently been demonstrated to have efficacy in the treatment of pediatric primary hlh [ ] and is also being evaluated in the treatment of covid- (nct ). a c c e p t e d m a n u s c r i p t current data demonstrate that the vast majority of pediatric patients with acute covid- recover from their initial illness without significant morbidity or 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interferon expression by sars-cov- supports clinical trials of interferon lambda to treat early covid- triple combination of interferon beta- b, lopinavir-ritonavir, and ribavirin in the treatment of patients admitted to hospital with covid- : an open-label, randomised, phase trial premature discontinuation of interferon plus ribavirin for adverse effects: a multicentre survey in 'real world' patients with chronic hepatitis c neuropsychiatric adverse effects of interferon-alpha: recognition and management lambda interferon restructures the nasal microbiome and increases susceptibility to staphylococcus aureus superinfection disease-promoting effects of type i interferons in viral, bacterial, and coinfections emapalumab in children with primary hemophagocytic lymphohistiocytosis the authors would like to thank drs. jason newland, kathleen chiotos, and mari nakamura and the sharing antimicrobial reports for pediatric stewardship (sharps) collaborative for their support in developing this document a c c e p t e d m a n u s c r i p t a c c e p t e d m a n u s c r i p t table . suggested covid- illness severity categories (modified from [ ] ) no new or increased supplemental oxygen requirement. new or increase from baseline supplemental oxygen requirement without need for new or increase in baseline non-invasive/invasive mechanical ventilation. new or increased requirement for invasive or non-invasive mechanical ventilation, sepsis, or multi-organ failure; or rapidly worsening clinical trajectory that does not yet meet these criteria.comments: non-invasive mechanical ventilation includes high-flow nasal canula, continuous positive airway pressure (cpap), or bilevel positive airway pressure (bipap).a c c e p t e d m a n u s c r i p t we suggest that immunomodulatory therapy only be used for pediatric patients in the setting of confirmed critical covid- (sars-cov- pcr positive) with evidence of hyperinflammation (table ). in addition, pediatric covid- patients with hyperinflammation whose pace of illness progression suggests imminent progression to critical covid- may be considered for immunomodulatory treatment. use of immunomodulation for pediatric covid- should be performed in consultation with specialists familiar with these medications. iii. which immunomodulatory agents should be considered?  there are no immunomodulators with proven efficacy for the treatment of covid- in pediatric patients as of july . therefore, no guidance can be provided to support the use of one immunomodulatory therapy over another.  if immunomodulators are used in the treatment of covid- , patients should be monitored for adverse effects. note: the order of discussion of each category below does not denote an order of preference. a. il- inhibition il- inhibition may be considered in the care of pediatric patients with critical covid- with priority given to clinical trial enrollment if available. b. il- inhibition il- inhibition may be considered in the care of pediatric patients with critical covid- with priority given to clinical trial enrollment if available. if il- inhibition is used as a treatment modality for pediatric covid- patients, we suggest the use of anakinra based on its safety profile and favorable pharmacokinetics. given their pleiotropic effects, glucocorticoids are used in a variety of inflammatory conditions and settings. as such, it is difficult to definitively support or discourage the use of glucocorticoids in all situations. therefore, we have provided guidance for specific situations in the following section.  glucocorticoid therapy is not currently indicated for outpatients or hospitalized patients with mild or moderate covid- .  glucocorticoid therapy may be considered for pediatric patients with critical covid- with preference for use in the setting of clinical trials, if available.  diagnosis with covid- does not preclude use of steroids when they are otherwise indicated (for example, in asthma or catecholamine-refractory shock).  elevated bilirubin, ggt, and/or transaminases renal  decreased creatinine clearance abbreviations: bnp, brain natriuretic peptide; crp, c-reactive protein; css, cytokine storm syndrome; pt, prothrombin time; ptt, partial thromboplastin time; ggt, gamma-glutamyl transferase key: cord- -l f hg authors: amor, sandra; fernández blanco, laura; baker, david title: innate immunity during sars‐cov‐ : evasion strategies and activation trigger hypoxia and vascular damage date: - - journal: clin exp immunol doi: . /cei. sha: doc_id: cord_uid: l f hg innate immune sensing of viral molecular patterns is essential for development of antiviral responses. like many viruses, sars‐cov‐ has evolved strategies to circumvent innate immune detection including low cpg levels in the genome, glycosylation to shield essential elements including the receptor binding domain, rna shielding and generation of viral proteins that actively impede anti‐viral interferon responses. together these strategies allow widespread infection and increased viral load. despite the efforts of immune subversion, sars‐cov‐ infection activates innate immune pathways inducing a robust type i/iii interferon response, production of proinflammatory cytokines, and recruitment of neutrophils and myeloid cells. this may induce hyperinflammation or alternatively, effectively recruit adaptive immune responses that help clear the infection and prevent reinfection. the dysregulation of the renin‐angiotensin system due to downregulation of angiotensin converting enzyme , the receptor for sars‐cov‐ , together with the activation of type i/iii interferon response, and inflammasome response converge to promote free radical production and oxidative stress. this exacerbates tissue damage in the respiratory system but also leads to widespread activation of coagulation pathways leading to thrombosis. here, we review the current knowledge of the role of the innate immune response following sars‐cov‐ infection, much of which is based on the knowledge from sars‐cov and other coronaviruses. understanding how the virus subverts the initial immune response and how an aberrant innate immune response contributes to the respiratory and vascular damage in covid‐ may help explain factors that contribute to the variety of clinical manifestations and outcome of sars‐cov‐ infection. the emergence in wuhan china of a novel severe acute respiratory syndrome coronavirus (sars-cov- ) triggered an epidemic of the coronavirus disease (covid- ) . as of september th , the confirmed , , cases including , deaths have been reported worldwide (worldometers.info/coronavirus). at the end of january , the who declared covid- a pandemic and a global health emergency. the family coronaviridae is subdivided into torovirinae and coronavirinae that contains the genera alphacoronavirus, betacoronavirus, gammacoronavirus, and deltacoronavirus. the human coronaviruses (hcov) belong to the αlpha-cov (hcov- e and hcov-nl ) and beta-cov (middle east respiratory syndrome coronavirus-mers-cov, sars-cov, hcov-oc and hcov-hku ) [ table ; ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ]. in comparison with most hcovs that cause mild upper respiratory tract infections, sars-cov, mers-cov and sars-cov- induce severe pneumonia ( ) . the clinical presentation of covid- ranges from mild 'flu-like' symptoms to severe respiratory failure and death although between . - % of sars-cov- infections are asymptomatic depending on the population ( ) . common symptoms include fever, cough, fatigue, shortness of breath, headache and pneumonia. in addition, some patients develop gastrointestinal problems ( ) , and neurological manifestations, including headache, dizziness, hyposmia and hypogeusia. age and comorbidities i.e., hypertension, chronic obstructive pulmonary disease, diabetes, obesity and cardiovascular disease predispose to more severe manifestations, including severe respiratory failure, septic shock, coagulation dysfunction, strokes, cardiovascular problems ( ) and neurological manifestations ( ) . although the origin and transmission of sars-cov- is unclear, genome sequencing reveals marked similarities with sars-cov ( ) . however, in comparison, sars-cov- spreads more quickly than sars-cov, likely due to the - % fold higher in infectivity and transmissibility during the initial non-symptomatic period ( - days) . in some cases, transmission has been reported after development of initial symptoms despite the presence of antibodies, ( ) indicating that both, neutralising antibodies and t cell responses, are necessary to prevent reinfection and for protection ( ) . this is further supported by studies showing pd + cd + t cell exhaustion, depletion or inactivation is associated with viral persistence in severe cases ( ) . sars-cov- is a positive-sense rna ( , nucleotides) enveloped virus of to nm diameter ( ) . the envelope is studded with homotrimers spike proteins of - nm length that are heavily decorated with n-glycans [figure ( , ) ]. similar to other hcovs, sars- this article is protected by copyright. all rights reserved non-structural proteins (nsp) while orfs - encode the viral structural proteins -spike, envelope, membrane and nucleocapsid, and the accessory proteins [figure b]. differences between the structural, non-structural and accessory proteins of sars-cov- and other coronaviruses help to explain the high infectivity rate and the range of pathologies observed ( , , ) . while knowledge of sars-cov is rapidly emerging, parallels with sars-cov, as well as ongoing sequencing data and antigenic typing will be crucial to understand the dynamics of the pandemic. sars-cov- cell entry is similar to sars-cov being mediated by the binding of the receptor-binding domain (rbd) of the s protein, to the angiotensinconverting enzyme- (ace- ), although other receptors such as cd and cd-sign have been reported [ table ]. docking of the rbd to the receptor and the action of furin, a serine protease that separates the s and s proteins exposes a second binding domain on s allowing membrane fusion. binding of the s protein to ace- requires priming by cell proteases -primarily tmprss , however, tmprss is expressed by a subset of ace + cells supporting the notion that the virus likely uses other host enzymes such as tmprss , lysosomal cathepsins and neuropilin- ( ) to augment the impact of furin and expose the rdb thus promoting sars-cov- entry ( ) . the structural proteins m, e and n are crucial for stability of the viral genome and viral replication. the nsp and accessory proteins ( ) encoded by open reading frames (orfs) have differing functions during viral replication [table and many also act to deviate the innate immune response thus augmenting viral replication and spread. the degree to which the innate immune system is suppressed and evaded clearly determines the viral load and the host's outcome to infection, the clinical symptoms and the severity of the disease. following infection, viral rna is sensed by several classes of pattern recognition receptors (pprs). the retinoic acid-like receptors (rlrs) include retinoid inducible gene i (rig-i) and melanoma differentiation-associated gene (mda ), toll-like receptors (tlr) -classically , and that trigger ifn pathways and cytokines production [ figure ]. once engaged these pprs act downstream via the kinases tank-binding kinase- (tbk ) and inhibitor-b kinases (ikks). such triggering leads to the activation of the transcription factors interferonregulatory factor- (irf ) and (irf ) and nuclear factor kappa-light-chain-enhancer of activated b cells (nfb). these subsequently induce expression of type i ifns (ifnα/β) and interferon stimulated genes (isgs) [figure ] many of which have potent antiviral activities, as well as other proinflammatory mediators e.g. cytokines, chemokines and antimicrobial peptides that are essential to initiate the host innate and adaptive immune response. in addition, the absent in melanoma (aim )-like receptors and nod-like receptors (nlrs) this article is protected by copyright. all rights reserved trigger the inflammasome and il-  and il- production leading to pyroptosis [ figure ]. immune responses include c-type lectins and the stimulator of interferon genes (sting). while the cgas/sting pathway is commonly associated with sensing cytosolic dna, it is also activated following binding of enveloped viruses to host cells and cytosolic viral rna ( , ) . similar to tlrs and rlr, downstream, sting engages tbk to active irf and/or nfb inducing type i ifn and/or proinflammatory cytokines [ figure ]. coronaviruses have evolved several strategies to escape such innate immune recognition allowing widespread replication. such evasion includes evolution of low genomic cpg, rna shielding, masking of potential key antigenic epitopes as well as inhibition of steps in the interferon type i/iii pathways. generally, the zinc finger antiviral protein (zap) specifically binds to and degrades cpg motifs in genomes of rna viruses. in comparison with other viruses, sars-cov- has evolved the most extreme cpg deficiency of all betacoronavirus [ ( ) . another strategy to protect mrna used by the host and many viruses is the processing of capping the ′ end. for both host and virus rna, capping limits degradation and importantly blocks recognition by cytosolic pprs. like many rna viruses sars-cov- has exploited several mechanisms to protect the ′ ends by a cap structure of rna generated during replication. while some viruses snatch the caps from host rna, sars-cov- , like other coronaviruses uses its own capping machinery composed of nsp , nsp and the this article is protected by copyright. all rights reserved proteins of - nm length that are heavily decorated with glycans. each spike protein comprises of two subunits (s and s ) that each bear glycan groups ( ) . cell entry of the highly glycosylated s protein of sars-cov is promoted by dc-sign possibly augmenting virus uptake or aiding capture and transmission of sars-cov by dcs and macrophages ( ) ( ) ( ) . similar to the spike protein, the other structural, non-structural and accessory proteins are also modified by glycosylation, palmitoylation, phosphorylation, sumoylation and adpribosylation ( ) . conversely, some viral proteins e.g. nsp , possess deubiquitinating (dub) and deisgylation activity thereby interfering with host functions targetting those that are critical for signalling transduction of innate immunity ( ) . insertion of the spike protein into cell membranes during replication is a key step for virus budding. whilst this takes place in the rtc [suppl figure ], receptor-bound spike proteins interact with tmprss expressed on the uninfected cell surface, mediates fusion between infected and uninfected cells promoting the formation of syncytia allowing the virus to spread to adjacent uninfected cells while evading detection by the immune response ( ) . in addition to strategies to evade ppr recognition, sars-cov- has also evolved strategies to inhibit steps in the pathway leading to type i/iii ifn production. this may be especially relevant in the lungs where type ifn iii (lambda) is considered to be more effective in controlling viral infections and critically affected in covid- . knowledge arising from the study of other coronaviruses, especially sars-cov and mers, has shown that many of the non-structural, structural and accessory proteins interfere with elements of the ifn pathway [ table , figure ], essential for the development of effective immunity. ifn antagonism has been attributed to several of the structural, non-structural and accessory proteins that interfere with stimulator of interferon genes (sting)-traf -tbk complex, thereby blocking sting/tbk /ikkε-induced type i ifn production, stat- / translocation to the nucleus, irf , nfb signalling as well as interfering with the actions of the isg products including ifits [ table ]. as examples, nsp , and , and orf interfere with stat- / signalling while nsp , and cap the viral rna [ table ] preventing recognition by rig-i, mda and ifits. nsp also acts by dub proteins thereby preventing their activity such as rig-i and other steps in the ifn pathways for which ubiquitination is essential. cov plpro (nsp ) also interrupts the stimulator of interferon genes sting.traf . tbk complex thereby blocking sting/tbk /ikkε-type i ifn production ( , ) . as well as subversion of the ifn pathway, sars-cov orf a (also present in sars-cov- ) blocks the activity of tetherin also known as bone marrow stromal antigen (bst- ) ( ). bst acts by tethering budding viruses to the this article is protected by copyright. all rights reserved cell membrane thus preventing its release from the cells. orf a removes this inhibition aiding the release of mature virions. in summary, emerging evidence from sars-cov- , and comparison with other sars-cov and mers, reveals many strategies used to evade the innate immune response and subvert the interferon pathway. while this facilitates widespread viral replication increasing the viral load also promotes the viral cytopathic effects leading to tissue damage described below, likely leads to exacerbation and hyperinflammation of the innate immune response once triggered. despite immune evasion and subverting innate immune responses during early infection, sars-cov- effectively initiates immune signalling pathways. this is likely due to the increased viral load that exponentially produces viral rna and viral proteins (pathogen associated molecular patterns -pamps), also induces cell damage that release damage associated molecular patterns (damps) both of which trigger innate immune pathways. like sars-cov and nl , sars-cov- uses the angiotensin (ang)-converting enzyme- (ace ) as a cell receptor [ table ], expressed on epithelia in renal, cardiovascular and gastrointestinal tract tissues, testes and on pneumocytes and vascular endothelia ( ) . ace regulates the renin-angiotensin system (ras) by balancing the conversion of angiotensin i to angiotensin - and angiotensin ii to angiotensin - . binding of sars-cov- to ace leads to endosome formation, reducing ace expression on the cell surface [figures and ] and pushing the ras system to a pro-inflammatory mode triggering production of reactive oxygen species, fibrosis, collagen deposition and a proinflammatory environment including il- and a balanced response to infection via tlr pathway is essential to trigger a protective response to sars-cov ( ) . this study also supports the idea that in addition, pamps, this article is protected by copyright. all rights reserved immune pathways triggered by damps such as oxidised phospholipids, high mobility group box (hmgb ), histones, heat shock proteins and adenosine triphosphate released by damaged cells may contribute to covid- outcome [figures and ] . in addition to rig-i, mda and mavs, rna viruses are also sensed by the stimulator of interferon genes (sting) that is activated by cgamp when enveloped rna viruses interact with the host membranes ( ) . downstream, sting engages tbk that actives irf and/or nfb inducing type ifn and/or proinflammatory cytokines. that hyperactivation of sting contributes to severe covid- as has been hypothesised by berthelot and lioté ( ) . these authors present several lines of evidence, the strongest being that gain of function mutations of sting associated with hyperactivation of type i ifn induces the disease savi (sting-associated vasculopathy with onset in infancy). affected children with savi present with pulmonary inflammation, vasculitis and endothelial-cell dysfunction that mimics many aspects of covid- ( ) . furthermore, sting polymorphisms are associated with ageing-related diseases such as obesity and cardiovascular disease, possibility explaining the impact of comorbidities and development of severe covid- ( ) . also, in bats in which sars-cov- may have arisen, sting activation and thus consequently ifn is blunted ( ), likely aiding viral replication and spread as observed in early sars-cov- infection in humans. that damps released due to viral cytotoxicity may contribute to severe covid- which is best exemplified by hmbg released by damaged and dying cells as well as activated innate immune cells especially in sepsis ( ) . depending on its conformation hmgb triggers tlr , tlr and tlr , the receptor for advanced glycation end-products (rage) and triggering receptor expressed in myeloid cells (trem- ) [ figure ]. in mice, intratracheal administration of hmgb activates mitogen-activated protein kinase (mapk) and nfκb, inducing proinflammatory cytokines, activating the endothelium and recruiting neutrophils in the lung -key pathological features of severe covid- ( , ) . hmgb , and especially the platelet-derived source may play a crucial role in sars-cov- vascular damage since hmgb -/mice display delayed coagulation, reduced thrombus formation and platelet aggregation ( ) . furthermore, blocking hmgb is beneficial in experimental lung injury and sepsis, suggesting therapies targeting hmgb might also be beneficial in severe covid- ( ) . studies of peripheral blood and post-mortem tissues from severe covid- cases reveal high levels of il- β and il- and increased numbers of cd +il- β monocytes, suggesting activation of the nod-like receptor family, pyrin domain-containing (nlrp ) inflammasome pathway ( ) . activation of the nlrp inflammasome, essential for effective antiviral immune responses, is elicited by several factors associated with sars-cov infection including ras this article is protected by copyright. all rights reserved disbalance, engagement of ppr, tnfr and ifnar, mitochondrial ros production, complement components including mac, as well as sars-cov viral proteins such as orf a, n and e [figure , table ]. as a consequence, nlrp interaction with adaptor apoptosis specklike protein (asc) recruits and activates procaspase- , processing pro-il- β and pro-il- to the activate forms [ figure ]. this drives the propyroptotic factor gasdermin d (gsdmd) formation of pores in the cell membrane, i.e. pyroptosis that facilitates the release of proinflammatory cytokines. the pores also aid the release of cellular damps such as hmgb , and viral pamps that further exacerbate inflammation suggesting that targeting the nlrp pathway might be beneficial in severe covid- cases. the delayed interferon response, increased viral load and virus dissemination, coupled with the release of damps and pamps lead to activation of several innate immune pathways. following infection, pneumocytes, epithelial and alveolar cells, and infiltrating monocytemacrophages and neutrophils likely produce the first wave of tnfα, il- , ip- , mcp- , mip-  and rantes production ( , ) . hyperinflammation is likely promoted by comorbidities due to increased ace expression, concurrent bacterial infections and ageing as well as a direct effect of sars-cov- replication since virus-host interactome studies reveal that sars-cov- nsp regulates the nfb repressor factor nkrf, facilitating il- production ( ). this is followed by a second wave of cell recruitment including nk cells that produce ifn and further recruitment of (alternatively activated) monocytes/macrophages and neutrophils sars-cov- exploits many strategies to subvert innate immune responses allowing the virus to replicate and disseminate within the host. the extent to which the virus replicates within this article is protected by copyright. all rights reserved the host, and the efficacy of the host innate immune response to eradicate the infection and trigger effective adaptive immune responses, but not hyper-responsiveness of innate immunity, strongly determines the disease outcome [ table ]. the severity of infection has been linked to age, smoking, comorbidities such as cancer, immune suppression, autoimmune diseases, inflammatory disease, neurodegenerative diseases, obesity, gender and race ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) . for example, in a large cohort of , cases the case fatality ratio for over years was . % versus . % in the total cohort ( ) . this is likely higher due to inflamm-ageing, an aberrant innate immune response such as lower production of ifnβ ( ), increased oxidative stress ( ) and sensence of macrophages that become less effective in their reparative functions with age ( ). likewise, viral load, obesity, gender, race, blood groups and comorbidities have all been reported to influence the response to sars-cov- infection, [ table ; ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ] although few studies have fully examined the extent to which subversion and activation of innate immune components contribute to susceptibility in these cases. understanding the innate immune factors that exacerbate the vascular complications will be crucial to control severe disease following sars-cov- infection. rapidly emerging studies reveal the extent to which therapeutic approaches for other viral infections and inflammatory diseases can be repurposed to target innate immunity to treat covid- patients ( , ) . likewise, novel approaches have been put forward to target the susceptible ageing population or those with comorbidities. one approach under investigation is to re-establish the youthful function of macrophages and repair mechanisms using metformin, a drug used in type diabetes that has been shown to attenuate hallmarks of ageing ( ) . in a retrospective study of , subjects tested for covid- while diabetes was reported to be an independent risk factor for covid- -related mortality ( ) this article is protected by copyright. all rights reserved upr -unfolded protein response; zap -zinc finger antiviral protein. this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved (b) and hyperinflammation with influx of macrophages, nk cells and neutrophils (c). this self-augmenting cycle triggers further cell damage and damps and pamps release as well as ros production. d) activation of neutrophils induces neutrophil extracellular traps (net) aided by the n protein and generated in response to ros-induced endothelial cell damage. disruption of the vascular barrier and endothelial cell exposure to proinflammatory cytokine and ros increases expression of p-selectin, von willebrand factor (vwf) and fibrinogen, that attract platelets triggering expression of tissue factor. together this sequence activates the complement system, one of many pathways that crucially activates the coagulation cascade leading to thrombi formation. human aminopeptidase n is a receptor for human coronavirus e tmprss activates the human coronavirus e for cathepsin-independent host cell entry and is expressed in viral target cells in the respiratory epithelium infection of primary cultures of human neural cells by human coronaviruses e and oc hla class i antigen serves as a receptor for human coronavirus oc human and bovine coronaviruses recognize sialic acid-containing receptors similar to those of influenza c viruses dc-sign and dc-signr interact with the glycoprotein of marburg virus and the s protein of severe acute respiratory syndrome coronavirus cd l (l-sign) is a receptor for severe acute respiratory syndrome coronavirus human coronavirus nl employs the severe acute respiratory syndrome coronavirus receptor for cellular entry identification of major histocompatibility complex class i c molecule as an attachment factor that facilitates coronavirus hku spike-mediated infection host cell entry of middle east respiratory syndrome coronavirus after two-step, furin-mediated activation of the spike protein a 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in this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved accepted article key: cord- -p twx a authors: lau, arthur chun-wing; yam, loretta yin-chun; so, loletta kit-ying title: management of critically ill patients with severe acute respiratory syndrome (sars) date: - - journal: int j med sci doi: nan sha: doc_id: cord_uid: p twx a severe acute respiratory syndrome (sars) is frequently complicated with acute respiratory failure. in this article, we aim to focus on the management of the subgroup of sars patients who are critically ill. most sars patients would require high flow oxygen supplementation, – % required intensive care unit (icu) or high dependency care, and – % developed acute respiratory distress syndrome (ards). in some of these patients, the clinical course can progress relentlessly to septic shock and/or multiple organ dysfunction syndrome (mods). the management of critically ill sars patients requires timely institution of pharmacotherapy where applicable and supportive treatment (oxygen therapy, noninvasive and invasive ventilation). superimposed bacterial and other opportunistic infections are common, especially in those treated with mechanical ventilation. subcutaneous emphysema, pneumothoraces and pneumomediastinum may arise spontaneously or as a result of positive ventilatory assistance. older age is a consistently a poor prognostic factor. appropriate use of personal protection equipment and adherence to infection control measures is mandatory for effective infection control. much of the knowledge about the clinical aspects of sars is based on retrospective observational data and randomized-controlled trials are required for confirmation. physicians and scientists all over the world should collaborate to study this condition which may potentially threaten human existence. in , an outbreak of severe acute respiratory syndrome (sars) caused by the sars-associated coronavirus involved countries and patients, resulted in deaths [ ] . thereafter, sars has re-emerged sporadically in both laboratory and community settings. its clinical spectrum varies from minimal respiratory symptoms to severe respiratory failure. we have previously contributed to an overview on the contemporary treatment of sars [ ] , and the whole topic has also been reviewed elsewhere [ ] . in this article, we aim to focus on the management of a subgroup of critically ill sars patients with more significant respiratory failure. critically ill sars patients frequently demonstrate the following clinical features: persistent pyrexia (occasionally from admission but often recurring after an initial period of defervescence), tachycardia (infrequently bradycardia), tachypnoea and significant oxygen desaturation. more than onethird of all the sars patients required high flow oxygen therapy [ ] , - % required intensive care unit (icu) admission or high dependency care, and - % developed acute respiratory distress syndrome (ards) [ , ] . the clinical course of some of these patients can progress relentlessly irrespective of all attempts at pharmacological treatment, eventually resulting in septic shock and/or multiple organ dysfunction syndrome (mods). lymphocytopaenia, neutrophilia and thrombocytopenia are frequently seen in critically ill sars patients. neutrophilia could be due to sars per se, to superimposed infection or related to corticosteroid administration. pancytopaenia, if present, could be due to haemophagocytosis syndrome [ ] or reactivation of latent human parvovirus (unpublished data). prolonged activated partial thromboplastin time and picture of disseminated intravascular coagulation has been reported [ ] . coinfections with other agents including chlamydia-like agents [ ] , metapneumovirus [ ] or influenza virus (unpublished data) have been reported. persistent and increasing elevations of creatine kinase, lactate dehydrogenase, and transaminases levels are common [ , , ] . associated lung damage is believed to be the result of a virally-triggered inflammatory reaction mediated by a host of cytokines [ , ] . in sicker patients, levels of pro-inflammatory cytokines (il- beta, il- , il- , il- , tnf-α) and tgf-β were higher, with slower decline on clinical recovery [ ] . radiographic abnormalities in the chest usually progress upwards from initial unilateral or bilateral lower-to mid-zone peripheral ground-glass shadows, to focal, multifocal or diffuse consolidation. peak radiographic changes occurred at . days after fever onset, with . % showing two peaks at . and . days, and % showing relentless progression [ ] . cavitation is rare but may be associated with superimposed infection in patients with a prolonged illness course and who are mechanically ventilated [ ] . high-resolution computer tomography (hrct) of the thorax showed focal ground-glass and scattered "crazy paving" patterns at presentation, followed by development of interstitial thickening, consolidation, pleural reaction, and scarring and fibrosis in later stages [ , ] . small (< cm) pulmonary cysts may be detected even if the patient is not receiving ventilatory assistance [ ] . subcutaneous emphysema, pneumothoraces or pneumomediastinum are distinct complications of severe sars [ ] . hrct features of late-stage ards caused by sars are similar to those arising from other causes [ ] . lung biopsy and postmortem studies [ , ] showed acute-phase diffuse alveolar damage (dad), airspace edema, bronchiolar fibrin, increased numbers of interstitial macrophages (with focal haemophagocytosis) and alveolar macrophages in patients with shorter duration (< days) of illness. on the other hand, histology after > days of illness showed organizing-phase dad with increased fibrosis, hyperplasia of type ii pneumocyte, squamous metaplasia, multinucleated giant cells, and acute bronchopneumonia [ ] . in patients who died late in the course of this disease, high loads of viral rna were detectable by reverse transcriptase polymerase chain reaction (rt-pcr) in the lungs, bowel, lymph nodes, spleen, liver, and kidneys [ ] . general principles anti-bacterial therapy for community-acquired pneumonia in accordance with standard guidelines [ ] should always be administered before laboratory confirmation of sars-cov infection. where effective anti-viral therapy is available, it should be started as early as possible after diagnosis, and even empirically if suspicious clinical features and especially epidemiological links are present. since critically ill patients are deemed to have already progressed from the viral replicative phase to the immunopathological phase [ ] , concomitant institution of an immunomodulatory therapy should also be considered [ ] . since there are no consensus regarding the most optimal treatment regimen in these respects, we will thus review the more commonly used agents and discuss their relative merits based on published reports. when respiratory failure eventually sets in, oxygen supplementation, assisted ventilation and intensive supportive treatments will be required. ribavirin was the most commonly used empirical antiviral agent for sars. it is a broad-spectrum purine nucleoside analogue which inhibits both rna and dna viruses by interfering with nucleic acid synthesis. there is experimental evidence to show that it has immunomodulatory effects in the treatment of mouse coronavirus hepatitis [ ] . subsequently, it was found that ribavirin has no direct in vitro activity against sars-cov [ ] . higher doses given intravenously resulted in more frequent and severe adverse effects including haemolytic anaemia, elevated transaminase levels and bradycardia [ ] . lopinavir-ritonavir co-formulation (kaletra ® , abbott laboratories, usa) is a protease inhibitor for the treatment of human immunodeficiency virus (hiv) infection. it can inhibit the coronaviral proteases, thus blocking the processing of viral replicase polyprotein and preventing the replication of viral rna. ritonavir inhibits lopinavir metabolism thus increasing its serum concentration, but it has no activity against sars-cov. in a retrospective analysis in hong kong [ ] , patients who had received kaletra as rescue therapy together with high dose corticosteroids had no difference in rates of oxygen desaturation, intubation and mortality compared with a matched cohort. however, when given as initial treatment in combination with ribavirin in another subgroup of patients, there were significant reductions in the need for rescue pulsed corticosteroid therapy, intubation rate and overall mortality. in addition to the prevalence of diarrhoea among these patients which may render oral drugs more appropriate and useful, synergism between kaletra and ribavirin might have contributed to the benefits since either drug alone has only weak anti-viral activities. another hong kong study of sars patients treated with a combination of lopinavir/ritonavir and ribavirin compared with patients (historical controls) treated with ribavirin only showed that adverse clinical outcomes (ards or death) were significantly lower in the treatment group than in the historical controls at day after symptom onset. further randomised placebo controlled trials are required [ ] . interferons are a family of cytokines with important roles in the cellular immune response. interferon α has been used for sars treatment in china and canada [ , , ] . in an open-label uncontrolled study [ ] , nine patients treated with corticosteroids plus interferon alfacon- (infergen ® , intermune inc., usa) showed better oxygen saturation, faster radiographic resolution and lesser need for supplemental oxygen compared to given corticosteroids alone. in vitro testing showed that interferon β was more potent than interferon α or γ, being effective even when administered after sars-cov infection in cell culture [ ] . traditional chinese herbal medicine has been used concomitantly with other drugs to treat sars in mainland china with good results reported [ ] . however, its value in critically ill patients has not been reported. glycyrrhizin, an active component derived from liquorice roots, is effective against sars-cov in vitro [ ] . its clinical utility remains uncertain. another herbal compound, baicalin, also demonstrates anti-sars-cov activity in vitro (unpublished data). in the absence of an effective antiviral agent in the outbreak, most physicians had opted to use immunomodulatory agents, most commonly corticosteroids, in the treatment of sars [ , , , ] it is generally agreed that corticosteroids should not be used during the early viral replicative phase, and that its administration should best coincide with the onset of the immunopathological phase [ ] . clinicoradiological surrogate criteria have been used to indicate the onset of this immune hyperactive phase, thus providing a practical guide to the timing of starting corticosteroids [ ] . corticosteroid dosages should be high enough, especially in the severe cases, to abort the cytokine storm, and maintained for long enough to prevent the rebound phenomenon [ , , ] . this may be achieved by using a weightadjusted [ ] and radiographic extent-modified dosages [ ] for a period of - weeks. in one-third to half of sars patients, fever may recur while on immunomodulatory treatment due to superimposed infections, too rapid tailing of corticosteroids or persistently severe and uninhibited cytokine storm. empirical anti-pseudomonal antibiotics should then be given first. if there is no apparent clinical response, opportunistic infections like fungal infection should be excluded. if fever is accompanied by obvious respiratory deterioration in the absence of superimposed pulmonary or systemic infection, most patients can be presumed to be suffering from a severe recrudescence of the sars illness. in such critically ill sars patients, further escalation of immunomodulation is warranted. such deterioration could sometimes occur very rapidly; immediate administration of pulsed methylprednisolone therapy at - mg per day intravenously for days, followed by tapering doses in the subsequent weeks, has been associated with improved outcome [ , ] . up to one-third to one-half of critically ill sars patients may benefit from this strategy [ , , ] . because radiographic abnormalities may lag behind clinical improvement, persistent radiographic shadows per se, when accompanied by clinical improvement, do not warrant additional corticosteroids [ ] . human gamma immunoglobulins have been used in selected sars patients who continued to deteriorate despite treatment [ , ] . an igm-enriched immunoglobulin product (pentaglobin ® , biotest pharma gmbh, germany) has been used in hong kong and mainland china [ , , ] . pentaglobin at mg/kg/day for three days given to patients who deteriorated despite repeated rescue methylprednisolone and ribavirin therapy had shown some improvement in radiographic scores and oxygen requirement [ ] . it has been reported that the use of combined methylprednisolone and highdose intravenous immunoglobulin ( . g/kg) daily for three consecutive days in probable sars patients with acute lung injury (ali) or ards had resulted in lower mortality and a trend towards earlier recovery [ ] . randomized controlled trials in larger numbers of patients are required to confirm its efficacy. based on the assumption that the neutralizing immunoglobulins in convalescent plasma can curb increases in viral load, convalescent plasma collected from recovered sars patients has been used in hong kong to treat severely ill patients not responding to corticosteroids. some clinical benefits were reportedly observed in a small number of patients [ ] . despite all efforts, at least % of sars patients would still develop acute hypoxemic respiratory failure, with up to % requiring supplemental oxygen [ ] overall, - % of patients had been admitted into icu, and - % eventually required intubation and mechanical ventilation [ ] . both non-invasive and invasive ventilatory support has been applied to critically ill sars patients. niv delivers continuous positive airway pressure (cpap) or bi-level pressure support through a tight-fitting facial or nasal mask. it was commonly employed in many chinese hospitals [ , , , , ] and our own centre in hong kong [ , , ] . early application may be beneficial because it could rapidly improve vital signs, oxygenation and tachypnoea [ , ] , and may reduce the need for increasing dosages of corticosteroids for progressive respiratory failure. it could avoid intubation and invasive ventilation in up to two-thirds of critically ill sars patients [ , , ] . use of niv in immunocompromised subjects of other diseases has reported similarly reduced rates of endotracheal intubation and serious complications [ ] . niv in sars may be of particular benefit, since high dose corticosteroids per se would already predispose to ventilator-associated pneumonia, and risks to healthcare workers (hcw) could also be markedly reduced through obviating the need for intubation, a potentially highly infectious procedure. patients who respond to niv will usually do so within hours, non-responders who will eventually need endotracheal intubation can thus be identified early [ ] . niv is indicated in the presence of ali and early ards when oxygen saturation (spo ) could not improve to more than % despite > litres per minute of oxygen; persistent tachypnoea of at least breaths per minute; and progressive radiographic deterioration in the lungs [ ] . the usual contraindications to niv apply, including impaired consciousness, uncooperative patient, high aspiration risk, and haemodynamic instability [ ] . sars-related respiratory failure responds readily to niv given at low pressures. cpap of - cm h o, or bi-level pressure support with inspiratory positive airway pressure (ipap) of < cm h o and expiratory positive airway pressure (epap) of - cm h o are reasonable starting pressures [ ] . higher pressures should be avoided whenever possible, because it may increase the risk of pneumothorax and pneumomediastinum, which are frequently spontaneous complications of sars even without assisted positive pressure ventilation [ ] . when patients do not improve within one to two days of niv or continue to deteriorate, or if niv is contraindicated, endotracheal intubation and mechanical ventilation should be considered. most centres [ ] adopted a ventilatory strategy similar to that recommended for ards from other causes [ ] . both pressure and volume control ventilation may be employed [ ] . the tidal volume should be kept low (e.g. - ml/kg predicted body weight), and plateau pressures maintained below cm h o. because of a higher risk of barotraumas in sars, the lowest positive end-expiratory pressure (peep) which could achieve satisfactory alveolar recruitment and oxygenation, usually - cm water, should be employed. other adjunctive measures employed in the usual ards cases had been tried in sars, including: prone positioning [ , ] , high frequency oscillatory ventilation [ , ] , nitric oxide [ ] , high peep and regular lung recruitment [ ] , but their efficacy is uncertain. tracheostomy is required in patients requiring prolonged mechanical ventilation and icu stay. strict adherence to infection control guidelines is mandatory in performing tracheostomy in the icu or operating room, as well as during subsequent changes of the tracheostomy tube [ , ] . critically ill sars patients on high dose corticosteroids and mechanical ventilation are particularly susceptible to superimposed bacterial and opportunistic infections. their peripheral blood cd +, cd + and cd + were also lower than normal [ , ] . ventilator-associated infection with organisms like pseudomonas aeruginosa, methicillin-resistant staphylococcus aureus, acinetobacter baumanii, as well as invasive mucor sp [ ] and aspergillosis [ , ] have been reported. strict control of hyperglycaemia during corticosteroid administration is essential to reduce the chance of septic complications [ ] . spontaneous subcutaneous emphysema, pneumothoraces and pneumomediastinum are common complications that are potentially aggravated by noninvasive or invasive ventilation [ ] . while chest drain insertion is useful to relieve pneumothoraces, prolonged air leak may sometimes occur. by itself, sars predominantly results in single organ failure of the lungs. other complications reported are more likely the result of sepsis and its attending problems, including acute renal failure ( %), acute liver failure ( %), rhadomyolysis, cardiovascular dysfunction, or of prolonged immobilization and underlying co-morbidities, including deep vein thrombosis, pulmonary embolism, ischaemic strokes, etc [ ] . the case-fatality ratio (cfr) of sars has been estimated to range from % to > % depending on the age group affected. the overall cfr is approximately % [ ] . variability may be due to different host and viral factors as well as treatment strategies. cfr may also be significantly affected by the duration of follow-up and inclusion of different mixes of suspected, probable and laboratory confirmed cases in different series [ ] . based on the treatment principles presented above, we have developed a standard treatment protocol early on in the outbreak, comprising initially high (but not pulsed) dose methylprednisolone with tapering over three weeks [ ] . this protocol was eventually applied to consecutively admitted sars patients [ ] . their mean age was years, with % having laboratory-confirmed sars. a low overall mortality of . % ( / ) was obtained, with all three deaths occurring in patients over the age of years. twenty four percent required icu admission: % received niv (bi-level pressure support) alone and % had both niv and invasive mechanical ventilation. hrct thorax in all survivors taken days after commencement of treatment showed most did not have clinically significant lung scarring. another multi-centered study comparing four treatment regimens in guangzhou, china, also found that a regimen of high dose corticosteroids adjusted according to clinical and radiological severity, coupled with nasal cpap ventilation, produced the best result: zero mortality in all clinically-defined sars patients, mean age . years. with % treated with cpap and none requiring mechanical ventilation. subsequently, very low mortality was again recorded among a further patients treated with the same regimen [ ] . many prognostic factors have been reported to independently predict adverse outcome in sars. they include advanced age [ , , , ] , diabetes [ , , ] , heart disease [ , ] , other significant coexisting conditions [ , , ] , shortness of breath on admission [ ] , degree of hypoxaemia [ ] , high total leukocyte count on admission [ , , ] , high initial lactate dehydrogenase [ , , ] , low platelet counts [ ] , and use of pulsed doses of corticosteroid [ , ] . compared to patients with nasopharyngeal aspirates negative for sars-cov by rt-pcr, pcr-positive ones are more likely to require icu care and mechanical ventilation, develop acute renal failure and die [ ] . in particular, mortality was high among icu patients: -day icu mortality was variously reported to be - % [ , , ] . older age, severity of illness, lymphocyte count, decreased steroid dose, positive fluid balance, chronic disease or immunosuppression, and nosocomial sepsis were associated with poor icu outcome [ ] . patients who had diarrhoea were more likely to require ventilatory support and icu care [ ] . higher serum sars-cov concentration in the early stage of the disease was a prognostic indicator for later icu admission [ ] . patients presenting with more extensive radiographic involvement also predicted the need for icu care or death [ ] . age alone is a consistent and strong prognostic factor in all series. age-stratified death rates were estimated to be < % in patients below years of age, % between and years, % between and years, and > % in elderly patients over years old [ ] . corresponding estimates in hong kong were % in those below years of age, and % in those over years [ ] . the cause of death in sars is usually progressive respiratory failure with or without concomitant sepsis. sudden cardiac arrest is also possible, and has been hypothesized to be due to hypoxemia (which would worsen during activities including defaecation), direct viral myocardial injury and extreme anxiety, all of which may lead to electrical instability in the myocardium and induction of arrhythmia [ ] . sars is primarily transmitted by direct or indirect contact of mucous membranes (eyes, nose, or mouth) with infectious respiratory droplets or fomites [ , ] . transmission risks increase with duration and proximity of contact. infection control precautions in the icu are shown in appendix [ ] . endotracheal intubation should be considered earlier and in anticipation of impending deterioration, so that ample time is available for preparation. it should be performed by the most skilful airway practitioner in a negative-pressure room behind closed doors. should the operator choose to wear additional personal protective equipment like the airmate hepa powered air purifying respirator system ( m, mn, usa), he/she must be familiar with its mode of operation and the precautions required for gowning and degowning, and must be assisted by a colleague with similar knowledge [ ] . a "modified awake" intubation technique has been suggested as the best possible compromise between patient and operator safety by administration of a combination of midazolam, fentanyl and lidocaine until the patient reaches the desired level of sedation [ ] . the patient is then paralysed after intubation to minimize coughing. alternatively, the "rapid sequence induction" technique with intravenous administration of midazolam and suxamethonium can also minimize patient coughing. it should however be emphasized that, unless there is prior preparation for a surgical airway, neuromuscular paralysis should be avoided in anticipated difficult intubation in order to maintain spontaneous respiration [ ] . both bronchoscopy and niv should be performed in a negative pressure room. although there is widespread fear of infective risk by niv [ , ] , centres with such experience, including ours, have found that its use is safe if the necessary precautions are taken [ , , , ] . finally, strict adherence to infection control measures in the form of strict isolation and effective cohorting, early diagnosis and contact tracing, timely reporting and institution of public health measures, as well as enhancement of environmental ventilation is key components in the effective management of infectious diseases. managing critically ill sars patients is a challenging task. most, if not all, knowledge 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prognostic factors in patients with severe acute respiratory syndrome in hong kong department of communicable disease surveillance and response. consensus document on the epidemiology of severe acute respiratory syndrome severe acute respiratory distress syndrome (sars): a critical care perspective outcome of coronavirus-associated severe acute respiratory syndrome using a standard treatment protocol outcomes and prognostic factors in patients with severe acute respiratory syndrome in hong kong prognostic factors for severe acute respiratory syndrome: a clinical analysis of cases short term outcome and risk factors for adverse clinical outcomes in adults with severe acute respiratory syndrome (sars) coronavirus-positive nasopharyngeal aspirate as predictor for severe acute respiratory syndrome mortality enteric involvement of severe acute respiratory syndrome-associated coronavirus infection quantitative analysis and prognostic implication of sars coronavirus rna in the plasma and serum of patients with severe acute respiratory syndrome prognostic significance of the radiographic pattern of disease in patients with severe acute respiratory syndrome acute respiratory syndrome in critically-ill patients with severe acute respiratory syndrome short-term outcome of critically ill patients with severe acute respiratory syndrome update -sars case fatality ratio epidemiological determinants of spread of causal agent of severe acute respiratory syndrome in hong kong cardiac arrest in severe acute respiratory syndrome: analysis of cases effectiveness of precautions against droplets and contact in prevention of nosocomial transmission of severe acute respiratory syndrome (sars) sars: ventilatory and intensive care a practical approach to airway management in patients with sars infection control precautions in the icu effective staff education in infection control, emphasizing on • precautions to be used in high-risk procedures and alternative procedures to reduce risks • limit opportunities for exposure, e.g., avoid aerosol generating procedures & limit number of health care workers (hcws) present, alternative nursing practices to limit number of hcws exposed to each patient • effective use of time during patient contact • "gowning" and "degowning" without contamination precautions: disposable gloves, gown, cap • eye protection with non-reusable goggles and face-shield • powered air purification respirators (papr) are optional ppe when performing high-risk procedures • pens, paper, personal items and medical records should where feasible, increase to ≥ ach + re-circulate air through hepa filter • preferred: negative pressure isolation rooms with antechambers • a viral/bacterial filter should be placed at the expiratory port of bag-valve mask • place two filters per ventilator: between expiratory port and the ventilator, and another on the exhalation outlet of the ventilator • use closed-system in-line suctioning for endotracheal/tracheostomy tubes • handle contaminated heat and moisture exchangers (hme) and heated humidifiers carefully • scavenger system for exhalation port of ventilator is optional if negative pressure with high air change (> /hour) is achieved • preoxygenate patient and temporarily switch off machine whenever ventilator circuit disconnection is required the authors have declared that no conflict of interest exists. key: cord- - noy z authors: desai, aakash; kulkarni, amit; rajkumar, s vincent; gyawali, bishal title: clinical trial endpoints in severe covid- date: - - journal: mayo clin proc doi: . /j.mayocp. . . sha: doc_id: cord_uid: noy z nan since the first outbreak in wuhan, china in late december , the coronavirus disease (covid- ) pandemic caused by severe acute respiratory syndrome coronavirus- (sars-cov- ) has already tolled , deaths. although most of the patients are asymptomatic or have minimal symptoms, patients with severe covid often require hospitalization [ ] . mortality in patients admitted to intensive care units (icu) or those requiring mechanical ventilation is as high as % [ , ] . there is an urgent need to develop effective therapy for severe covid that improves mortality. however, clinical trials of agents tested for severe covid may not necessarily test for mortality outcomes as the primary endpoint, as was highlighted in the press release of the recent remdesivir trial. since drugs improving mortality in severe covid- is the most important endpoint to achieve both from clinical and public policy standpoint, we evaluated the type of primary endpoints currently being assessed in randomized controlled trials (rcts) in severe covid . we searched www.clinicaltrials.gov to identify clinical trials for "severe covid " as of april th, . we included only phase iii and interventional trials. we excluded prophylaxis, prevention, and treatment trials with no mention of "severe covid" in the title. we then extracted information on the primary endpoint of each eligible study. two authors (a.d and a.k) independently performed the data extraction and analysis. of the trials identified initially, trials (with study drugs) satisfied our inclusion and exclusion criteria and were included in the final analysis. of these, / ( . %) trials were randomized studies and / ( . %) of these were placebo-controlled trials. the most common study drugs in the intervention arm were hydroxychloroquine alone ( / , ( . %)), followed by remdesivir and methylprednisolone ( / , ( . %)) ( table ) . among the trials, we found different primary endpoints. all cause hospital mortality / ( %), change in pao to fio ratio / ( . %), and composite primary endpoints that included mortality and time to clinical improvement / ( . %) were the most common primary endpoints studied. (figure ). our analysis found that only / ( %) ongoing phase iii rct in severe covid have mortality as the standalone primary endpoint or a part of composite endpoint. this finding is surprising given the urgent need for effective drugs that can alter the natural history of severe covid and reduce mortality. most of the other primary endpoints being assessed are unproven surrogate clinical endpoints or biomarker-based endpoints. some surrogate clinical endpoints like length of time to clinical improvement ( . %), hospital stay ( . %) may offer meaningful information from a cost of care or resource perspective but these studies are not powered to detect mortality benefit. given that mortality is as high as % in severe covid who need icu care and average time to death is - days [ , ] , the number of mortality events needed and follow-up time do not pose an impediment for analysis of mortality as the primary endpoint. furthermore, as the pandemic subsides, we may lose valuable time to enroll patients into well-designed rcts to obtain definitive answers for treatment efficacy. although our search results may not be comprehensive given the non-uniformity in the definition of severe covid among all trials, our findings do raise serious concerns about the use of surrogate measures, in current and ongoing clinical trials for severe covid . any endpoint other than mortality reduction may not be relevant at the time of a pandemic with a virus that has high mortality rates. we must prioritize discovery of a drug that truly reduces mortality rather than squandering resources in finding a drug that may make us complacent but not improve any meaningful outcomes. clinical characteristics of coronavirus disease in china presenting characteristics, comorbidities, and outcomes among patients hospitalized with covid- in the new york city area baseline characteristics and outcomes of patients infected with sars-cov- admitted to icus of the lombardy region, italy key: cord- -zp aqpm authors: harrison, andrew g.; lin, tao; wang, penghua title: mechanisms of sars-cov- transmission and pathogenesis date: - - journal: trends immunol doi: . /j.it. . . sha: doc_id: cord_uid: zp aqpm the emergence of sars-coronavirus (sars-cov- ) marks the third highly pathogenic coronavirus to spill over into the human population. sars-cov- is highly transmissible with a broad tissue tropism that is likely perpetuating the pandemic. however, important questions remain regarding its transmissibility and pathogenesis. in this review, we summarize current sars-cov- research, with an emphasis on transmission, tissue tropism, viral pathogenesis, and immune antagonism. we further present advances in animal models that are important for understanding the pathogenesis of sars-cov- , vaccine development, and therapeutic testing. when necessary, comparisons are made from studies with sars to provide further perspectives on covid- , as well as draw inferences for future investigations. infection. first, viral entry may heavily depend on the expression of tmprss , as nearly undetectable amounts of ace still support sars-cov entry, so long as tmprss is present [ ] . second, the mrna expression of cellular genes such as escrt (endosomal sorting complex required for transport) machinery gene members (including chmp , chmp , chmp a, and vps b) related to pro-sars-cov- lifecycle is higher in a small population of human type ii alveolar cells with abundant ace , relative to ace -deficient cells [ ] . this suggests that sars-cov- hijacks a small population of type ii alveolar cells with high expression of ace and other pro-viral genes for its productive replication. third, the lungas the main tropism of sars-covs-may be contingent on the regulation of ace at the transcriptional and protein levels [ , , ] [ , ] . for example, in human airway epithelial cells, ace gene expression is upregulated by type i and ii interferons [ , ] during viral infection. lastly, compared to other sars-covs, sars-cov- spike contains a unique insertion of rrar at the s /s cleavage site [ ] [ ] . this site can be pre-cleaved by furin, thus reducing the dependence of sars-cov- on target cell proteases (tmprss /cathepsin l) for entry [ ] [ ] and potentially extending its cellular tropism, given that proteolytically active furin is abundantly expressed in human bronchial epithelial cells [ , ] . one of the distinctions between sars-cov and sars-cov- is the latter's ability to efficiently infect the upper respiratory tract (urt), such as nasopharyngeal (np) and/or oropharyngeal (op) tissues, possibly due to its higher affinity for ace , which is expressed in human nasal and oral tissues [ ] [ ] [ , , ] . the readily detectable titers of sars-cov- in the urt mucus of covid- patients during prodromal periods might contribute to explaining the more rapid and effective transmissibility of sars-cov- relative to sars-cov [ ] . human covs often cause enteric infections, with variable degrees of pathogenicity [ ] . indeed, ace and tmprss are abundantly expressed within the human and many other mammalian intestinal tracts, specifically the brush border of intestinal enterocytes [ , , , ] . accordingly, gastrointestinal illness has been frequently reported in covid- patients [ , ] , consistent with the recovery of sars-cov from sars patients' stool samples [ ] , suggesting a potential fecal-oral route of transmission for these two covs. of note, ~ % of covid- patients examined have had detectable sars-cov- rna in feces, even after respiratory symptoms subsided, suggesting that sars-cov- titers might be prolonged in the intestinal tract [ ] . although further testing is warranted, these data suggest the possibility that fecal-oral transmission of sars-cov- might occur. evidently, robust epidemiological studies are needed to conclusively demonstrate if covid- patients recovering from respiratory illness are able to spread sars-cov- . human covs are transmitted primarily through respiratory droplets, but aerosol, direct contact with contaminated surfaces, and fecal-oral transmission were also reported during the sars epidemic [ ] [ ] [ ] . early reports of patients with cough, lung ground glass opacities, and symptom progression to severe pneumonia, suggested communicability of sars-cov- via the respiratory route (figure ) [ ] [ ] [ ] . direct transmission by respiratory droplets is reinforced by productive sars-cov- replication in both the urt and lrt, and the increasing number of reports indicating human-to-human spread among close contacts exhibiting active coughing ( figure ) [ , [ ] [ ] [ ] . so far, the basic reproduction number (r ) is ~ . , based on early case j o u r n a l p r e -p r o o f journal pre-proof tracking in the beginning of the pandemic, with a doubling time of days [ ] [ ] . furthermore, there is now evidence for non-symptomatic/pre-symptomatic spread of sars-cov- , which is in contrast to the transmission dynamics of sars-cov [ ] . this finding underscores the ability of sars-cov- to colonize and replicate in the throat during early infection [ , , ] . based on these apparent disparities in virus transmission, one study modeled the transmission dynamics of sars-cov- in pre-symptomatic individuals, and indicated that the pre-symptomatic r has approached the threshold for sustaining an outbreak on its own (r > ); by contrast, the corresponding estimates for sars-cov were approximately zero [ ] . similarly, asymptomatic spread of sars-cov- has been documented throughout the course of the pandemic [ ] [ , [ ] [ ] [ ] [ ] . understanding the relative importance of cryptic transmission to the current covid- pandemic is essential for public health authorities to make the most comprehensive and effective disease control measures that include mask-wearing, contact tracing, and physical isolation. for sars-cov- , various modes of transmission have been proposed, including aerosol, surface contamination, fecal-oral route, representing confounding factors in the current covid- pandemic; thus, their relative importance is still being investigated (figure ) [ ] . aerosol transmission (spread > m) was implicated in the amoy gardens outbreak during the sars epidemic, but the inconsistency of these findings in other settings suggested that sars-cov was likely an opportunistic airborne infection [ , ] . similarly, no infectious sars-cov- virions have been isolated, though viral rna was detectable in the air of covid- hospital wards [ ] . generation of experimental aerosols carrying sars-cov- (comparable to those that might be generated by humans) have offered the plausibility of airborne transmission, but the aerodynamic characteristics of sars-cov- during a natural course of infection is still an area of intense inquiry [ ] . nonetheless, deposition of virus-laden aerosols might contaminate j o u r n a l p r e -p r o o f objects (e.g. fomites) and contribute to human transmission events [ , ] . finally, fecal-oral transmission has also been considered as a potential route of human spread, but this route remains an enigma despite evidence of rna-laden aerosols being found nearby toilet bowls, along with detectable sars-cov- rna in rectal swabs during the precursor epidemic of covid- in china [ , , ] . in general, common cold covs tend to cause mild urt symptoms and occasional gastrointestinal involvement (figure ) . on the contrary, infection with the highly pathogenic covs, including sars-cov- , causes severe 'flu'-like symptoms that can progress to acute respiratory distress (ards), pneumonia, renal failure, and death [ , , , ] . the most common symptoms are fever, cough and dyspnea, accounting for %, % and % of covid- patients (n= ), respectively in one epidemiological study [ ] . the incubation period in covid- is rapid, ~ - days, versus - days in sars-cov infections [ , , ] . as the pandemic is progressing, it has become increasingly clear that covid- encompasses not only rapid respiratory/gastrointestinal illnesses, but can have long-term ramifications such as myocardial inflammation [ ] . furthermore, severe covid- is not restricted to the aged population as initially reported; children and young adults are also at risk [ ] . from a diagnostic perspective, covid- presents with certain 'hallmark' laboratory and radiological indices, which can be helpful in assessing disease progression ( table ) it is widely accepted that the aging process predisposes individuals to certain infectious diseases [ ] . in the case of covid- , older age is associated with greater covid- morbidity, admittance to the icu, progressing to ards, higher fevers and greater mortality rates [ ] [ , ] . moreover, lymphocytopenia, neutrophilia, elevated inflammation-related indices, and coagulation-related indicators have been consistently reported in older (≥ years old) relative to young and middle-aged covid- patients [ table ;( [ , ] ) [ , , , , ] . at the cellular level, a lower capacity of cd + and cd + t-cells to produce ifn-γ and il- , as well as an impairment in t-cell activation from dendritic cells (dcs) in acute covid- patients (≥ years old), might potentially compromise an optimal adaptive immune response [ ] . based on examples from mice, a productive cd + t-cell response relies heavily on lung resident dcs (rdcs) and abates sars-cov infection [ , ] . however, whether a reduction in the dc population in the lungs of older, more severe patients causes sub-optimal t-cell activation during sars-cov- infection remains to be robustly investigated. higher proportions of proinflammatory macrophages and neutrophils have also been observed in the bronchoalveolar lavage fluid (balf) of covid- patients with severe symptoms compared with those exhibiting mild symptoms (key figure, figure ) [ ] . accordingly, proinflammatory cytokines (e.g. il- , il- ) are elevated in the balf of severe j o u r n a l p r e -p r o o f journal pre-proof covid- patients, along with higher expression of inflammatory chemokines (e.g. ccl ) in macrophages relative to non-severe covid- patients [ ] [ ] [ ] [ ] . indeed, similar inflammatory milieux have been associated with severe lung pathology in sars patients, along with the notable 'cytokine storm' that can present in critically ill covid- patients [ , ] [ , [ ] [ ] [ ] . these proinflammatory mediators can, in turn, perpetuate lung disease by elevating creactive protein (crp) from the liver ( table ) products [ ] . specifically, covs can avoid immune sensing via i) the formation of dmvs that sequester viral nucleic acid from being recognized by prrs and ii) direct ablation of the functionality of immune signaling molecules by viral proteins [ , ] . the structural and functional conservation of these proteins across the betacoronavirus genus and in nsps between sars-cov and sars-cov- , suggests that some of these suppressive mechanisms might be employed by sars-cov- (see below) [ ] . indeed, patients with severe covid- have reported an imbalanced immune response with high concentrations of inflammatory cytokines/chemokines, but little circulating ifn-β or ifn-λ, resulting in persistent viremia [ ] . of note, among several respiratory viruses tested, sars-cov- has demonstrated to most potently suppress type i and type iii ifn expression in both human bronchial epithelial cells and ferrets [ ] . thus, evasion of ifn signaling by sars-cov- and impaired ifn production in j o u r n a l p r e -p r o o f human peripheral blood immune cells might contribute to the productive viral replication, transmission, and severe pathogenesis during covid- , although further testing is warranted to fully dissect these putative evasion pathways [ ] . with regard to functional conservation of viral proteins, sars-cov and mers-cov nsps and accessory proteins circumvent viral rna-sensing pathways at multiple stages (e.g. rig-i, mda- ) through proteasomal degradation and/or prevention of protein activation ( figure ) [ ] . functional conservation between sars-cov and mers-cov pl pro (encoded by nsp ) proteins has been reported, where these proteins target the initial prr signaling cascade at multiple levels of the pathway includingbut not limited to-rig-i, mavs, tbk , irf and nf-k b ( figure ) [ ] [ ] [ ] . the sars-cov pl pro also targets the dna-sensing pathway at sting ( figure ) ; antagonizing this pathway might be important as mitochondrial stress during dengue virus infection triggers ifn-β production that is dependent on sting activation [ , ] . recent evidence suggests the sars-cov- pl pro might also inhibits ifn-i expression in human kidney epithelial cells, yet the mechanisms remain to be defined [ ] . moreover, nsp of highly pathogenic hcovs, including sars-cov and mers-cov displays a pleiotropic effect, targeting several components of ifn-i signaling ( figure ) [ , ] . this potent suppressive function of nsp also appears to be maintained in sars-cov- , primarily through shutdown of translational machinery and prevention of immune gene expression [ , , ] . furthermore, because there are only five accessory genes in the mers-cov genome compared to eight and seven in the sars-cov and sars-cov- genomes, respectively, similar immunosuppressive mechanisms may exist but appear to be mediated via different proteins [ , ] . for example, sars-covs orf can inhibit irf activation and stat nuclear translocation, whereas this same effect is obtained by orf a/b and orf of mers-cov j o u r n a l p r e -p r o o f ( figure ) [ , ] . coincidently, the apparent loss of these proteins may provide evidence for why mers-cov is more sensitive to ifn treatment than sars-covs in primary and continuous cells of the human airways [ ] . the sars-cov- proteins appear to have stronger inhibitory effects than their counterparts of highly pathogenic sars-and mers-cov [ ] . in light of these findings, sars-cov- has replicated more efficiently than sars-cov in ex vivo human lung explants, possibly through the greater suppression of ifn-i/iii cytokines [ ] ; further work given that sars-cov- uses the same ace entry receptor as sars-cov, rapidly deploying mouse models for pathogenesis studies were well underway within weeks of the pandemic's inception. however, various impediments remain for sars-cov- in productively infecting mice in these models, as it is unable to bind mouse ace (mace ) [ ] . to overcome these prerequisites, several mouse models have been developed that recapitulate certain components of j o u r n a l p r e -p r o o f human covid- . one of these strategies is to genetically modify mice to express human ace (hace ) (humanized mice) under the epithelial cell-specific cytokeratin- (krt ) promoter [ ] , a universal chicken beta-actin promoter [ ] , or the endogenous mace promoter [ ] . all these mice are susceptible to sars-cov- infection, but phenotypic disease varies because of differential hace tissue expression [ ] [ ] [ ] . for instance, krt -hace and betaactin-hace -transgenic mice rapidly succumb to sars-cov- infection with lung infiltration of inflammatory immune cells inducing severe pulmonary disease, accompanied by evident thrombosis and anosmia, which partially recapitulate human covid- [ ] [ ] . as the onset of severe histopathological changes occurs days after peak virus infection, these models recapture the delayed morbidity seen in covid- patients as a result of inflammatory cell infiltration [ ] . therefore, employing humanized mouse models of severe sars-cov- infection might be useful for testing the efficacy of antiviral drugs, vaccines, and immune therapeutics that ablate hyperinflammation [ ] . however, the broad expression of hace in these models significantly expands sars-cov- tissue tropisms and might alter its pathogenic mechanisms [ ] [ ] . for example, both sars-cov and sars-cov- infection lead to encephalitis in these mouse models, which is not common in covid- patients [ , , ] . considering the fact that the majority of human sars-cov- infections are asymptomatic or mild, mice originally bearing mace that is replaced by hace may be more appropriate for assessing pathogenesis and tissue tropism [ ] . this model develops mild lung pathology, with sars-cov- infection being restricted to the lung and intestine [ ] . in addition to the transgenic modification, mice can also be sensitized to sars-cov- infection via transient transduction of adenovirus (ad )-or adeno-associated virus (aav)-expressing hace in respiratory tissues, akin to the approach previously used for mers-cov infection [ ] [ ] [ ] . these mice develop viral pneumonia, weight loss, severe pulmonary pathology, and a high viral load in the lung, consistent with human covid- [ ] . this approach might be quickly adapted to many genetically modified mouse strains that might provide mechanisms of sars-cov- pathogenesis and protective immune responses. this model is limited, however, by the transient ectopic expression of hace from the ad /aav vector that can induce mild bronchial inflammation and expand cell tropism of sars-cov- and thus, presumably alter disease pathogenesis [ ] . rather than genetic modification in host animals, viruses can also be genetically modified and be used in model animals [ , ] . for instance, in one study, serial passaging of sars-cov- in mice led to enrichment of a n y viral mutant that elicited interstitial pneumonia and inflammatory responses in both aged and young wild-type balb/c mice [ ] . another mouseadapted sars-cov- strain (ma ) carrying three mutations in the rbd of spike protein caused severe lung pathology and ards in mice, characteristic of severe covid- [ ] . despite the three mutations in the rbd of the mouse-adapted spike, vaccination with full length sars-cov- spike elicited robust neutralizing antibody titers and complete protection against a secondary challenge with ma [ ] ; these findings suggest that this strain may be applicable to pathogenesis studies, as well as antiviral drug and vaccine testing in rodents. the role of non-human primates (nhp) in evaluating coronavirus pathogenesis cannot be understated. depending on the nhp model utilized, clinical signs/symptoms may be mild or absent entirely [ ] [ ] [ ] . in rhesus macaques, several studies have noted reduced appetite, j o u r n a l p r e -p r o o f transient fevers ( day post infection: dpi) and mild weight loss without overt signs of respiratory distress or mortality [ ] [ ] [ ] . by contrast, cynomolgus macaques did not display any observational signs of disease in another study [ ] . although certain nhps appear to only mimic mild disease (if any), rhesus macaques have exhibited high viral loads in nasal swabs, throat samples, and balf early post inoculation, and viral rna was still measurable by qpcr in the trachea and lung on day p.i., highlighting the apparent tropism of sars-cov- for the urt and lingering viral nucleic acid in respiratory tissues after resolution of disease [ , ] . sars-cov- has also been detected in nasal swabs at dpi in nhps, consistent with the prolonged urt shedding of virus in covid- patients at ~ dpi [ , , , ] . the tropism of sars-cov- for the lrt in nhps has also been recapitulated by the development of multifocal lesions and interstitial pneumonia, supporting the hypothesis that lung injury is driven by increased infiltration of neutrophils and macrophages into the lung following viral infection such as thickened alveolar septum and diffuse severe interstitial pneumonia when compared to young macaques ( - years old) [ ] . therefore, these studies highlight the importance of also considering the age factor, as an additional variable, when selecting animal models that might closely, or accurately, recapitulate human disease. evaluating efficacious vaccine candidates in nhps will also be important for understanding correlates of protection against sars-cov- . accordingly, reports of antibodydependent enhancement, as well as of non-neutralizing humoral responses to the conserved regions of sars-cov- , raise concerns on our future ability to effectively administer an immunogen without inducing immunopathology [ , ] . furthermore, upon viral challenge, lymphocytes have expanded in rhesus macaque models around dpi with complementary b-cell responses against sars-cov- spike appearing - dpi in blood samples [ ] ; expansion of these adaptive immune compartments was analogous to those observed in covid- patients [ , , [ ] [ ] [ ] . subsequent re-challenged rhesus macaque have presented a rapid anamnestic immune response characterized by significantly higher neutralizing antibody (nab) titers than the primary infection macaques [ ] . thus, protective efficacy seems to depend primarily on nab titers, at least in nhps, and so far, t-cell numbers have not substantially increased following re-challenge in the serum of these animals, and in a secondary study, cd + and cd + cytokine (e.g. ifn-γ) responses did not correlate with immune protection from dna vaccines with different components of the sars-cov- spike protein [ ] [ ] . although these animals have failed to manifest overt signs of infection and respiratory compromise, nhps still represent the 'gold standard' for evaluating the protective efficacy of human-bound sars-cov- vaccines based on parallels to humans in terms of viral tropism, immunopathology, and correlates of protection [ ] . further research is urgently needed to j o u r n a l p r e -p r o o f explore the durability of immune responses to sars-cov- , considering reports of waning immunity to other covs and the detection of pre-existing cross-reactive 'common-cold' cov tcells with sars-cov- in naïve humans (see outstanding questions) [ , ] . the emergence of sars-cov- as the most recent example of zoonotic virus spillovers into in the last hours leading up to death, all patients which were included for this metric had a prothrombin time of > . s. aerosol: suspension of fine solid or liquid droplets in the air (or a gas medium), such as dusts, mists, or fumes. anamnestic immune response: memory immune response to a previously encountered antigen. angiotensin-converting enzyme (ace ): cell surface enzyme of endothelial, epithelial, and other cells, with a well-defined function in maintaining normal blood pressure. anosmia: partial or complete loss of the sense of smell. antibody-dependent enhancement: phenomenon by which antibodies against a virus are suboptimal to the virus and enhance its entry into host cells. convalescence period: the time of gradual recovery after an illness or injury. correlates of protection: quantifiable parameters such as antibodies, indicating that a host is protected against microbial infection. cytokine storm: severe immune reaction in which the body releases too many cytokines into the blood too quickly. d-dimer: fibrin degradation product in the blood after a clot is degraded by fibrinolysis. disseminated intravascular coagulation (dic): condition in which blood clots form throughout the body and block small blood vessels, leading to multiorgan failure. fecal-oral transmission: route of disease transmission by which an infectious agent in fecal materials is passed to the mouth of another. fomite: inanimate object (clothes, utensil, and furniture etc.) that, when contaminated with an infectious agent, can transfer the infectious agent to a new host. furin: proprotein convertase that cleaves a precursor protein into a biologically active state. incubation period: timeframe elapsed between when a host is first exposed to an infectious agent and when signs or symptoms begin to appear. lung ground glass opacity: nonspecific radiological description of an area of increased opacity in the lung through which vessels and bronchial structures are still visible. neutralizing antibody (nab): an antibody that binds a pathogen with high affinity and prevents the latter from exerting its biological effect. neutrophil extracellular traps (nets): networks of extracellular fibers, primarily composed of dna from neutrophils due to chromatin decondensation, which can 'trap' extracellular pathogens. pattern recognition receptor: germline-encoded host sensor that recognizes a signature pattern in microbial molecules. prodromal period: the time immediately following the incubation period of a microbial infection in which a host begins to experience symptoms or changes in behavior/functioning. prothrombin time: measurement of the extrinsic pathway of coagulation. r (reproductive number): the expected number of new disease cases generated by one case. an r > indicates the outbreak will expand; r < the outbreak will die out. respiratory droplet: small aqueous droplet produced by exhalation, consisting of saliva or mucus and other matter derived from respiratory tract surfaces. zoonotic disease: infectious disease caused by a pathogen that has crossed a species barrier from animals to humans. the ongoing covid- pandemic has resulted in numerous accounts of different transmission routes between humans. droplet transmission (> μm) is the most pronounced and heavily implicated mode of transmission reported during the pandemic. direct contact spread from one infected individual to a second, naïve person has also been considered a driver of human-to-human transmission, especially in households with close interactions between family members. the contagiousness of sars-cov- after disposition on fomites (e.g. door handle) is still under investigation, but is likely a compounding factor for transmission events, albeit less frequently than droplet or contact-driven transmission. both airborne and fecal-oral human-to-human transmission events were reported in j o u r n a l p r e -p r o o f the precursor sars-cov epidemic but have yet to be observed in the current crises. solid arrows show confirmed viral transfer from one infected person to another with a declining gradient in arrow width 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coronavirus comparative replication and immune activation profiles of sars-cov- and sars-cov in human lungs: an ex vivo study with implications for the pathogenesis of covid- ribose ′-o-methylation provides a molecular signature for the distinction of self and non-self mrna dependent on the rna sensor mda the pathogenicity of sars-cov- in hace transgenic mice lethal infection of k -hace mice infected with severe acute respiratory syndrome coronavirus severe acute respiratory syndrome coronavirus infection of mice transgenic for the human angiotensin-converting enzyme virus receptor k -hace mice for studies of covid- treatments and pathogenesis including anosmia sars-cov- infection of human ace -transgenic mice causes severe lung inflammation and impaired function severe acute respiratory syndrome coronavirus infection causes neuronal death in the absence of encephalitis in mice transgenic for human ace a sars-cov- infection model in mice demonstrates protection by neutralizing antibodies rapid generation of a mouse model for middle east respiratory syndrome generation of a broadly useful model for covid- pathogenesis vaccination, and treatment adenovirus vectors for gene therapy, vaccination and cancer gene therapy a mouse-adapted sars-coronavirus causes disease and mortality in balb/c mice a mouse-adapted sars-cov- model for the evaluation of covid- medical countermeasures a sars-cov- protein interaction map reveals targets for drug repurposing a mouse-adapted sars-cov- induces acute lung injury (ali) and mortality in standard laboratory mice respiratory disease in rhesus macaques inoculated with sars-cov- comparative pathogenesis of covid- , mers, and sars in a nonhuman primate model sars-cov- infection protects against rechallenge in rhesus macaques cynomolgus macaque as an animal model for severe acute respiratory syndrome age-related rhesus macaque models of covid- immunization with sars coronavirus vaccines leads to pulmonary immunopathology on challenge with the sars virus anti-spike igg causes severe acute lung injury by skewing macrophage responses during acute sars-cov infection antibody responses to sars-cov- in patients of novel coronavirus disease antibody responses to sars-cov- in patients with covid- deep immune profiling of covid- patients reveals distinct immunotypes with therapeutic implications dna vaccine protection against sars-cov- in rhesus macaques seasonal coronavirus protective immunity is short-lasting targets of t cell responses to sars-cov- coronavirus in humans with covid- disease and unexposed individuals type-i, type-iii ifns then signal in an autocrine or paracrine manner through the janus kinase (jak )/signal transducer and activator of transcription and (stat / ) pathway, culminating in antiviral interferon-stimulated gene (isg) transcription. listed here are sars-cov (abbreviated cov), sars-cov- (abbreviated cov- ) and mers-cov (abbreviated m-cov) ifn-i antagonists, which make these viruses resistant to interferon responses. ifn-iii is also implicated in exhibiting potent antiviral effects in lung/intestinal tissues, but the underlying evasion strategies of this pathway for these viruses are currently unknown. sars-cov proteins are highlighted in blue, while functions of sars-cov- and mers-cov proteins are highlighted in red and green, respectively. question mark symbol (?) denotes sars-cov- protein bound a member of that signaling pathway in [ ], but further work is necessary to confirm its immunological mechanism. sars-cov- proteins with * denotes functional conservation with sars-cov which animal(s) serves as the natural reservoir of sars-cov- ?  does active replication of sars-cov- in the upper respiratory tract contribute to enhanced transmissibility in humans?  is intestinal sars-cov- infection a source of virus transmission?  which sars-cov- proteins antagonize innate and adaptive immune responses? do the sars-cov- proteins with more potent antagonistic immune functions increase virulence in humans compared to other hcovs?  why do some recovered patients fail to develop neutralizing antibodies?  what are the host and/or viral factors driving  what are the underlying mechanisms contributing to an inadequate ifn response to sars-cov- ?  what are the correlates of immune protection for sars-cov and will they provide sterilizing immunity?  will candidate vaccines against sars-cov also be effective in elderly subpopulations this work was supported by a national institutes of health grant r ai to p.w.the authors declare no competing financial/non-financial interest.j o u r n a l p r e -p r o o f journal pre-proof j o u r n a l p r e -p r o o f key: cord- -tah jdw authors: zhang, shen-ying; zhang, qian; casanova, jean-laurent; su, helen c. title: severe covid- in the young and healthy: monogenic inborn errors of immunity? date: - - journal: nat rev immunol doi: . /s - - - sha: doc_id: cord_uid: tah jdw severe covid- is rare in previously healthy individuals who are less than years of age, affecting probably no more than in , such infected individuals. we suggest that these patients may become critically ill because of monogenic inborn errors that disrupt protective immunity to sars-cov- . a key scientific issue for researchers studying infectious diseases is the immense clinical variability that occurs among infected individuals . similar to other human-tropic viral infections, clinical severity following infections with sars-cov- extends from asymptomatic to life-threatening, and there are various clinical forms of covid- . the data emerging from different countries suggest a global case fatality rate of ~ % in virus pcr-confirmed cases of all ages, ranging from < . % for children aged - years to > % for those aged over years. however, recent serological data suggest that the infection fatality rate may be at least an order of magnitude lower globally. there is compelling evidence for a much higher proportion of casualties in those aged over years and in those with underlying health conditions. however, a small number (~ % of all severe cases) of previously healthy adults under the age of years present with severe covid- , including severe pneumonia and, more rarely, encephalitis, cardio vascular disease and other presentations, including the recently reported kawasaki-like disease (also known as paediatric inflammatory multisystem syndrome or multisystem inflammatory syndrome in children). why do these younger, healthier patients get severe covid- ? four hypotheses could account for such severe 'idiopathic' cases . first, these patients may be infected with larger amounts of virus or a more virulent sars-cov- strain. higher virulence is unlikely, because it would lead to clusters among close contacts, through direct contagion, and there is little evidence for such clusters. a higher initial viral load appears more likely, and this hypothesis is supported by more than a century of experimental inoculations of animals with various viruses. higher inoculum levels are generally associated with more severe disease. however, it remains unknown whether the most heavily exposed humans (spouses of severe cases or health-care workers treating patients with covid- , particularly those with insufficient protection) account for a large proportion of severe idiopathic cases. these people clearly have a much higher risk of infection, and probably of disease, but may not be at higher risk of severe disease if they are young and healthy. second, particular environmental conditions, such as the presence or absence of an element in the air inhaled by the patient, may aggravate the infection. the finger has recently been pointed at global pollution, but clinical heterogeneity exists in both polluted and unpolluted areas. season or climate may also affect the infection process and immunity. nevertheless, environmental differences may account for epidemic differences between the northern and southern hemispheres, for example, between ecuador and iceland, but are less likely to account for differences within single countries or over smaller scales (for example, within the confines of a single city, cruise ship or household). third, an inevitable 'somatic transformation' of cells occurs in individual human hosts. such genetic and epigenetic processes are responsible, for example, for the steady increase in the incidence of shingles after the age of years. acquired covert illnesses may weaken some individuals prematurely, or the absence of an acquired, protective process may be detrimental. pulmonary cells and leukocytes may be affected in different ways. an agnostic attitude is therefore essential. prior infectious history may also be a somatic determinant of disease severity, through the accumulation of immunological memory via the t and b lymphocytes governing adaptive immunity. for example, previous infection with another coronavirus, such as epidemic sars-cov or endemic hcov- e, might be protective. alternatively, previous infections with related viruses may be deleterious, as reported for dengue, which is typically silent during the first infection and severe during subsequent infections with different serotypes, owing to antibody-dependent enhancement . detailed serological studies are required to investigate this aspect of covid- . severe covid- in the young and healthy: monogenic inborn errors of immunity? shen-ying zhang , , ✉ , qian zhang , , ✉ , jean-laurent casanova , , , , ✉ , helen c. su and the covid team* severe covid- is rare in previously healthy individuals who are less than years of age, affecting probably no more than in , such infected individuals. we suggest that these patients may become critically ill because of monogenic inborn errors that disrupt protective immunity to sars-cov- . finally, severe covid- in previously healthy children and adults may result from monogenic predisposition. this hypothesis is supported by the rarity of severe cases among such individuals during primary infection with sars-cov- . studies since have identified a number of monogenic inborn errors of immunity (ieis) underlying life-threatening infectious diseases, including specific viral diseases, in previously healthy patients , - . two of these ieis are classic mendelian disorders (monogenic with complete penetrance) underlying viral illnesses, including familial disease. in autosomal recessive epidermodysplasia verruciformis, mutations in tmc , tmc or cib result in high susceptibility to skin-tropic beta human papillomaviruses (hpvs). meanwhile, increased susceptibility to epstein-barr virus is seen in patients with x-linked lymphoproliferative syndrome (caused by mutations in the x chromosome genes sh d a and xiap) and in patients with biallelic mutations in cd , cd , itk or magt . moreover, since , other monogenic ieis have been shown to underlie sporadic (as opposed to familial) viral diseases, with incomplete penetrance (that is, non-mendelian). the first example is herpes simplex virus (hsv- ) encephalitis, attributable in about - % of cases to mutations affecting the tlr or snorna pathways (forebrain infection) or dbr (brainstem infection). another example is influenza a virus (iav) pneumonia caused by mutations in tlr , irf or irf that impair interferon immunity. other ieis underlying severe viral diseases, with incomplete or unknown penet rance, have since been reported , , . the dis coveries of these ieis demonstrated that severe disease due to primary infection with a common virus that is benign in the general population can result from a monogenic 'hole' in human immunity. in patients with these monogenic ieis, disease immuno pathogenesis can involve an impairment of immune mechanisms specific to the virus (for example, cib mutations affect control of hpv in keratinocytes) or specific to the tissue site (for example, dbr mutations affect immunity to various viruses in the brainstem). molecularly, these disorders disrupt antiviral immunity through known (for instance, type i and type iii interferon pathways in iav or human rhinovirus pneumonia) or unknown (for example, snorna deficiency in hsv- encephalitis) mechanisms. they may result in an insufficient immune response (for example, patients with ifnar deficiency can develop severe adverse reactions to live attenuated vaccines) or an excessive immune response (for example, patients with il- bp deficiency develop excessive inflammation and fulminant viral hepatitis). at the cellular level, ieis can affect leukocytes or other tissue-resident cells (for example, ieis in pulmonary epithelial cells affect the control of iav in the lung). studies of inborn errors of non-haematopoietic cell-intrinsic immunity have suggested that keratinocytes, pulmonary epithelial cells and cortical neurons are essential for tissue-specific protective immunity to various viruses, scaling immunity to infections up from the immune system to the whole organism . these different immunological scenarios may also underlie severe covid- in patients with monogenic disorders. pneumonia, encephalitis and kawasaki-like disease may be caused by different types of disorders. the search for monogenic ieis conferring predisposition to severe covid- in previously healthy children and young or even middle-aged adults should therefore involve the genome-wide, agnostic testing of genetic hypotheses (see also covid human genetic effort) . there may be autosomal and x-linked disorders, and recessive, dominant or co-dominant traits. genetic variants may be loss of function or gain of function. there may be both genetic and physiological homogeneity and heterogeneity. clinical penetrance may be complete or incomplete, and incomplete penetrance may suggest digenic or even oligogenic disorders. the discovery of monogenic ieis to sars-cov- should help unravel the mechanistic basis of the immunopathogenesis of severe covid- in young, previously healthy individuals. monogenic disorders can provide a basis for genetic diagnosis and counselling, while paving the way for preventive and therapeutic interventions. they also provide mechanistic hypotheses that can be tested in other patients who are critically ill who are older or have comorbidities. the human genetic determinism of life-threatening infectious diseases: genetic heterogeneity and physiological homogeneity? igg antibodies to dengue enhanced for fcγriiia binding determine disease severity human inborn errors of immunity to infection affecting cells other than leukocytes: from the immune system to the whole organism genomics is rapidly advancing precision medicine for immunological disorders human inborn errors of immunity: update of the iuis phenotypical classification human inborn errors of immunity: update on the classification from the international union of immunological societies expert committee new immunodeficiency syndromes that help us understand the ifn-mediated antiviral immune response human inborn errors of immunity to herpes viruses shared and distinct functions of type i and type iii interferons & covid human genetic effort. a global effort to define the human genetics of protective immunity to sars-cov- infection the authors contributed equally to all aspects of the article. the authors declare no competing interests. covid human genetic effort: https://www.covidhge.com key: cord- -vuuxthx authors: deng, ming; qi, yongjian; deng, liping; wang, huawei; xu, yancheng; li, zhen; meng, zhe; tang, jun; dai, zhe title: obesity as a potential predictor of disease severity in young covid‐ patients: a retrospective study date: - - journal: obesity (silver spring) doi: . /oby. sha: doc_id: cord_uid: vuuxthx objective: to explore the indicators for severity in young covid‐ patients age between to . methods: this retrospective cohort study includes consecutively admitted covid‐ patients age between to in zhongnan hospital of wuhan university. among them, were moderate cases, were severe or critical cases. epidemiological, clinical and laboratory characteristics and treatment data were collected. a multivariate logistic regression analysis was implemented to explore risk factors. results: the severe/critical cases have obviously higher bmi (average . vs. . kg/m( )) and lower liver ct value (average . vs. . mu) than moderate cases group. the severe/critical cases have higher fasting glucose, alanine aminotransferase (alt) , aspartate aminotransferase (ast) , and creatinine (cr) compared with moderate cases (all p< . ) . more severe/critical cases ( . % vs. . %) have positive urine protein. the severe/critical cases will experience a significant process of serum albumin decline. logistic regression analysis showed that male, high body mass index (especially obesity), elevated fasting blood glucose and urinary protein positive are all risk factors for severe young covid‐ patients. conclusion: obesity is an important predictor of severity in young covid‐ patients. the main mechanism is related to the damage of liver and kidney. the institutional ethics board of zhongnan hospital of wuhan university (no. ). a total of patients were included in the analysis. all cases were classified as moderate, severe, or critical according to the guidance for corona virus disease ( th edition) released by the national health commission of china [ ] . moderate cases have fever and respiratory tract-related symptoms as well as visible signs of pneumonia on imaging. cases are classified as severe if they meet any of the following criteria: ( ) shortness of breath, according to a respiratory rate (rr) ≥ times/min; ( ) in the resting state, an oxygen saturation ≤ %; ( ) arterial blood oxygen partial pressure (pao )/oxygen concentration (fio ) ≤ mmhg. critical case need to meet any of the following criteria: ( ) respiratory failure occurs and mechanical ventilation is required; ( ) shock occurs; ( ) organ failure in addition to respiratory distress requires intensive care unit (icu) monitoring and treatment. the need for written informed consent was waived owing to the rapid emergence of this infectious disease. clinical data were collected up to march , , and data were collected for one critically ill patient until april , after transfer to leishenshan hospital. we use a standard case report form for data collection. past medical history, family history, anthropometric data, laboratory test results, treatment details, and oxygen support data during hospital admission were collected from the patients' medical records by two independent researchers. if any key data were this article is protected by copyright. all rights reserved missing, we contacted the physician in charge of the patient's care to request further information. data for metabolic indexes (i.e., fasting blood glucose [ the presence of the sars-cov- virus in respiratory samples was confirmed with real-time reverse transcription polymerase chain reaction (rt-pcr) using a nucleic acid detection kit developed by daan gene co., ltd. of sun-yatsen university. this kit is designed to qualitatively detect the orf ab and n genes of the sars-cov- virus. in this study, the intelligent evaluation system (hangzhou yitu healthcare technology co., ltd.) was employed as the ct image analysis tool for the detection of covid- -related pneumonia. the system was used for segmentation of the left and right lungs and detection of patchy shadows, and from the collected images, quantitative parameters including the inflammatory volume and proportion of the inflammatory volume among the total lung volume were computed. this article is protected by copyright. all rights reserved axial ct images were reconstructed with slice thicknesses of mm and mm. the thickness of subcutaneous fat was measured on the axial images under the midline of the anterior abdominal wall, and the thickness in the largest transverse view was taken as the final measurement. the ct value of the liver was determined by selecting the left lateral lobe, the left medial lobe, and the right hepatic lobe on the transverse images of the th thoracic vertebra using a digital imaging and communications in medicine (dicom) viewer. during the measurement, the area of interest (roi) as positioned as far away from the blood vessels as possible. the averages of the three measurements was used as the ct value of the liver. in addition, the evaluation criteria for fatty liver was a liver density lower than that of the spleen (< hounsfield units [hu]) [ ] . ct density of epicardial fat and visceral fat were measured by standard methods. we assessed the volume of epicardial fat volume using the original chest ct data as the basic data. the pixel from the upper edge of the outer pericardium to the heart fat was extracted layer by layer, and the volume of the fat was accumulated according to the chest ct image thickness ( figure s ). this small-sample study had unbalanced numbers of patients in the moderate and severe/critical groups (severe and critical cases were combined in one group for analysis). for descriptive statistical analysis, categorical variables were expressed as frequencies and percentages, and continuous variables were expressed as medians with quartiles (q , q ). continuous variables were also transformed into categorical variables for further analysis. differences in categorical variables between the moderate and severe/critical groups were assessed by fisher's exact test, and differences in continuous variables between the two groups were tested using the wilcoxon rank-sum test (nonparametric test). many factors presented statistical significance in the initial difference analysis. however, the sample sizes for the different conditions limited their simultaneous inclusion in multiple logistical regression. to maintain the interpretability of the multiple regression, variable selection was this article is protected by copyright. all rights reserved performed for a set of binary predictors using lasso logistic regression. multivariable logistical regression was performed by exact logistic regression and firth's logistical regression, which are standard approaches for the analysis of binary outcomes with a small sample size. exact logistic regression was selected to explore the effect of the selected variables. sex and bmi were considered as forced-in covariates, and selected variables from lasso regression were included in turn. to assess the robustness of the results, these binary predictors were replaced by their continuous types, and firth's logistical regression was performed instead of exact logistical regression. the data were analyzed using sas software (sas institute inc, cary, nc) and r version . . software (the r foundation for statistical computing, vienna, austria) with the glmnet package. two-sided p values of less than . were considered statistically significant. a total of covid- cases among young adults were included in the analysis, of which were moderate cases and were severe or critical cases. because only three cases were classified as critical, we combined the severe cases (n= ) and critical cases (n= ) for comparison with moderate cases (table ). notable findings included that all severe/critical cases were male, making the proportion of male patients in this group significantly higher than that in the moderate group ( . % vs. . %). the age distribution and proportions of patients who reported cigarette smoking and alcohol drinking were similar between the two groups. few patients in either group had a history of chronic disease or a related family history before admission. the systolic pressure and diastolic pressure were comparable between the moderate cases (average / mmhg) and severe/critical cases (average / mmhg). before hospital admission, moderate cases and severe/critical cases experienced a similar duration of symptoms. more severe/critical cases had diarrhea ( %), but the frequencies of other symptoms were similar this article is protected by copyright. all rights reserved between the two groups. evaluation of metabolic and inflammatory indicators showed that the severe/critical group had higher levels of fbg ( . vs. . mmol/l), lp(a) ( . vs. . mg/l), crp ( . vs. . mg/l), and ldh ( . vs. . u/l) and a lower level of hdl-c ( . vs. . mmol/l) compared with moderate cases ( table ). the neutrophil count also was higher in the severe/critical group than in the moderate group ( . × vs. . × /l). coagulation indexes including d-dimer and aptt were comparable between the two groups at admission (table ). indexes for dysfunction of different organs also were measured and compared between the two groups. with regard to cardiovascular functioning, the ck-mb levels of the moderate and severe/critical groups were similar ( . vs. . ng/ml). because cardiac tni and bnp data were available for only a few patients, these two indicators could not be included in the analysis. among the cases with test results, all had a normal bnp level, and only one critically ill patient who experienced cardiac arrest before hospitalization had an elevated tni concentration ( pg/ml). we also found from ct imaging that all patients had good coronary vascular conditions. from the ct images, we observed that even the patient with highest bmi had no coronary calcification similar to the normal-weight patient ( figure s ). with regard to liver functioning, the severe/critical group had higher levels of alt ( . vs. . u/l) and ast ( . vs. . u/l) than did the moderate group. because the increases in alt and ast were nearly parallel (r= . ), we considered that the elevated alt and ast levels were caused by impaired liver function. with regard to renal functioning, the severe/critical group have higher levels of cr compared with the moderate group. in addition, the frequency of urine protein positivity was greater among severe/critical cases than among moderate cases ( . % vs. . %). because immune cell counts after admission were available for only a few moderate cases and less than half of severe/critical cases, we did not include this indicator in the analysis. however, among those patients, we observed a decreased trend in t cell counts in severe/critical cases (table s ). from the evaluation of obesity among the study population, the severe/critical group had an this article is protected by copyright. all rights reserved obviously higher mean bmi than the moderate group (average . vs. . kg/m ; table ). in addition, the liver ct value was less for the severe/critical group compared with the moderate group ( . vs. . mu), which indicates more fat deposition in the liver ( table ). the epicardial fat volume is significantly higher in severe/critical group ( vs. mm ) . the subcutaneous fat thicknesses were similar between the two groups.ct density of epicardial fat and viseral fat were similar between the two groups. according to ct examination, severe/critical cases had a larger intrapulmonary lesion volume compared to moderate cases ( . % vs. . %) ( table ) . as treatment for covid- , more cases in the severe/critical group received glucocorticoids ( . % vs. . %) and albumin supplementation ( % vs. . %). more cases in the severe/critical group required oxygen support, and accordingly, the frequencies of nasal catheter use ( . % vs. . %) and extracorporeal membrane oxygenation (ecmo, % vs. . %) were higher in the severe/critical group than in the moderate group (table ) . we observed the distribution of different indicators of obesity among young adult covid- according to the different in disease severity ( figure ). according to bmi measurements, both severely and critically ill patients were distributed in the overweight/obesity interval. among them, all three critically ill patients were with obesity. according to the liver ct value, all the severely and critically ill cases were in the fatty liver interval. consistent with the results of the comparative analysis, subcutaneous fat thickness did not have a detectable effect on the severity of disease. the serum albumin level was examined twice during hospitalization for four patients with this article is protected by copyright. all rights reserved severe disease, and in these patients, the serum albumin level dropped rapidly in a short period of time ( figure ). in addition to changes in serum albumin, the d-dimer level in critically ill patients also changed significantly. interestingly, the decline in the serum albumin level in critically ill patients and the rise in the d-dimer level showed an essentially parallel inverse relationship, even when anticoagulant drugs were used (figure ). from the detailed data for the three critically patients (tables s -s ) , we observed that the changes in serum albumin levels were also parallel with the changes in the lung lesions. notably, all three of the critically ill patients experienced acute respiratory distress syndrome (ards). however, cardiovascular, liver, and kidney function did not show significant changes during the course of the disease. in critical patient , the elevated levels of tni and renal and liver function indexes at the time of admission may have been related to the occurrence of cardiac arrest before admission, and these indexes quickly recovered and stabilized after treatment. obvious changes in coagulation indexes were observed in all of the critically ill patients during the disease process. critically ill patient eventually experienced disseminated intravascular coagulation (dic). logistic regression analysis was applied to assess the potential risk factors for the severity of young covid- patients (tables and ). the analysis showed that a high body mass index (especially obesity), an elevated fbg level, an elevated ldh level, and urinary protein positivity were all risk factors for severe covid- in these young patients. in particular, urinary protein positivity could increase the risk of severe disease by more than -fold. in the analysis of relevant risk factors for severe covid- published in china before april, little was mentioned about the correlation between obesity and severe illness in patients with covid- [ ] . however,more and more updated researchs indicate that the role of obesity is very important [ ] [ ] [ ] [ ] . a study from nyu school of medicine showed that in the population this article is protected by copyright. all rights reserved under years old, the proportion of people with a bmi over kg/m who were hospitalized and admitted to the icu was more than double that among patients with a lower bmi [ ] . a recently published study from china also showed that in metabolic-associated fatty liver disease patients, obesity can increase the risk for severe covid- by about -fold [ ] . the present study provide more detailed information about the role of obesity in the severity of young covid- patients. our findings show that obesity is an important predictor of severe covid- among young patients. further analysis found that obesity mainly affects the severity of covid- through the deposition of ectopic fat in multiple organs, which in turn damages the organ function. according to our data, in young patients with severe covid- , the organs most likely to be affected were the liver and kidneys. in our study, the liver ct value in people with overweight and obesity (average mu) is similar with previous study [ ] . only one critically ill patient in this study had myocardial damage, and it was most likely caused by cardiac arrest. notably, in the present study, all of the severely or critically ill covid- patients were males, an observation which may also be related to the distribution of obesity in china. among people under the age of years, the number of men with obesity is three times greater than the number of women [ ] . although all patients in the current study had comparable albumin levels on admission, we observed a rapid decline in albumin concentration in severely and critically ill patients after admission. because the albumin level was not tested a second time in moderate cases during hospitalization, we cannot be sure that the moderate patients did not also experience a decline in albumin concentration. a study evaluating patients with covid- after weeks of hospitalization found that patients with aggravated conditions had significantly reduced albumin levels, while those with stable or improved conditions had an average albumin level of . g/l [ ] , suggesting that moderate cases are less likely to experience a significant decrease in albumin. additional studies have suggested that hypoalbuminemia on admission can help predict the progression of covid- cases to severe or critical condtiion [ ] [ ] [ ] . chronic liver disease due to synthesis disorders and chronic kidney disease due to glomerular leakage are common causes of hypoalbuminemia. even without liver and kidney dysfunction, obesity itself will be accompanied this article is protected by copyright. all rights reserved by hypoalbuminemia [ ] . but abnormal albumin function is more common reported in fatty liver disease [ ] . this study suggests that young patients with proteinuria at admission are more than times more likely to develop severe and critical illness. a renal histopathological analysis of deceased covid- patients showed that sars-cov- can directly infect and damage proximal tubular epithelial cells and podocytes [ ] . podocyte damage is usually associated with the formation of proteinuria [ ] . hypoproteinemia may be very obvious in some critically ill patients. we observed that in one critically ill patient who continued to be treated with endotracheal intubation and mechanical ventilation at this time of this reporting, normal albumin levels could only be maintained by continuous large infusions of albumin. our understanding of the functions of albumin continues to increase. albumin can not only effectively maintain plasma osmotic pressure, but also has functions in transport, anti-thrombosis, immune regulation and endothelial stability [ ] . in severe and critically ill covid- patients, low albumin levels may also be accompanied by impaired organ functioning. in this study, it was considered that the rapid onset of hypoalbuminemia may be related to the attack of the virus on the organs, because almost all young covid- patients had normal serum albumin levels at admission. pre-existing damage to liver and kidney function may cause further damage to organ functions under the attack of sars-cov- infection. in the present study, we also observed that although the difference in d-dimer between moderate and severe/critical covid- cases was not significant at admission, the level of d-dimer continued to increase in critically ill cases. some studies have suggested that d-dimer may be a risk factor for severe covid- [ , ] , and the risk of thrombosis is high for severe covid- patients [ , ]. our follow-up of three critically ill patients showed that even when anticoagulant drugs were administered, the decrease in albumin and increase in d-dimer were nearly parallel. in a critically ill patient who later died, dic eventually appeared. hypoalbuminemia has been considered a high risk factor for thrombosis [ , ] , and albumin itself also has anti-thrombotic effects due to its capacity to bind nitric oxide (no) and prolong the biologic activity of no [ ] . of course, the increased d-dimer level in critically ill covid- this article is protected by copyright. all rights reserved patients in this study could also be related to other factors, including the hypercoagulable state combined with obesity, viral infection and later combined bacterial infections, and multiple organ dysfunction. consistent with previous studies [ ] , this study also suggests that elevated fasting blood glucose levels at admission are also risk factors for critical illness in young covid- patients, although most people have no previous history of diabetes. considering that previous studies have suggested the role of inflammation in severe covid- cases [ ] , and that fat density in people with obesity may be related to inflammation of adipose tissue [ , ], we evaluated epicardial fat and visceral fat ct density in the study population, but find the difference between moderate and severe/critical group were not significant. because the sample size of this study was small, and the distribution of the two groups was uneven. this has been considered as much as possible in the analysis. two methods were used in the logistic analysis for identification of risk factors. in moderate covid- cases, the absence of multiple albumin test results during hospitalization made it impossible to accurately determine the changes in albumin levels among moderate cases. in addition, immune cell numbers were not tested for many cases, making it impossible for us to thoroughly assess the role of the immune response in young covid- patients. in addition, we need to note that chinese definition of overweight and obesity is different from the united states and europe [ ] . therefore, when applying the conclusions of this study, it is necessary to consider the possible impact of this difference.our research suggests that the liver ct value is helpful for predicting severe cases of young covid- . however, we should note that liver ct value is an imaging index, which can not reflect pathophysiological changes. therefore, we also need further research to help clarify low liver ct value associate with severe covid- cases. this article is protected by copyright. all rights reserved the present study investigated predictive factors for severe covid- among young patients and has several major findings: ( ) this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved data are presented as n(%) or median(q , q ). abbreviation: ct, computed tomography. this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved or, odds ratio. ci, confidence interval. bmi, body mass index. fbs, fast blood sugar. ldh, lactate dehydrogenase. cr, creatinine. +∞, positive infinity. ne, not estimated. this article is protected by copyright. all rights reserved or, odds ratio. ci, confidence interval. bmi, body mass index. fbs, fast blood sugar. ldh, lactate dehydrogenase. cr, creatinine. +∞, positive infinity. who coronavirus disease (covid- ) situation dashboard a pathological report of three covid- cases by minimally invasive autopsies covid- autopsies review and prospect of pathological features of corona accepted article this article is protected by copyright. all rights reserved virus disease analysis of heart injury laboratory parameters in covid- patients in one hospital in wuhan, china sars-cov- inflames the heart. the importance of awareness of myocardial injury in covid- patients liver injury in covid- : management and challenges covid- and kidney failure in the acute care setting: our experience from seattle pathological evidence of pulmonary thrombotic phenomena in severe covid- clinical characteristics of deceased patients with coronavirus disease : retrospective study the clinical and chest ct features associated with severe and critical covid- pneumonia characteristics of and important lessons from the coronavirus accepted article this article is protected by copyright. all rights reserved disease (covid- ) outbreak in china: summary of a report of cases from the chinese center for disease control and prevention clinical characteristics of medical workers infected with new coronavirus pneumonia zhonghua jie he he hu xi za zhi covid- in critically ill patients in the seattle region -case series factors associated with hospitalization and critical illness among , patients with covid- disease in new york city high prevalence of obesity in severe acute respiratory syndrome coronavirus- (sars-cov- ) requiring invasive mechanical ventilation obesity in patients younger than years is a risk factor for covid- hospital admission new coronavirus pneumonia prevention and control program liver fat accumulation assessed by computed tomography is an independent risk factor for diabetes mellitus in a population-based study: sessa (shiga epidemiological study of subclinical atherosclerosis) obesity and covid- severity in a obesity in patients younger than years is a risk factor for covid- hospital admission obesity as a risk factor for greater severity of covid- in patients with metabolic associated fatty liver disease sex differences in the prevalence and adverse outcomes of sarcopenia and sarcopenic obesity in community dwelling elderly in east china using the awgs criteria analysis of factors associated with disease outcomes in hospitalized patients with novel coronavirus disease risk factors for disease severity, unimprovement, and mortality of covid- patients in wuhan, china clinical and biochemical indexes from -ncov infected patients linked to viral loads and lung injury accepted article this article is protected by copyright. all rights reserved a tool to early predict severe corona virus disease (covid- ) : a multicenter study using the risk nomogram in wuhan and guangdong obesity and morbid obesity associated with higher odds of hypoalbuminemia in adults without liver disease or renal failure impaired albumin function: a novel potential indicator for liver function damage? renal histopathological analysis of postmortem findings of patients with covid- in china the role of podocytes in proteinuria. nephrology (carlton) albumin in chronic liver disease: structure, functions and therapeutic implications clinical characteristics in urolithiasis formation according to body mass index clinical course and risk factors for mortality of adult inpatients with covid- in wuhan, china: a retrospective cohort study accepted article key: cord- -ltmu ncu authors: pfitscher, l. c.; cecatti, j. g.; pacagnella, r. c.; haddad, s. m.; parpinelli, m. a.; souza, j. p.; quintana, s. m.; surita, f. g.; sousa, m. h.; costa, m. l. title: severe maternal morbidity due to respiratory disease and impact of h n influenza a pandemic in brazil: results from a national multicenter cross-sectional study date: - - journal: bmc infect dis doi: . /s - - -z sha: doc_id: cord_uid: ltmu ncu background: the aim of this study was to assess the burden of respiratory disease, considering the influenza a pandemic season (h n pdm ), within the brazilian network for surveillance of severe maternal morbidity, and factors associated with worse maternal outcome. methods: a multicenter cross-sectional study, involving referral maternity hospitals in five brazilian regions. cases were identified in a prospective surveillance by using the who standardized criteria for potentially life-threatening conditions (pltc) and maternal near miss (mnm). women with severe complications from respiratory disease identified as suspected or confirmed cases of h n influenza or respiratory failure were compared to those with other causes of severe morbidity. a review of suspected h n influenza cases classified women as non-tested, tested positive and tested negative, comparing their outcomes. factors associated with severe maternal outcome (smo = mnm + md) were assessed in both groups, in comparison to pltc, using pr and % ci adjusted for design effect of cluster sampling. results: among cases of severe maternal morbidity, ( %) had respiratory disease. respiratory disease occurred in one-quarter of mnm cases and two-thirds of md. h n virus was suspected in cases with respiratory illness. around % of these women were tested, yielding confirmed cases. confirmed h n influenza cases had worse adverse outcomes (mnm:md ratio < ( . : ), compared to : in cases due to other causes), and a mortality index > %, in comparison to . % in other causes of severe maternal morbidity. delay in medical care was associated with smo in all cases considered, with a two-fold increased risk among respiratory disease patients. perinatal outcome was worse in cases complicated by respiratory disease, with increased prematurity, stillbirth, low birth weight and apgar score < . conclusions: respiratory disease, especially considering the influenza season, is a very severe cause of maternal near miss and death. increased awareness about this condition, preventive vaccination during pregnancy, early diagnosis and treatment are required to improve maternal health. background improvement in maternal health aiming a reduction in maternal mortality is a priority worldwide, in an attempt to accomplish the established millennium development goals set for [ ] [ ] [ ] . however, the expected reduction in maternal mortality is still far from ideal and varies widely across regions [ , ] . most recently, to better comprehend the burden of disease on female health and complement mortality inquiries, an alternative approach has been to study maternal morbidity. maternal morbidity can have an impact on both low-income and highincome settings. in , the world health organization (who) standardized the definitions of potentially life-threatening conditions (pltc) and maternal near miss (mnm) [ ] . pltc is defined by the number of maternal complications, including hemorrhagic (e.g., abruption placenta, ruptured uterus, atony and others), hypertensive disorders (e.g., severe preeclampsia, eclampsia, hellp syndrome), management indicators of severity (e.g., blood transfusion, intubation, intensive care unit admission) and other complications (e.g., pulmonary edema, cardiac disease and sepsis). maternal near miss (mnm) is any situation in which a woman survives a very severe complication with proven organ dysfunction, during pregnancy or puerperium ( days after childbirth), with at least one of the following criteria: clinical (e.g., shock or clotting disorder), laboratory (lactate > , pao /fio < mmhg) or management (hysterectomy due to infection or hemorrhage and blood transfusion ≥ units of packed red blood cells). severe maternal outcome (smo) accounts for cases of mnm plus maternal deaths (md) [ ] . recently, the concept of "obstetric transition" was incorporated into the study of maternal morbidity and mortality. the concept illustrates a global trend in which a pattern of high maternal mortality rates with predominantly direct obstetric causes (e.g., hemorrhage, preeclampsia or uterine infection) has been replaced by lower maternal mortality rates with an increasing proportion of indirect causes (preexisting disorders or those aggravated by pregnancy, such as cardiac disease, kidney disease or infection due to urinary or pulmonary foci), institutionalization and medicalization of childbirth and increased rate of obstetric interventions [ ] . obstetric transition is important to help understand the occurrence of severe maternal morbidity and provide patients with the appropriate treatment in different settings. among the indirect causes of maternal morbidity and mortality, respiratory disease plays a significant role, either due to the presence of severe infection or complications of the underlying conditions, such as asthma and heart disease. physiological and anatomical changes that occur during pregnancy to provide accommodation for the growing uterus, can affect the known clinical presentation of respiratory signs and symptoms. adequate diagnosis and treatment of respiratory disease may be delayed [ , ] . in addition, it is recognized that pregnancy may increase the risk of severe influenza-associated complications [ , ] . it became clear throughout the h n influenza pandemic [termed a(h n )pdm ] worldwide [ ] [ ] [ ] [ ] [ ] [ ] that pregnant women were a highly vulnerable group. from july to january , , , cases of the disease and deaths were reported in brazil, [ ] . however, the total number of cases and deaths were likely much higher than the notified number. we proposed a novel approach to analyzing the burden of h n influenza virus infection and other respiratory disease among patients with severe maternal morbidity. cases complicated by severe respiratory disease were compared to cases with morbid conditions due to other causes (such as hemorrhage and hypertension). in addition, factors possibly associated with a higher risk of smo were evaluated by using the who standardized definitions of morbidity in referral maternity hospitals. this study is a secondary analysis of the brazilian network for surveillance of severe maternal morbidity including referral maternity hospitals in brazil. the study evaluated severe maternal morbidity cases, from a prospective surveillance, according to the who newly publicized criteria for these conditions [ ] . the methodological details of the original study have already been published elsewhere [ , ] . briefly, this multicenter study included referral maternity hospitals distributed among the five brazilian geographical regions. from july to june , all women admitted to participating centers, who were identified as having any life-threatening condition, near miss or maternal death, according to the who definition, were included in the study. data collection, by the study team, was acquired through medical chart review after hospital discharge or death of the patient. if any doubt on diagnosis considered, the treating doctors were further contacted for clarifications. information was entered into the openclinica® electronic platform (version . . -waltham, ma, usa) through a structured form completed by the local coordinator from each participating center. this was not a population based study, however, there was a concern to reduce the impact of nonrandom sampling and an effort to consider representativeness of the national territory (with health facilities from all five macro-regions of the country) and of facilities from public and private sectors, university and non-university hospitals. all selected hospitals had to provide information concerning their characteristics, including location, complexity of level of care, population covered, number of maternity beds and availability of resources for severe cases. quality control was carried out during various phases of the study. initially, training was provided to the entire team participating in the study, using a detailed operations manual, with the definition of each variable. meetings were held between the local research team and the coordinating team of the study to standardize data. case review was conducted by the local investigator. subsequently, the coordinating team of the study performed random reviews of manual and electronic forms for data consistency in visits to monitor the centers' performance. periodically, review of the electronic system was carried out to check for data inconsistency, along with systematic case review. some reported conditions were delay or substandard care, which had been previously reported [ ] . reasons for the delay in treatment were the woman or family member (including delay in identifying the condition, seeking care and refusing to accept treatment), health service (difficulties in obtaining equipment or medical supplies) or health professional (delays in identifying the correct diagnosis and providing appropriate patient treatment). sample size was determined by the prevalence of about maternal near miss cases per births and a maternal mortality ratio of / , live-born infants ( % confidence interval). it was predicted that , births [ ] needed to be monitored. for the present analysis, we considered severe respiratory disease as a suspected or confirmed case of influenza or acute respiratory failure, defined as incapacity of the respiratory system to promote adequate gas exchange, with arterial blood gas parameters: pao < mmhg or peripheral saturation < %, associated or not with paco > mmhg. clinical parameters such as tachypnea (respiratory rate-rr > ) or bradypnea (rr < ), use of accessory respiratory muscles, nasal flaring, associated with torpor or agitation were also considered. for suspected or confirmed cases of a(h n )pdm , the definition of cases considered only those with severe morbidity, including acute respiratory insufficiency, sepsis, intensive care admission, intubation and others. cases of h n without severe complications were not included. a review of all h n influenza cases was necessary to confirm whether laboratory tests had been performed and to obtain the results of these tests, since data in the original study had not been collected in detail. case review was requested from each local center and new data were distributed into three groups: non-tested, positive and negative cases for h n influenza virus. initially, the prevalence of pltc, mnm and md was calculated per group, as well as the respective health indicators related to maternal morbidity and mortality: maternal near miss ratio, severe maternal outcome ratio, mortality index and maternal mortality ratio, according to the who definition [ ] . to evaluate the progression of severe maternal morbidity in cases complicated by respiratory disease throughout the study, maternal outcomes (pltc, mnm and md) were measured for each month studied. the risk of smo associated with procedures used to manage the severity of conditions was estimated for the group with severe respiratory disease and other causes of severe maternal morbidity, using prevalence ratios plus their respective % ci adjusted for the design effect of cluster sampling. subsequently, we performed an analysis considering the total number of cases with severe respiratory disease versus cases with other causes of severe maternal morbidity. in each group, pltc (less severe cases) and severe maternal outcome (smo: mnm + md) cases were compared to evaluate the factors potentially associated with more severe disease, including delay in obstetric care, also using the prevalence ratios plus their respective % ci adjusted for the design effect of cluster sampling. the prevalence of sociodemographic, obstetric and perinatal factors were evaluated between the two groups using chi-square tests. values statistically significant were considered those with a p-value under . . the statistical procedures for analysis were performed with spss and stata. during the -month study period, , women were screened. of these, had criteria for severe maternal morbidity. among these women, only ( %) had severe respiratory disease. however, in this group with respiratory illness, symptom severity progressed more rapidly, if compared to other causes of severe morbidity ( fig. ), such as bleeding or hypertensive disorders, and may be times more lethal. among the total number of women with respiratory disease, patients with suspected h n influenza a virus infection had more severe disease ( . % mnm and . % md) than those without suspected h n influenza a virus (prevalence of mnm: . %, md: . %) ( fig. ) . about % of cases of suspected h n influenza a were tested. women who tested positive ( cases) for h n had more severe disease, with a higher prevalence of smo. figure shows the distribution of cases with severe respiratory disease, according to progression of severity during the study period, based on date of admission in participating centers. there was a higher incidence of cases in the first months considered, especially july, august and september . national guidelines and availability of vaccination during pregnancy were instituted in march/ . considering health indicators, disease was more severe among cases tested and positive for h n (table ) . mortality rate was higher than % among positive cases for a(h n )pdm . the death rate was about % in cases testing negative for h n and . % in non-tested cases. in contrast, the mortality rate was only . % in morbid disorders due to other causes. the maternal near miss to mortality ratio was . : , . : and . : , among positive, negative and non-tested groups for a(h n )pdm , respectively, compared to a value of . : for other causes of severe maternal morbidity. more than % of patients with severe respiratory disease had three diagnostic criteria for near miss: laboratory, clinical and management, while for the remaining causes of severe maternal morbidity, around % of patients only had criteria for laboratory or management diagnosis ( table ). all procedures for management of severity were associated with a worse outcome in both groups, women with severe respiratory disease and those with severe maternal morbidity due to other causes ( table ) . analysis of sociodemographic and obstetric characteristics ( there was an association to smo and non-white color, history of diabetes, low weight and substance abuse (use of psychoactive substances, including alcohol and illicit drugs), in addition to delay in care. in contrast, the group of cases due to other causes of morbidity, low maternal age, first pregnancy, history of maternal obesity and lack of a partner were identified as having lower association to smo, while hospitalization in a non-public institution, parity, history of caesarian section, drug abuse, complication occurrence at an earlier gestational age and mainly in the postpartum period, in addition to any type of delay in obstetric care, were associated with smo. concerning characteristics of pregnancy and perinatal results (table ) , the group with severe respiratory disease had a higher rate of early preterm births, between and weeks of gestation, low birthweight (< g), apgar < at five minutes of life, stillborn and the need for hospital admission/transference of the newborn infant, compared to the group with severe maternal morbidity due to other causes. neonatal death increased threefold in women with severe respiratory disease. a statistically significant difference was observed in the groups compared, when the mode of delivery and onset of labor were taken into consideration (p < . ). the number of women who did not undergo pregnancy resolution and remained pregnant during the severe morbid event was much higher in the respiratory disease group. around % were "still pregnant" compared to % in the group with severe maternal morbidity due to other causes. our study presents the burden of severe respiratory diseases among cases of severe maternal morbidity and results of the h n influenza pandemic, considering referral maternity hospitals in brazil. overall, the prevalence of respiratory disease was rare ( %). nevertheless, respiratory disease accounted for one-quarter of mnm cases and two-thirds of md. worse adverse outcomes occurred among cases of confirmed a(h n )pdm , with an impressive mnm:md ratio below one, meaning that there were more deaths than near miss cases in this group. the mortality index (mi) was over % in the h n group, compared to . % for other causes of severe maternal morbidity. the mi is known to correlate to quality of care and when the index is above %, it represents substandard care [ ] . numbers of mi over % most likely reflect that poor outcomes were not only due to the severity of disease, but also to substandard care, including delays in diagnosis and management of the considered cases. our data further confirmed that the increased risk of smo was linked to delays in health care (delays due to women/family members, health services or health professionals). considering the impact of the a(h n )pdm on maternal health [ ] , a great effort towards prevention occurred worldwide, with strong recommendations for vaccination during pregnancy and empirical antiviral *adjusted for design effect of cluster sampling therapy, as soon as possible in case of suspected disease [ ] . brazil followed these recommendations and launched a national vaccination campaign before the winter of , targeted at high-risk groups, including pregnancy. the vaccine was available in all public health facilities, at no cost for the patient and reached very high coverage (around %), most likely due to the long term experience in the national immunization program for children and due to the awareness about the severity of the disease, among health professionals and among the society [ ] . we cannot evaluate the impact of those preventive measures in our study, since we lack information on the total number of cases and specific data on individual history of vaccination or treatment, however, from our fig. , we can see on the linear traces that there is a trend towards decrease in numbers of severe cases, through time, especially after vaccination. clinical evaluation should determine treatment, in order to ensure timely and effective interventions. in our study, around % of suspected cases of h n influenza a virus were tested. in accordance with previous reports, symptoms were more severe in positive cases [ ] . the majority of cases in brazil occurred during cold weather (july, august and september), period of increased infections by respiratory viruses and influenza outbreak in the country (brazil declared a pandemic in mid-july ). over half of the reported cases of severe respiratory disease were not due to suspected influenza infection. acute respiratory failure was the cause, including a broad number of conditions, as follows: pulmonary edema, cardiac disease community-acquired pneumonia, aspiration, pulmonary embolism, asthma exacerbation or venous embolism. unfortunately we do not have detailed information on each of the mentioned causes. nevertheless, these complications include mostly indirect causes of maternal morbidity and mortality, which represent novel or preexisting health problems unrelated to pregnancy, such as cardiac disease and asthma. asthma is the most common medical condition that may worsen during pregnancy and it is often underdiagnosed and under-treated [ ] . direct causes of maternal morbidity and mortality can also lead to respiratory failure, such as systemic consequences of sepsis due to uterine infection and severe preeclampsia and eclampsia, complicated by pulmonary edema [ , ] . it is very important to understand all differential diagnosis, since timely and adequate interventions can potentially improve maternal outcome. future studies focusing on the specific differences in diagnosis and management of causes of acute respiratory failure should consider the main aspects on diagnosis and management of these conditions. pneumonia in pregnancy and postpartum, for example, is the leading cause of fatal none obstetric infection and can be caused by bacteria, virus (at risk of secondary bacterial infection), fungus and mycobacteria and the clinical features include fever, cough, dyspnea and hypoxia [ ] . another important cause of severe complications is pulmonary edema, which can be consequence of left ventricular systolic or diastolic dysfunction, or due to the use of tocolytic agent, fluid overload, severe hypertension or severe renal disease. the clinical presentation of pulmonary edema is normally dyspnea, tachypnea, tachycardia, chest pain and diffuse crackles. there can be evidence of cardiac dysfunction, specific alterations in the electrocardiogram and radiographic abnormalities [ ] . during labor or immediate postpartum, a rare and feared complication is the aspiration of gastric contents, if needed intubation for general anesthesia, due to increased intraabdominal pressure and predisposing physiological changes of pregnancy such as relaxation of the lower esophageal sphincter and delayed gastric emptying. however, in the last decades, the incidence of aspiration significantly declined, even with food intake during labor [ ] . the definitions of severe respiratory complications that are usually reported can be rather confusing and sometimes difficult to incorporate [ ] . recent onset of fever and respiratory symptoms, including cough is the clinical definition of severe acute respiratory syndrome. in the setting of an epidemic, this definition is very useful to raise awareness and ensure prompt treatment, as soon as a suspected case is identified [ ] . ards is another acronym for acute respiratory distress syndrome, a different condition that represents hypoxemic respiratory failure and bilateral radiographic opacities, without congestive heart failure. this diagnosis depends on oxygenation deficit measurements and chest imaging [ , ] . in the current study, we couldn't accurately establish any of the above conditions, since we did not collect data on clinical symptoms (fever, cough) neither obtained the results of those specific laboratory findings or imaging. the diagnosis of h n influenza was also not standardized through all hospitals included. we understand that timing of sample collection, quality of sample and laboratory procedures are key for the accurate diagnosis and unfortunately we do not have data on details regarding these procedures. another limitation was the lack of data on the use of antiviral therapy or vaccination. for cases of smo, complicated by documented organ dysfunction, ards would probably be the diagnosis of respiratory disease. nevertheless, confirmation was lacking for all cases. in addition, we do not have a control group, with no underlying complication, what would be key to access risk factors. we only have data on severe maternal morbidity cases, comparing less severe (pltc) to more severe cases (smo). factors associated with smo, included non-white color, history of diabetes, low weight and substance abuse, along with delay in care, were reported for the majority of conditions under study. substance abuse associated with increased risk of severity in cases of respiratory disease, is in agreement with previous reports [ , ] . drug-related severe respiratory complications can occur, resulting from parenchymal (infectious and non-infectious pneumonitis, aspiration-related events, hemorrhage, pulmonary edema and pneumothorax), pulmonary vascular insults (endovascular infection, hemorrhage, and vasoconstriction) or airway (bronchospasm and hemorrhage) abnormalities. diabetes was also associated with an increased risk of smo among cases complicated by severe respiratory disease. previous studies had demonstrated this fact, even in the brazilian population. diabetes is one of the main risk factors for death from h n influenza a [ ] virus infection. medical history, including known factors related to worse outcomes should be highlighted and the awareness among patients and health professionals towards targeted cases could impact in the final outcome. pregnancy characteristics and perinatal outcomes according to the main cause of severe morbidity showed that pregnancies complicated by respiratory disease present an increased rate of preterm delivery and worse perinatal outcomes. this finding had already been demonstrated [ , ] . studies have shown that vaccination during the first trimester of pregnancy can improve those outcomes and decrease stillbirth rates without increasing the risk of malformations, which is a common concern among health practitioners and pregnant women [ ] . severe respiratory disease, especially considering the influenza season, is one of the most serious causes of maternal near miss and death. increased awareness of this condition, preventive vaccination during pregnancy, early diagnosis and treatment are required to improve maternal health. abbreviations smm: severe matenal morbidity cpr: cardiopulmonary resuscitation; pr: prevalence ratio a(h n )pdm : h n influenza pandemic; ards: acute respiratory distress syndrome; ci: confidential interval; md: maternal deaths; mi: mortality index mnm: maternal near miss; pltc: potentially life-threatening conditions rr: respiratory rate; smo: severe maternal outcome; who: world health organization hospital das clınicas da universidade federal de pernambuco united nations department of economic and social affairs maternal mortality for countries, - : a systematic analysis of progress towards millennium development goal targets and strategies for ending preventable maternal mortality: consensus statement. geneva: world health organization ending preventable maternal deaths: the time is now trends in maternal mortality: to . geneva: world health organization who working group on maternal mortality and maternal morbidity classifications. maternal near miss -towards a standard tool for monitoring quality of maternal health care obstetric transition: the pathway towards ending preventable maternal deaths surveillance for emerging respiratory viruses respiratory disease in pregnancy h n influenza and pregnancy- years later clinical aspects of pandemic influenza a (h n ) virus infection maternal mortality due to pandemic influenza 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a national multicenter cross-sectional study incidence and predictors of severe obstetric morbidity: case control study a(h n ) vaccination in pregnant women in brazil: identifying coverage and associated factors a cross-sectional analysis of symptom severity in adults with influenza and other acute respiratory illness in the outpatient setting the pulmonary edema preeclampsia evaluation (pepe) study pulmonary edema in severe preeclampsia-eclampsia: analysis of thirty-seven consecutive cases acute pulmonary edema in pregnancy effect of food intake during labour on obstetric outcome: randomised controlled trial personal view of sars: confusing definition, confusing diagnoses clinical management and infection control of sars: lessons learned critical illness in pregnancy: part ii: common medical conditions complicating pregnancy and puerperium ards in pregnancy the large spectrum of pulmonary complications following illicit drug use: features and mechanisms acute respiratory failure from abused substances risk factors for death from influenza a(h n )pdm , state of são paulo, brazil influenza and pregnancy in the united states: before, during, and after h n effect of influenza vaccination in the first trimester of pregnancy the authors declare that they have no competing interests.ethics and consent to participate the study complied with ethical principles guiding human research described in the declaration of helsinki and was approved by the institutional review board of each local center and research coordinating center (conep, brazilian ministry of helath: / ). the informed consent form was waived, since data were obtained from hospital records after patient discharge, without any contact with research subjects. the review boards of the following institutions reviewed and approved this study: maternidade cidade nova dona nazarina daou (manaus, am), consent to publish not applicable. all the data supporting the presented findings is contained within the manuscript. the idea for the study and this specific analytical approach arose in a group discussion among all the authors. analyses were planned by lcp, mlc and jgc. the first version of the manuscript was drafted by lcp and mlc. subsequently, all remaining authors complemented with suggestions. all authors contributed to the development of the study protocol and approved the final version of the manuscript. key: cord- -r ser c authors: matricardi, paolo maria; dal negro, roberto walter; nisini, roberto title: the first, holistic immunological model of covid‐ : implications for prevention, diagnosis, and public health measures date: - - journal: pediatr allergy immunol doi: . /pai. sha: doc_id: cord_uid: r ser c the natural history of covid‐ caused by sars‐cov‐ is extremely variable, ranging from asymptomatic or mild infection, mainly in children, to multi‐organ failure, eventually fatal, mainly in the eldest. we propose here the first model, explaining how the outcome of first, crucial ‐ days after infection, hangs on the balance between the cumulative dose of viral exposure and the efficacy of the local innate immune response (natural iga and igm antibodies, mannose binding lectin ). if sars‐cov‐ runs the blockade of this innate immunity and spreads from the upper airways to the alveoli in the early phases of the infections, it can replicate with no local resistance, causing pneumonia and releasing high amounts of antigens. the delayed and strong adaptive immune response (high affinity igm and igg antibodies) that follows, causes severe inflammation and triggers mediator cascades (complement, coagulation, and cytokine storm) leading to complications often requiring intensive therapy and being, in some patients, fatal. low‐moderate physical activity can still be recommended. however, extreme physical activity and hyperventilation during the incubation days and early stages of covid‐ , facilitates early direct penetration of high numbers of virus particles in the lower airways and the alveoli, without impacting on the airway’s mucosae covered by neutralizing antibodies. this allows the virus bypassing the efficient immune barrier of the upper airways mucosa in already infected, young and otherwise healthy athletes. in conclusion, whether the virus or the adaptative immune response reach the lungs first, is a crucial factor deciding the fate of the patient. this “quantitative and time‐sequence dependent” model has several implications for prevention, diagnosis, and therapy of covid‐ at all ages. most serological data currently available in the literature refer to patients examined mostly in the acute phase of the disease. therefore, they are insufficient to exactly establish durability of the antibody titers of each isotype peak when they eventually disappear. the levels of serum igg antibodies, however seems to be proportional to the intensity of the viral load and to the symptom severity. [ , ] the efficacy of specific ig and their role in limiting viral spread may be indirectly assumed by observations demonstrating that plasma from subjects recovered from covid- showed a therapeutic efficacy if passively transferred to patients.[ , ] similar effectiveness had been this article is protected by copyright. all rights reserved already demonstrated for plasma from patients having recovered from sars-cov and mers-cov. [ , ] consequently, infusion of plasma from convalescent individuals to critically ill covid- patients is a therapeutic option that is being investigated. although controlled clinical trials are not yet available, several papers report the efficacy of this treatment and the lack of serious adverse events. [ , ] convalescent plasma was administered in patients with a severe disease, and it is unclear whether earlier administration might have been associated with different clinical outcomes [ ] and with the prevention of respiratory distress. only a small proportion of humans younger than , among those who get infected by sars-cov- , suffer from moderate and severe covid- . [ - ] among them, hospital doctors frequently exposed to covid- patients are, unfortunately, highly represented. [ ] dr. li wenliang, the first man alerting china and the world of the new infection, died from covid- at the age of . [ ] similarly, dr. carlo urbani, i.e. the first man alerting the of sars-cov, died of sars at the age of . [ ] both doctors cared for weeks patients with severe pneumonia with no personal protection. [ , ] in italy, doctors exposed to sars-cov- , have so far ( th april, ) died of covid. [ ] the case fatality ratio among doctors working in hospitals and caring patients developing severe covid- has been therefore much higher than among their age and gender matched peers. [ , ] (table- observations in previous viral epidemics, further clarify this aspect. the reliability of high viral loads in nasopharyngeal specimens as a prognostic indicator of respiratory failure or mortality, with or without a high viral load in serum, has been previously characterized in sars. [ ] a link has been established between the initial dose and subsequent severity of the disease to the - spanish flu pandemic. it was demonstrated by simulation models that the number of this article is protected by copyright. all rights reserved simultaneous contacts a susceptible person has with infectious ones are correlated with the infectious dose; that severe cases of influenza result from higher infectious doses of the virus; and that a susceptible person can be easily exposed to very high infectious doses of influenza in over-crowded places. [ ] the viral replication is more active and prolonged in patients suffering from severe influenza. viral clearance is slow when host defenses are weakened, however it is enhanced when antivirals start within the first days of illness. [ ] among over thousand chinese with covid- , most were aged over years ( %), while only % were aged years or younger, % were aged to years an % aged to years. [ ] moreover, most of the relatively few pediatric cases were classified as mild ( %), only % severe and % critical. [ this article is protected by copyright. all rights reserved covid- mortality has been lower among chinese females than males. [ ] in italy, mortality and hospitalization rates have been also more frequent among males than among females. [ ] moreover, patients with blood group and a have slightly lower and a slightly higher risk, the first diagnosis of covid- in europe has been confirmed in a year-old italian healthy male who regularly participated in running events and soccer games. one day before starting covid- symptoms, he had been training sport. the time-lapse between the onset of upper airways symptoms and pneumonia was days only. only days after the onset of covid- , the patient was admitted to the intensive care unit of the policlinico san matteo in pavia because of respiratory failure. after weeks of intubation and supportive treatment, the patient luckily recovered and could be discharged in good conditions. the italian first covid- case is worldwide famous but, surprisingly, no official study on it has been so far published. the example of this physically active, young patient offers room for reasoning with regard to the importance of sport for virus transmission and course of disease. indeed, other cases of covid- in (semi-) professional athletes have been described. the pattern of breathing during strenuous exercise changes dramatically by a tremendous increase of ventilation (i.e.: inspiratory and expiratory volumes of air), and of alveolar ventilation in particular. obviously, these changes mostly attain to whatever kind of runners belonging to all sport disciplines, being semi-professional and professional athletes particularly exposed (such as much more than individuals of common population) due to their frequent practice of extreme and long-lasting exercise. furthermore, the majority of these athletes have their lungs that usually work in perfect physiological conditions, such as very close to those of the "ideal lung". in other words, in the absence of any anatomical or physiological factor causing a significant unevenness in distribution of their alveolar ventilation. paradoxically, these pre-existing ideal this article is protected by copyright. all rights reserved conditions significantly favor the deep inhalation of several irritants, allergens, infectious agents. even the sars-cov- can then spread more easily to the deepest areas of the lungs (alveolar bronchioles and alveoli) during strenuous exercise, and there starts its aggressive action. not by chance, a great proportion of professional football players claimed the occurrence of fever, dry cough and malaise (and dyspnea in some cases) immediately after, or a few hours following their last official match. in covid- , the occurrence of pneumonia requiring oxygen therapy is a critical event discriminating asymptomatic or mild cases, whose infection remains mostly confined to the upper airways, from those with severe disease, who experience massive viral invasion of their lower airways. [ ] what makes the difference? what prevents the virus from rapidly reaching the lungs and then causing severe pneumonia? what makes covid- pneumonia a life-threatening disease? ) pneumonia may start before adaptive immune response develops; [ ] ) serious complications begin together with the adaptive immune response. the first two weeks after infection are crucial. [ , ] innate immunity is the only first-line, early defense against the new sars-cov- virus. consequently, the early confrontation between host's innate immunity and sars-cov- , at exposure and during the following two weeks, decides the natural history of the disease. this confrontation also decides whether the infection will be efficiently blocked in upper airways, or how many virus particles reach the lungs, and when. to understand which part of the innate immunity involved in early protection from sars-cov- , we have: ) examined which primary immune deficiencies are associated with pneumonia. ) examined the patterns of risk factors for covid- severity: dose of exposure to sars-cov- ; age, gender, abo group; ) identified the innate immunity components fitting the same patterns of risk factors; this article is protected by copyright. all rights reserved ) examined the biological plausibility that the candidate molecules, emerging from the previous reasoning, are really essential in limiting the consequences of sars-cov- infection to upper airways or to mitigate the course of pneumonia. these data suggest that the lack of natural igm and iga in the upper respiratory airways may have contributed to the rapid viral spread to lungs, causing pneumonia. unexpectedly in immunodeficient individuals, agammaglobulinemic patients, who are unable to develop specific sars-cov- igs, did not develop severe pneumonia, suggesting that the serious complications observed in other patients may be related to the development of acquired immunity. under the circumstances described above, innate immunity become an obvious candidate to act as very first barrier protecting of children, almost all adults and most elders from sars-cov- . innate immunity is essential to control virus replication early enough, before a very effective adaptive immune response is generated. [ ] accepted article this article is protected by copyright. all rights reserved we focused on humoral components and, in particular on natural antibodies and mbl, to ascertain whether these players of the innate immunity fit all the epidemiological and clinical pre-conditions presented in the last three months by sars-cov- . finally, we tentatively describe mechanisms beyond the most severe cases of pneumonia as a possible consequence of the development of adaptive immunity in individuals with an early high viral spread in lungs. anti-glycan natural antibodies are detected in serum in the absence of previous immunization, are observed also in gnotobiotic animals, and belong mostly to the igm isotype [ ] but also to the iga and igg isotype. (b) old patients -viral exposure is probably higher (the source of contagion is also an old person) but the innate immunity is much weaker; a high number of viral particles can reach the alveoli and replicate in type ii pneumocytes in coincidence or even much before the this article is protected by copyright. all rights reserved expansion of the specific immune response leading to a more severe and symptomatic pneumonia; (c) young but highly exposed patients -the exposure to an excessive cumulative viral dose (i.e. unprotected health care personnel) will overcome their efficient innate immunity. viral particles will reach the alveoli in early stages and cause symptomatic pneumonia; this article is protected by copyright. all rights reserved macrophages via fc-receptors. in addition, ig binding to the s protein of sars-cov- may cause its conformational changes that make the binding to the ace- receptor more effective for the viral fusion with the cell membrane. the local high concentration of cytokines and chemokines that contribute to recruitment of inflammatory cells and vasodilatation, permits to serum natural igs and mbl to maintain a vicious circle of inflammation with complement activation and immunocomplexes deposition. in this light, it cannot be excluded that mbl or igm mediated immunocomplexes contribute to activation of platelets or tissue factor leading to coagulation and microthrombosis that have been described in covid- patients with acute respiratory failure. in this phase of the disease, natural igm and mbl that circulate in serum may have no protective role, but, rather may contribute to tissue damage. moreover, during this second phase of the disease, the adaptive response is also progressively on the increase. this may be one side protective against further virus spread in the lungs,but may also reinforce the immunological and coagulation cascades provoking complications. mbl binds to polymeric iga and initiates complement cascade, a defense against invading pathogens in mucosal immunity. polymeric iga also has a role in activating lectin-mediated complement signaling. the complement cascade links the innate and the adaptive immune system, protecting against invading pathogens during the first phase of the diseases. in this sense, ab-mediated complement activation flows in parallel between mbl and c q. additionally, it can boost proinflammatory effects of iga deposition with the same mechanism that is supposed to occur in the glomerulus, and results in renal injury. this article is protected by copyright. all rights reserved evidence-based medicine should also apply for covid- patients, so that "new" or off-label drugs or treatment regimens should only be given in a clinical study context, following approval by relevant national or international agencies. however, we believe that these points are first priorities for intensively and focused clinically oriented research. microarras and other tests aimed at measuring natural antibodies, that might be protective against sars-cov- and other viruses, should be developed. individuals with low natural antibody and mbl levels should be identified and specifically protected. moreover, governments promoting herd immunity must protect individuals, even if young, who may have low levels of natural antibodies. these individuals should be not exposed to the virus, especially if shed at high doses. including strenuous sport activities requiring high respiratory volumes and flows, should be avoided during the early stage of infection. when the adaptive immune response is this article is protected by copyright. all rights reserved still not initiated. particular precautions should be given to athletes performing fatiguing sports, since a portion of sub-micrometer-size, aerosolized particles, are expired by the runner or eliminated by cough or nasal secretions and may contain viruses if the athlete is an asymptomatic but sars-cov infected individual. these droplets or aerosol might be re-inhaled and facilitate the spread of the virus from the upper to the lower airways. this article is protected by copyright. all rights reserved moieties flanking sars-cov- s -rbs may be useful, among elders, to identify those at higher risk of severe disease. glycan microarrays will be instrumental for this target. paucisymptomatic individuals could be pursued by measuring sars-cov- specific serum igg and iga that are expected to persist as a memory response to infection. appropriate and high performing validated tests should be used to retrospectively evaluate the seroconversion status to estimate the herd immunity of a given population. immunization strategy: innate and adaptive immunity . . while effective vaccines are being developed, produced, tested, and validated, a strategy to stimulate innate immunity and natural igm antibody production in particular, would empower the defences of at-risk elderly population. these measures may include influenza, 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sars-cov in human lungs: an ex vivo study with implications for the pathogenesis of covid- level of il- predicts respiratory failure in hospitalized symptomatic covid- patients off-label use of tocilizumab in patients with sars-cov- infection pathological study of the novel coronavirus disease (covid- ) through postmortem core biopsies abnormal coagulation parameters are associated with poor prognosis in patients with novel coronavirus pneumonia accepted article thrombomodulin is associated with organ failure and mortality in children with acute respiratory distress syndrome (ards): a prospective observational cohort study analytical performances of a chemiluminescence immunoassay for sars-cov- igm/igg and antibody kinetics microvascular covid- lung vessels obstructive thromboinflammatory syndrome (microclots): an atypical acute respiratory distress syndrome working hypothesis unexpected receptor functional mimicry elucidates activation of coronavirus fusion accepted article the text contains the personal opinion of the authors, not of their institutions: charité universitaetsmedizin berlin, germany (pmm) and istituto superiore di sanità, rome, italy (rn).all authors declare no conflict of interest. key: cord- -go cs r authors: wang, yafei; zhou, ying; yang, zhen; xia, dongping; hu, yi; geng, shuang title: clinical characteristics of patients with severe pneumonia caused by the sars-cov- in wuhan, china date: - - journal: respiration doi: . / sha: doc_id: cord_uid: go cs r background: a new virus broke out in wuhan, hubei, china, that was later named severe acute respiratory syndrome coronavirus (sars-cov- ). the clinical characteristics of severe pneumonia caused by sars-cov- are still not clear. objectives: the aim of this study was to explore the clinical characteristics and risk factors of severe pneumonia caused by the sars-cov- in wuhan, china. methods: the study included patients hospitalized at the central hospital of wuhan who were diagnosed with covid- . clinical features, chronic comorbidities, demographic data, laboratory examinations, and chest computed tomography (ct) scans were reviewed through electronic medical records. spss was used for data analysis to explore the clinical characteristics and risk factors of patients with severe pneumonia caused by sars-cov- . results: a total of patients diagnosed with covid- were included in the study, including with severe pneumonia and with nonsevere pneumonia. statistical analysis showed that advanced age, increased d-dimer, and decreased lymphocytes were characteristics of the patients with severe pneumonia. moreover, in the early stage of the disease, chest ct scans of patients with severe pneumonia showed that the illness can progress rapidly. conclusions: advanced age, decreased lymphocytes, and d-dimer elevation are important characteristics of patients with severe covid- . clinicians should focus on these characteristics to identify high-risk patients at an early stage. in december , a new type of unexplained pneumonia was reported in wuhan, hubei, china, which appeared to be related to the huanan seafood wholesale market [ ] [ ] [ ] . the disease spread rapidly from wuhan to the surrounding provinces and cities, earning the attention of all levels of government and the administrative departments of health. the chinese center for disease control and prevention (cdc) promptly organized the relevant disease control agencies, medical units, and research institutes to carry out investigations and treat- doi: . / ment. a new type of coronavirus was detected by researchers in a patient's bronchoalveolar lavage fluid sample on january , [ ] . the world health organization (who) named it severe acute respiratory syndrome coronavirus (sars-cov- ), and on january , it was announced that the new coronavirus epidemic had been listed as a public health emergency of international concern. the disease caused by sars-cov- was named coronavirus disease (covid- ) on february , . as of : on april , , there were , confirmed cases, suspected cases, , cured cases, and , deaths in china as well as , , confirmed cases, , cured cases, and , deaths collectively in all other countries. the sars-cov- , which belongs to the genus betacoronavirus, is a single-strand, positive-strand rna virus that appears to be distinct from but is actually related to other coronaviruses, such as severe acute respiratory syndrome-related coronavirus (sarsr-cov) and middle east respiratory syndrome coronavirus (mersr-cov) [ ] [ ] [ ] [ ] [ ] . current studies have shown that sars-cov- has a sequence similarity with bat sars-like-covzxc of around % and a sequence similarity with human sars-cov of around % [ ] . just as the epidemics of sars and mers have challenged populations and global health care systems over the last years, so does sars-cov- [ ] . covid- is highly contagious, and may rapidly develop into severe pneumonia, acute respiratory distress syndrome (ards), multiple organ dysfunction syndrome (mods), and even death. therefore, the top priority for clinicians is to identify and treat the most severely affected patients in the disease's early stage. the aim of this study was to assess the potential high-risk factors for severe covid- and provide evidence for effective screening of severely afflicted patients. all patients in this study were hospitalized at the central hospital of wuhan, from january to february , . the patients were all admitted to the hospital because they were infected with sars-cov- and suffered from various symptoms, including fever, dyspnea, cough, and fatigue. every patient had completed the relevant laboratory examinations, including common pathogen detection tests and chest computed tomography (ct) scans. all patients were local residents of wuhan. moreover, most of the patients had a history of exposure to the huanan seafood wholesale market, or else had been in contact with people who had been either confirmed or suspected to have contracted the illness. sars-cov- nucleic acid detection results were positive for some patients and negative for others. these differing results could have been caused by the immaturity of the methods used which led to false-negative results. high-resolution ct scans with a scan layer thickness of mm and reconstruction of a thin layer ( - . mm) are recommended for radiological examination of covid- . based on patients' exposure history, clinical symptoms, laboratory examinations, and chest ct scans, all of them received a clinical diagnosis of covid- in accordance with who interim guidance [ ] . for patients suspected to have the illness, senior respiratory doctors made the diagnosis together. the patients were divided into groups. those considered to have severe pneumonia had the following severe manifestations: fever or suspected respiratory infection, in addition to a respiratory rate > breaths/min, severe respiratory distress, or spo < % on room air. patients with ards, sepsis, or septic shock were also included in this group. patients without the above severe signs were defined as having nonsevere pneumonia. patients who had covid- combined with other bacterial, fungal, or viral infections and those with missing data were excluded. data were extracted from the central hospital of wuhan, a tertiary teaching hospital responsible for the treatment for patients with covid- , as assigned by the chinese government. clinical features, chronic comorbidities, demographic data, laboratory examinations, and chest ct scans were reviewed using electronic medical records. laboratory examinations included routine blood tests as well as tests of liver and kidney function, and electrolyte, b-type natriuretic peptide, d-dimer, c-reactive protein (crp), and procalcitonin values. we obtained the lymphocyte absolute values of patients with severe pneumonia on the first and the third day after admission. d-dimer data of the patients with severe pneumonia were collected on the first, third, and seventh day after admission. chest ct scans were reviewed on the first and the third day for patients with severe pneumonia. additionally, albumin data were collected on the first and seventh day after admission for the severe pneumonia group. for patients with nonsevere pneumonia, we only collected data on the first day after admission because the relevant items were not frequently reviewed. for patients admitted to the ricu, the acute physiology and chronic health evaluation ii (apache-ii) score and a sequential organ failure assessment (sofa) were determined on the first day. the data were acquired by physicians. all data were checked by a second researcher to ascertain their accuracy. to reflect the progression of severe covid- , we calculated the difference in lymphocyte values between day and day , the difference in serum albumin values between day and day , and the difference in d-dimer values between days , , and . continuous variables were described with the mean ± standard deviation (sd) and compared using t tests if they were normally distributed. nonnormally distributed variables were described using the median and interquartile range (iqr). the mann-whitney u test was used for comparisons. categorical variables were expressed as n (%) and compared by a χ test or fisher's exact test. logistic regression analysis was used to assess the risk factors for severe pneumonia. the difference of a certain indicator in the same patient at different periods was shown by a bar chart. a twosided α < . was considered statistically significant. we used spss software v . for statistical analysis. . there was no significant difference in the history of smoking or alcohol consumption between the groups. the incidence of diabetes and cerebrovascular disease was also similar in both groups. according to the medical records, common symptoms at the onset of co-vid- were fever, fatigue, dry cough, and dyspnea; the initial symptoms of the patients with severe pneumonia were more commonly fever and dyspnea, but the difference was not statistically significant. the temperature range was divided into low (≤ ° c), moderate (≥ . ° c, ≤ ° c), and high (≥ . ° c) fever. however, there was no statistical difference between the groups with regard to temperature. there were a number of differences in the laboratory findings between the patients with severe pneumonia and those with nonsevere pneumonia (table ) , including a lower lymphocyte count, platelet count, and serum albumin in severely afflicted patients. the levels of serum creatinine, blood urea nitrogen, aspartate aminotransferase, crp, serum procalcitonin, d-dimer, and b-type natriuretic peptide were higher in the patients with severe pneumonia. the white blood cell count, neutrophil count, hemoglobin, and alanine aminotransferase results did not differ between the groups. statistically, some parameters differed between the groups, but were still within the normal range, including platelet count, and the level of serum creatinine, blood urea nitrogen, aspartate aminotransferase, and serum procalcitonin. binomial logistic regression analysis was used to assess the risk factors for severe pneumonia. the variables with statistical differences between the groups were incorporated into the logistic regression equation. to better understand the correlation between the lymphocyte and d-dimer measurements in the patients with severe pneumonia, we divided the values by their sds. the results revealed that, after adjusting for other confounding factors, age and d-dimer values were independent risk factors for severe pneumonia ( table ) . patients > years of age and those in the age group - years had a significantly higher risk of developing severe pneumonia than those < years of age. for every -sd increase in d-dimer value, the risk of developing severe pneumonia increased by about -fold. in addition, lymphocytes were found to be independent protective factors for severe pneumonia. the incidence of severe pneumonia in patients with a . increase in lymphocytes decreased by about . %. both the apache-ii and sofa scores were assessed in patients with severe pneumonia, with results of . (iqr - ) and (iqr - ), respectively (table ) . severe pneumonia usually progresses rapidly, and many clinical indicators can change in a short time, especially lymphocyte count, d-dimer and serum albumin values, and chest ct manifestations. to better identify severe pneumonia early, we calculated the difference in lymphocyte count and serum albumin and d-dimer values at different points in time (table ; fig. - ) . the results showed that the average difference in the lymphocyte and serum albumin values between day and day after admission were . (sd . ) and . (sd . ), respectively. the median difference in d-dimer values between day and day after admission was . (iqr . - . ), and the difference between day and day was sex, age, copd, hypertension, lymphocyte count, platelet count, serum creatinine, blood urea nitrogen, aspartate aminotransferase, serum albumin, c-reactive protein, serum procalcitonin, d-dimer, and b-type natriuretic peptide were used in the logistic regression equation. the result suggests that advanced age, lymphocyte decline, and d-dimer elevation are independent risk factors for patients with severe covid- . p value refers to the test results of each variable in logistic regression analysis; p < . indicates that the variable has an independent correlation with severe covid- . otherwise, there is no independent correlation. the sd of the absolute lymphocyte count is . ; the sd of the ddimer is . . values express mean (sd) or median (iqr). Δly: change in lymphocyte absolute value from day to day after admission; Δalb: change in serum albumin value from day to day after admission; Δd : difference in d-dimer value between day and day after admission; Δd : difference in serum d-dimer value between day and day after admission. pao , pressure of arterial oxygen; fio , fraction of inspired oxygen; apache-ii, acute physiology and chronic health evaluation ii score; sofa, sequential organ failure assessment score. . these results suggested that the d-dimer absolute value increased at a constant rate from day to day . the results showed that in the early stage of covid- , lymphocyte and serum albumin levels decreased but d-dimer levels increased with the progression of the disease. we have included chest ct scans from patient with severe pneumonia at different time periods; it illustrates the rapid progress of the disease (fig. ) . this was a cross-sectional study of the clinical characteristics of patients with covid- , especially those with severe pneumonia. our study suggested that advanced age, lymphocyte decline, and d-dimer elevation were more prominent in the patients with severe pneumonia, which is useful for the early identification of patients likely to develop severe covid- . the laboratory examinations showed that patients with severe pneumonia had depressed serum albumin, while serum creatinine, blood urea nitrogen, aspartate aminotransferase, crp, and b-type natriuretic peptide were all elevated. hypoproteinemia may be due to inadequate protein intake caused by poor appetite. a previous study reported that hypoalbuminemia is a potent, dosedependent predictor of poor outcomes for pneumonia with covid- infection [ ] . an elevated level of crp may be associated with the inflammatory response and cytokine storms caused by sars-cov- in the blood vessels [ ] . these results were consistent with a previous study, which showed that the crp level was positively correlated with the pneumonia severity [ ] . according to the results of the binomial logistic regression analysis, we found that age and d-dimer level were independent risk factors. these results suggested that d-dimer level was significantly positively correlated with severe covid- , also shown in another study [ ] . previous studies showed that sars-cov could bind to ace , downregulating the expression of ace , and resulting in an increased angiotensin ii level in mouse blood samples and increased signaling through angiotensin ii receptor , inducing acute lung injury [ ] [ ] [ ] . ace is a receptor protein of both sars-cov and sars-cov- ; it is abundantly present in the epithelia of the lung and small intestine [ ] . it was reported that sars-cov- binds to ace in the same way as sars-cov [ ] , inducing damage to the pulmonary arteries and leading to the extensive embolization in the alveolar terminal capillaries [ , ] . these changes eventually lead to an increase in d-dimer [ ] . a significant decline in lymphocytes with the progression of severe pneumonia was also observed, consistent with the results of huang et al. [ ] . a lower lymphocyte count suggests that sars-cov- may pri- marily attack the body's immune system; this especially the case with t lymphocytes, and similar to the action of sars-cov [ ] . after sars-cov- impairs the immune system, it is difficult to prevent virus replication by immediately forming the neutralizing antibody. nonspecific pulmonary secondary inflammation is triggered by the sars-cov- infection, inducing a cytokine storm and producing a series of immune responses as well as causing disorders of the lymphocyte subsets. the above factors may explain the decline of lymphocytes and the rise of d-dimer observed with the progression of severe pneumonia. in addition, pulmonary ct findings showed that severe pneumonia progressed rapidly. it is important to review chest ct scans in a timely fashion to learn about potential pulmonary lesions. this study had several limitations. first, only patients from a single hospital were included; a larger-scale study needs to be carried out to confirm our conclusions. second, most of the patients were still hospitalized when the manuscript was submitted, so we could not verify the efficacy of the therapeutic interventions and prognosis of the patients. in general, the results suggested that advanced age, decreased lymphocytes, and elevated levels of d-dimer were risk factors for the severe pneumonia caused by co-vid- . clinicians should pay close attention to these indicators and identify high-risk patients as early as possible. more studies are needed to explore the clinical characteristics and treatment options of critically ill patients. outbreak of pneumonia of unknown etiology in wuhan, china: the mystery and the miracle the continuing -ncov epidemic threat of novel coronaviruses to global health -the latest novel coronavirus outbreak in wuhan, china coronavirus infections -more than just the common cold a novel coronavirus from patients with pneumonia in china genomic characterisation and epidemiology of novel 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patients with novel coronavirus-infected pneumonia in wuhan, china angiotensin ii plasma levels are linked to disease severity and predict fatal outcomes in h n -infected patients angiotensin-converting enzyme protects from severe acute lung failure. nature angiotensin-converting enzyme protects from lethal avian influenza a h n infections tissue distribution of ace protein, the functional receptor for sars coronavirus. a first step in understanding sars pathogenesis discovery of a novel coronavirus associated with the recent pneumonia outbreak in humans and its potential bat origin immunothrombosis in acute respiratory distress syndrome: cross talks between inflammation and coagulation the respiratory system in autoimmune vascular diseases. respiration diagnostic value of ddimer in pulmonary embolism and pneumonia clinical features of patients infected with novel coronavirus in wuhan change of t lymphocyte and its activated subsets in sars patients. zhongguo yi xue ke xue yuan xue bao we thank letpub (www.letpub.com) for its linguistic assistance during the preparation of this manuscript. the study was approved by the ethics committee of wuhan central hospital (yuan lun han [ ] no. ). as this was a retrospective study, only clinical data of patients were collected; privacy data such as name, id number, and telephone number were not involved, so no informed consent was obtained. moreover, the data were only used for scientific research, not for other purposes. all authors report no conflicts of interest. the research received no funding. y.f.w. and y.z. had full access to all data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis. study concept and design: y.f.w. and s.g. acquisition, analysis, or interpretation of data: y. key: cord- -p jp fiq authors: lalloo, david g.; shingadia, delane; bell, david j.; beeching, nicholas j.; whitty, christopher j.m.; chiodini, peter l. title: uk malaria treatment guidelines date: - - journal: j infect doi: . /j.jinf. . . sha: doc_id: cord_uid: p jp fiq .malaria is the tropical disease most commonly imported into the uk, with – cases reported each year, and – deaths. . approximately three quarters of reported malaria cases in the uk are caused by plasmodium falciparum, which is capable of invading a high proportion of red blood cells and rapidly leading to severe or life-threatening multi-organ disease. . most non-falciparum malaria cases are caused by plasmodium vivax; a few cases are caused by the other species of plasmodium: plasmodium ovale, plasmodium malariae or plasmodium knowlesi. . mixed infections with more than one species of parasite can occur; they commonly involve p. falciparum with the attendant risks of severe malaria. . there are no typical clinical features of malaria; even fever is not invariably present. malaria in children (and sometimes in adults) may present with misleading symptoms such as gastrointestinal features, sore throat or lower respiratory complaints. . a diagnosis of malaria must always be sought in a feverish or sick child or adult who has visited malaria-endemic areas. specific country information on malaria can be found at http://travelhealthpro.org.uk/. p. falciparum infection rarely presents more than six months after exposure but presentation of other species can occur more than a year after exposure. . management of malaria depends on awareness of the diagnosis and on performing the correct diagnostic tests: the diagnosis cannot be excluded until more than one blood specimen has been examined. other travel related infections, especially viral haemorrhagic fevers, should also be considered. . the optimum diagnostic procedure is examination of thick and thin blood films by an expert to detect and speciate the malarial parasites. p. falciparum and p. vivax (depending upon the product) malaria can be diagnosed almost as accurately using rapid diagnostic tests (rdts) which detect plasmodial antigens. rdts for other plasmodium species are not as reliable. . most patients treated for p. falciparum malaria should be admitted to hospital for at least h as patients can deteriorate suddenly, especially early in the course of treatment. in specialised units seeing large numbers of patients, outpatient treatment may be considered if specific protocols for patient selection and follow up are in place. . uncomplicated p. falciparum malaria should be treated with an artemisinin combination therapy (grade a). artemether–lumefantrine (riamet(®)) is the drug of choice (grade c) and dihydroartemisinin-piperaquine (eurartesim(®)) is an alternative. quinine or atovaquone–proguanil (malarone(®)) can be used if an act is not available. quinine is highly effective but poorly-tolerated in prolonged treatment and should be used in combination with an additional drug, usually oral doxycycline. . severe falciparum malaria, or infections complicated by a relatively high parasite count (more than % of red blood cells parasitized) should be treated with intravenous therapy until the patient is well enough to continue with oral treatment. severe malaria is a rare complication of p. vivax or p. knowlesi infection and also requires parenteral therapy. . the treatment of choice for severe or complicated malaria in adults and children is intravenous artesunate (grade a). intravenous artesunate is unlicensed in the eu but is available in many centres. the alternative is intravenous quinine, which should be started immediately if artesunate is not available (grade a). patients treated with intravenous quinine require careful monitoring for hypoglycemia. . patients with severe or complicated malaria should be managed in a high-dependency or intensive care environment. they may require haemodynamic support and management of: acute respiratory distress syndrome, disseminated intravascular coagulation, acute kidney injury, seizures, and severe intercurrent infections including gram-negative bacteraemia/septicaemia. . children with severe malaria should also be treated with empirical broad spectrum antibiotics until bacterial infection can be excluded (grade b). . haemolysis occurs in approximately – % patients following intravenous artesunate treatment. haemoglobin concentrations should be checked approximately days following treatment in those treated with iv artemisinins (grade c). . falciparum malaria in pregnancy is more likely to be complicated: the placenta contains high levels of parasites, stillbirth or early delivery may occur and diagnosis can be difficult if parasites are concentrated in the placenta and scanty in the blood. . uncomplicated falciparum malaria in the second and third trimester of pregnancy should be treated with artemether–lumefantrine (grade b). uncomplicated falciparum malaria in the first trimester of pregnancy should usually be treated with quinine and clindamycin but specialist advice should be sought. severe malaria in any trimester of pregnancy should be treated as for any other patient with artesunate preferred over quinine (grade c). . children with uncomplicated malaria should be treated with an act (artemether–lumefantrine or dihydroartemisinin-piperaquine) as first line treatment (grade a). quinine with doxycycline or clindamycin, or atovaquone–proguanil at appropriate doses for weight can also be used. doxycycline should not be given to children under years. . either an oral act or chloroquine can be used for the treatment of non-falciparum malaria. an oral act is preferred for a mixed infection, if there is uncertainty about the infecting species, or for p. vivax infection from areas where chloroquine resistance is common (grade b). . dormant parasites (hypnozoites) persist in the liver after treatment of p. vivax or p. ovale infection: the only currently effective drug for eradication of hypnozoites is primaquine ( a). primaquine is more effective at preventing relapse if taken at the same time as chloroquine (grade c). . primaquine should be avoided or given with caution under expert supervision in patients with glucose- -phosphate dehydrogenase deficiency (g pd), in whom it may cause severe haemolysis. . primaquine (for eradication of p. vivax or p. ovale hypnozoites) is contraindicated in pregnancy and when breastfeeding (until the g pd status of child is known); after initial treatment for these infections a pregnant woman should take weekly chloroquine prophylaxis until after delivery or cessation of breastfeeding when hypnozoite eradication can be considered. . an acute attack of malaria does not confer protection from future attacks: individuals who have had malaria should take effective anti-mosquito precautions and chemoprophylaxis during future visits to endemic areas. . management of malaria depends on awareness of the diagnosis and on performing the correct diagnostic tests: the diagnosis cannot be excluded until more than one blood specimen has been examined. other travel related infections, especially viral haemorrhagic fevers, should also be considered. . the optimum diagnostic procedure is examination of thick and thin blood films by an expert to detect and speciate the malarial parasites. p. falciparum and p. vivax (depending upon the product) malaria can be diagnosed almost as accurately using rapid diagnostic tests (rdts) which detect plasmodial antigens. rdts for other plasmodium species are not as reliable. . most patients treated for p. falciparum malaria should be admitted to hospital for at least h as patients can deteriorate suddenly, especially early in the course of treatment. in specialised units seeing large numbers of patients, outpatient treatment may be considered if specific protocols for patient selection and follow up are in place. . uncomplicated p. falciparum malaria should be treated with an artemisinin combination therapy (grade a). artemetherelumefantrine (riamet Ò ) is the drug of choice (grade c) and dihydroartemisinin-piperaquine (eurartesim Ò ) is an alternative. quinine or atovaquone eproguanil (malarone Ò ) can be used if an act is not available. quinine is highly effective but poorly-tolerated in prolonged treatment and should be used in combination with an additional drug, usually oral doxycycline. . severe falciparum malaria, or infections complicated by a relatively high parasite count (more than % of red blood cells parasitized) should be treated with intravenous therapy until the patient is well enough to continue with oral treatment. severe malaria is a rare complication of p. vivax or p. knowlesi infection and also requires parenteral therapy. . the treatment of choice for severe or complicated malaria in adults and children is intravenous artesunate (grade a). intravenous artesunate is unlicensed in the eu but is available in many centres. the alternative is intravenous quinine, which should be started immediately if artesunate is not available (grade a). patients treated with intravenous quinine require careful monitoring for hypoglycemia. . patients with severe or complicated malaria should be managed in a high-dependency or intensive care environment. they may require haemodynamic support and management of: acute respiratory distress syndrome, disseminated intravascular coagulation, acute kidney injury, seizures, and severe intercurrent infections including gram-negative bacteraemia/septicaemia. . children with severe malaria should also be treated with empirical broad spectrum antibiotics until bacterial infection can be excluded (grade b). . haemolysis occurs in approximately e % patients following intravenous artesunate treatment. haemoglobin concentrations should be checked approximately days following treatment in those treated with iv artemisinins (grade c). . falciparum malaria in pregnancy is more likely to be complicated: the placenta contains high levels of parasites, stillbirth or early delivery may occur and diagnosis can be difficult if parasites are concentrated in the placenta and scanty in the blood. . uncomplicated falciparum malaria in the second and third trimester of pregnancy should be treated with artemetherelumefantrine (grade b). uncomplicated falciparum malaria in the first trimester of pregnancy should usually be treated with quinine and clindamycin but specialist advice should be sought. severe malaria in any trimester of pregnancy should be treated as for any other patient with artesunate preferred over quinine (grade c). . children with uncomplicated malaria should be treated with an act (artemether elumefantrine or dihydroartemisinin-piperaquine) as first line treatment (grade a). quinine with doxycycline or clindamycin, or atovaquoneeproguanil at appropriate doses for weight can also be used. doxycycline should not be given to children under years. . either an oral act or chloroquine can be used for the treatment of non-falciparum malaria. an oral act is preferred for a mixed infection, if there is uncertainty about the infecting species, or for p. vivax infection from areas where chloroquine resistance is common (grade b). . dormant parasites (hypnozoites) persist in the liver after treatment of p. vivax or p. ovale infection: the only currently effective drug for eradication of hypnozoites is primaquine ( a). primaquine is more effective at preventing relapse if taken at the same time as chloroquine (grade c). . primaquine should be avoided or given with caution under expert supervision in patients with glucose- -phosphate dehydrogenase deficiency (g pd), in whom it may cause severe haemolysis. . primaquine (for eradication of p. vivax or p. ovale hypnozoites) is contraindicated in pregnancy and when breastfeeding (until the g pd status of child is known); after initial treatment for these infections a pregnant woman should take weekly chloroquine prophylaxis until after delivery or cessation of breastfeeding when hypnozoite eradication can be considered. . an acute attack of malaria does not confer protection from future attacks: individuals who have had malaria should take effective anti-mosquito precautions and chemoprophylaxis during future visits to endemic areas. ª the british infection association. published by elsevier ltd. all rights reserved. malaria remains one of the most common imported infections in the united kingdom (uk). between and malaria cases are reported each year in the uk, although reviews of reporting suggest that this may represent about % of all cases that occur. approximately three-quarters of reported infections are due to plasmodium falciparum and there are between and deaths annually. children under account for around % of cases. two thirds of cases occur in people of african or south asian ethnic origin and over half of the cases occur in those who had been visiting friends and family in endemic areas. most patients with falciparum malaria acquire infection in africa and malaria accounts for about % of travellers from africa presenting to hospital with fever; west africa is the commonest geographical source. most plasmodium vivax infections are acquired in south asia. , this document offers guidance for the management of both uncomplicated and complicated malaria in the uk. it complements existing public health england (phe) guidelines on the prevention of malaria in uk travellers. https:// www.gov.uk/government/publications/malariaprevention-guidelines-for-travellers-from-the-uk. it has been based on a review of the available evidence by the phe advisory committee on malaria prevention (acmp), with input from other experts and expert bodies, and incorporates international guidance including who guidelines on treatment and definitions of severe malaria. , these guidelines will specifically present a uk perspective on management. other non-endemic countries including the usa, canada and europe have also developed their own guidelines. e these uk guidelines have been developed specifically for use in a non-endemic setting, but necessarily depend heavily upon evidence obtained from studies in endemic areas. more detailed information about individual drug regimens and contra-indications can be found in the british national formulary (http://www.bnf.org/). a short summary of key points in the initial assessment and management, for use in emergency departments, is available from the british infection association website (http://www.britishinfection.org/). major recommendations have been graded using a modified grade approach which grades both the strength of the recommendation and the level of evidence for the recommendations. a grade recommendation is a strong recommendation to do (or not do) something, where benefits clearly outweigh risks (or vice versa) for most patients. a grade recommendation is a conditional recommendation, where the risks and benefits are more closely balanced or are more uncertain. grade a evidence means high-quality evidence that comes from consistent results from high quality randomised controlled trials (rcts). grade b evidence means moderate-quality evidence from randomised trials that suffers from flaws in conduct or design or methodologically strong observational studies with consistent effects and exclusion of most sources of bias. grade c evidence is low-quality evidence from controlled trials with serious limitations or inconsistent results, or observational studies with limited evidence on effects. grade d evidence is based only on case studies or expert judgement or poor quality observational studies. assessment of the patient with suspected malaria (boxes and ) the crucial issue in the management of malaria is consideration of the possibility of this diagnosis. malaria should be suspected in anyone with a fever or a history of fever who has returned from or previously visited a malaria endemic area, regardless of whether they have taken prophylaxis. the minimum incubation period for naturally acquired infection is six days. most patients with falciparum infection present in the first month or months after exposure; almost all present within six months of exposure. vivax or ovale infections commonly present later than six months after exposure and presentation may be delayed for years. there are no specific symptoms of malaria: most patients complain of fever, headache and general malaise. gastrointestinal disturbances, jaundice or respiratory symptoms occasionally occur and are often responsible for misdiagnosis. most missed malaria infections are erroneously diagnosed as non-specific viral infections, influenza, gastroenteritis or hepatitis. children are less likely than adults to complain of chills, arthralgia/myalgia or headaches and more likely to present with non-specific symptoms (fever, lethargy, malaise, somnolence): gastrointestinal symptoms (nausea, abdominal pain, vomiting, diarrhoea) are particularly common. , the physical examination of patients with uncomplicated malaria is often unremarkable apart from a fever which is not invariably present. most patients have no specific fever pattern. children are more likely to have hepatomegaly, splenomegaly and somnolence than adults. , if the diagnosis of falciparum malaria has been delayed, severely ill patients may present with jaundice, confusion or seizures. if malaria is suspected, a blood test for malaria without delay is mandatory. unless rapid malaria testing can be achieved in primary care, the patient should be referred to hospital for testing. results should be communicated the same day: all positive tests should be telephoned back to the requesting doctor as soon as practicable and ideally within h of the test reaching the laboratory. the most important test is examination of thick and thin blood smears by microscopy. this is highly sensitive and specific in expert hands. however, because of a lack of expertise in many uk labs, particularly out of hours, rapid diagnostic tests (rdts) based upon detection of parasite antigens, are now commonly used in addition to blood slides. although slightly less sensitive than good quality blood films examined by experienced microscopists, they are easier for the non-expert to use to detect falciparum infections and are useful as an initial screen if expertise in reading slides is not immediately available. rdts are generally not as specific and sensitive for the detection of non-falciparum infections, although newer generation rdts perform well in diagnosing vivax infection. , rdts may be used in addition to, but not as a replacement for blood films and all patients with suspected malaria should have blood films prepared and examined as recommended in the british committee for standards in haematology guidelines. if falciparum or knowlesi malaria is diagnosed, the percentage of red blood cells that are parasitized should be estimated. if there is clinical suspicion of malaria, but initial blood films are negative, repeat films, with or without rdt, should be examined after e h and again after a further h. rdt use alone should be discouraged where good microscopy is available. thrombocytopaenia is suggestive of malaria in non-immune adults and children, both in nonfalciparum malarias and in p. falciparum, , although it can also occur in a number of other imported infections. malaria is unlikely if three negative specimens have been examined by a competent microscopist. empirical therapy for malaria should not be given unless a patient with a convincing exposure history demonstrates features of severe malaria and expert advice has been taken. in pregnancy, thick films can be negative, despite the presence of parasites in the placenta. expert advice should be sought if malaria is suspected. pcr techniques are used in reference laboratories to determine the species of malaria, but are not sufficiently standardised or validated to use for routine clinical diagnosis. cases of malaria should be notified to public health authorities. in england and wales, thick and thin films and a blood aliquot should be sent to the malaria reference laboratory for confirmation (which is performed free of charge). the scottish parasite and diagnostic reference laboratory provides a reference service for scotland. if malaria is diagnosed in a returned traveller, other members of the family or travelling group should be warned that they may have shared the same exposure risk and that they should seek medical attention if they develop symptoms. malaria, particularly severe malaria, should always be managed in consultation with someone experienced in managing the disease. non-falciparum malaria the distinction between falciparum malaria and other species of malaria is important. malaria caused by plasmodium ovale, p. vivax, and plasmodium malariae rarely causes life-threatening disease and can usually be managed on an outpatient basis, unless the patient has other comorbidities or cannot tolerate oral medication. however, severe presentations of plasmodium knowlesi infection and p. vivax are well recognised and severe p. ovale can occur in exceptional circumstances and clinicians should look out for these rare cases. e box . important considerations in the assessment of a patient with possible malaria. malaria is a medical emergency and patients with suspected malaria should be evaluated immediately. symptoms of malaria are often non-specific: fever/sweats/chills, malaise, myalgia, headache, diarrhoea, and cough. falciparum malaria is most likely to occur within months of return from an endemic area. the incubation period for malaria is at least days. malaria caused by other species may present more than a year after return from an endemic area. a careful exposure history is necessary: country and area of travel, including stopovers, and date of return. consider what malaria prophylaxis was taken (i.e. drug, dose & adherence, premature cessation); appropriate prophylaxis with full adherence does not exclude malaria. consider other travel-related infections: e.g. typhoid, hepatitis, dengue or other arboviruses, avian influenza, mers-cov, hiv, meningitis/encephalitis and viral haemorrhagic fevers (vhf). three negative diagnostic samples over a period of e h are necessary to exclude malaria. estimation of the haemoglobin concentration should be done, and in malaria caused by p. vivax or p. ovale, glucose- -phosphate dehydrogenase (g pd) activity should be measured, as concomitant primaquine therapy will be necessary to eliminate hypnozoites (dormant forms) from the liver. primaquine can cause haemolysis in patients with g pd deficiency. patients with a mixed infection that includes falciparum parasites or with an infection with an unidentified species should be treated as though they had falciparum infection in the first instance. patients with falciparum malaria should usually be admitted to hospital initially because of the risk of deterioration even after effective treatment has been initiated. there is limited evidence that some patients with falciparum malaria can be managed safely as outpatients in units that see large numbers of patients and use well defined protocols for assessment and follow up. e certain factors such as age, ethnicity and parasite count can predict the likelihood of severe malaria, but even patients who might be expected to be semi-immune may deteriorate rapidly and require intensive care treatment. e outpatient management of malaria in adults should only be undertaken by clinicians experienced in managing malaria with clear protocols and systems for assessing the likely risk of severe malaria and for rapidly reassessing patients. children with falciparum malaria should be observed in hospital initially for at least h, because of the possibility of rapid progression and also to ensure that they are tolerating oral therapies; treatment in children may be complicated by vomiting. , patients should be observed closely; certain categories such as pregnant women, infants and the elderly are more likely to develop severe disease or to deteriorate rapidly. , , the management of patients with falciparum malaria, especially if severe, should always be discussed with a specialist; mortality is higher in regions of the uk where malaria is less commonly managed. % of uk malaria cases are seen in centres with less than cases a year. patients with falciparum malaria (or a mixed infection which includes falciparum parasites) can be divided into those with uncomplicated and those with severe or complicated disease (box ). assessment of the patient should include careful clinical evaluation and review of investigations for the features of severe malaria detailed below. a full blood count, urea, creatinine and electrolytes, liver function tests and blood glucose should be done routinely. thrombocytopaenia is common and in isolation does not reflect severe disease. in ill patients, blood gases, blood culture, lactate and clotting studies should be also performed. urine dipstick and culture, stool culture and chest x-ray may be appropriate. lumbar puncture to exclude meningitis should be considered in febrile patients with impaired consciousness or repeated seizures. the initial parasite count is helpful in estimating the potential future severity of disease. although highly dependent upon the stage of the infection, if more than % of red blood cells are parasitized, there is an increased chance of developing severe disease even if the patient initially box . common errors in diagnosis or management of malaria (adapted from beeching nj et al. with permission). delayed patient presentation. failure of health care worker to take a travel history or consider diagnosis of malaria. belief that chemoprophylaxis prevents all malaria. belief that malaria is unlikely if patient does not remember being bitten by mosquitoes. belief that malaria presents with a classical fever pattern. failure to recognise nonspecific clinical presentations of malaria. failure to obtain immediate blood films or rdt. failure to repeat diagnostic tests if first tests are negative. failure to prescribe adequate and appropriate chemotherapy immediately. failure to anticipate or treat complications. impaired consciousness or seizures. renal impairment (oliguria < . ml/kg bodyweight per hour or creatinine > mmol/l). acidosis (ph < . ). hypoglycemia (< . mmol/l). pulmonary oedema or acute respiratory distress syndrome (ards). haemoglobin g/l. spontaneous bleeding/disseminated intravascular coagulation. shock (algid malaria e bp < / mmhg). haemoglobinuria (without g pd deficiency). parasitaemia > %. appears well, and a % parasitaemia is considered to represent severe disease. other important poor prognostic factors are: the presence of peripheral blood schizonts of p. falciparum, pigment deposits in peripheral polymorphonuclear leucocytes on the blood film, , metabolic acidosis or an elevated lactate level, e older age, coma and renal impairment. , treatment of uncomplicated falciparum malaria in adults there are now three main therapeutic options for the treatment of uncomplicated falciparum malaria in adults in the uk: artemisinin combination therapy (act), oral atovaquoneeproguanil or quinine plus doxycycline (or quinine plus clindamycin in certain circumstances) (see box for details of doses). two acts are licenced for use in the uk; artemetherelumefantrine or dihydroartemisininpiperaquine. although mefloquine is an effective treatment, the side effects and high rate of non-completion of courses means that we do not recommend this as therapy in the uk. acts are highly effective and clear parasites more rapidly than other options because they are effective throughout a broader range of the parasite life cycle. for this reason, they are now considered to be the drugs of choice in uncomplicated malaria (grade a). studies comparing acts with oral quinine in africa have demonstrated greater efficacy using acts, driven partly by poor compliance with quinine. there has been most experience with artemetherelumefantrine in a western setting and this seems to be well tolerated. , artemetherelumefantrine is ideally taken with a high fat meal to maximise absorption and is considered the act of choice (grade c). both acts need to be given for only three days. although dha piperaquine (dha-ppq) has the advantage of only needing single daily dosing, there has been concern about the potential for qtc prolongation with dha-ppq. until further data become available, it is recommended that dha-ppq is taken more than h after food and that patients should not eat for h after doses of dhapiperaquine to prevent excessive peak piperaquine levels. dha-ppq should not currently be used in patients with previous arrhythmias, cardiac conditions that predispose to arrhythmia, or those taking drugs that prolong the qt interval. ecgs should be obtained early in the course of treatment with dha-ppq and before and after the last daily dose of dha-ppq; more frequent monitoring may be indicated in those taking drugs which inhibit cyp a and potentially increase piperaquine levels, such as clarithromycin. atovaquoneeproguanil (malarone Ò ) has been used extensively in some western settings with high levels of efficacy, although parasite clearance is relatively slow ( % at three days). , almost a quarter of patients experienced gastro-intestinal side-effects and patients should be warned about these to ensure full adherence. in contrast to the three day regimens for acts and atovaquoneeproguanil, quinine needs to be taken for five to seven days, or until parasites have cleared, which requires daily monitoring. quinine is often associated with "cinchonism" (nausea, deafness and ringing in the ears), which may result in poor adherence. although international recommendations suggest that quinine should be taken for seven days in endemic areas, uk experience suggests that five days treatment is adequate for the vast majority of cases when combined with a second drug (doxycycline for adults or clindamycin in pregnant women and young children) to ensure complete eradication of parasites. the second drug can be taken either simultaneously with quinine or sequentially after the quinine. in view of increasing failure rates of anti-folate drugs in most part of the world, sulfadoxine-pyrimethamine (fansidar Ò ) should not be used routinely as a second drug to accompany quinine, except on specialist advice. chloroquine should not be used for the treatment of falciparum malaria. antibiotics, including tetracyclines, sulfa drugs, macrolides and clindamycin, should only be used in combination therapies and not used alone for the treatment of malaria. there is insufficient evidence to support the use of azithromycin either alone or in combination with other drugs for the treatment of malaria. treatment of severe or complicated falciparum malaria antimalarial therapy urgent appropriate parenteral therapy with antimalarials has the greatest impact on prognosis in severe malaria. treatment should not be delayed in patients with proven or strongly suspected malaria. parenteral treatment is indicated in all patients with severe or complicated malaria, those at high risk of developing severe disease (box ) or if the patient is vomiting and unable to take oral antimalarials. there is now substantial evidence for the superiority of intravenous artesunate over intravenous quinine with two large trials and a meta-analysis combining with other smaller trials demonstrating a mortality benefit in adults and children e for all patients with severe malaria. intravenous artesunate is therefore considered the treatment of choice for severe malaria. (grade a). the manufacturers of intravenous artesunate have not achieved good manufacturing practice (gmp) certification and artesunate is not licenced in the european union ( ). however, the factory manufacturing artesunate has achieved who prequalification standards and some importers of artesunate also carry out quality checks on imported batches. intravenous artesunate is now stocked by many infectious diseases units in the uk and can also be obtained from specialist tropical disease centres in london and liverpool (see below for contact details). we recommend that any centre regularly seeing patients with severe malaria should stock artesunate. however, treatment should never be delayed whilst obtaining artesunate: every patient with severe malaria should start quinine initially if artesunate is not immediately available as delay is very dangerous (grade a). there is no additional benefit from using artesunate in combination with quinine but it is safe to do so. following a minimum of h of intravenous artesunate, and once patients have improved and are able to take oral medication, a full course of an act (artemetherelumefantrine or dha-ppq) should be given. alternatively, a full course of quinine and doxycycline (clindamycin in children/ pregnant women) or atovaquoneeproguanil could be used. clearance of parasites should be checked by daily thick blood films. there has been increasing experience with the use of intravenous artesunate in western non-immune travellers, demonstrating rapid effectiveness and generally low adverse event rates compared to quinine. , however, there is clear emerging evidence of delayed haemolysis ( e days post treatment) following intravenous artesunate in approximately e % of adults or children, especially those with high parasite counts. e this appears to be self-limiting but patients should be warned to be aware of potential symptoms of anaemia and their haemoglobin level should be routinely checked approximately days after completing artesunate (grade c). the emerging evidence of reduced susceptibility to artemisinin derivatives in se asia does not alter our current recommendations for the use of artesunate as first line therapy for this region (grade c). however, as with all cases of severe malaria, monitoring of clearance of parasites and for recurrence of symptoms is recommended. artemether is an oil-based intramuscular artemisinin preparation, which is produced to gmp standards. however, despite generally favourable trends, several studies and meta-analyses have not shown a clear advantage of artemether over quinine in the management of severe malaria in either adults or children e and we do not recommend its use (grade b). intravenous quinine dihydrochloride is an alternative if artesunate is not immediately available. it should be given as an intravenous infusion with an initial loading dose of mg/kg in % dextrose or dextrose/saline over h to achieve high blood levels rapidly (see box ) . this should be followed by mg/kg infused over h every h. a loading dose should not be given if quinine or mefloquine therapy has been taken within the previous h (grade c). caution should be exercised in older patients or those with cardiac disease, because of the potential for quinine to lead to arrhythmias. these patients should have ecg monitoring during intravenous quinine treatment. when the patient is well enough to take oral medication, treatment should be completed with a full course of an oral box . other indications for parenteral therapy in adults. parasitaemia > % red blood cells parasitized. pregnant women following specialist advice. patients unable to swallow/retain tablets. box . drug treatment of severe or complicated malaria. artesunate regimen: . mg/kg given as an intravenous injection at , and h then daily thereafter. after completion of a minimum of h therapy (maximum five days), a full course of an oral act should be taken when the patient can tolerate oral medication. quinine: loading dose of mg/kg quinine dihydrochloride in % dextrose or dextrose saline over h. followed by mg/kg every h for first h (or until patient can swallow). frequency of dosing should be reduced to hourly if intravenous quinine continues for more than h. parenteral quinine therapy should be continued until the patient can take oral therapy when quinine sulphate mg should be given three times a day to complete five to seven days of quinine in total. quinine treatment should always be accompanied by a second drug: doxycycline mg (or clindamycin mg three times a day for children or pregnant women), given orally for total of seven days from when the patient can swallow. agent (box ). if quinine is used, a total combined iv and oral course of seven days is appropriate. if intravenous quinine needs to be continued for longer than h, or the patient is in renal failure or has severe hepatic dysfunction, quinine doses should be reduced by a third. , neither quinine nor artesunate levels are affected by haemofiltration; dose modification is not necessary. all patients with severe or complicated malaria should be managed in a high dependency unit. patients may deteriorate rapidly and close observation is vital. transfer to an intensive care unit should be considered for those with severe acidosis, high lactate levels, pulmonary oedema/acute respiratory distress syndrome, complicated fluid balance problems or renal impairment and those deteriorating despite appropriate treatment (box ). careful fluid balance is important to avoid over-filling, which may exacerbate the increased pulmonary capillary permeability that occurs in severe malaria (grade c). there is evidence that measurement of the central venous pressure is not useful in predicting true volume status in severe malaria and much of the lactic acidosis seen in severe malaria may be due to microvascular obstruction rather than hypovolaemia. hypoglycemia may occur in severe malaria, complicated by quinine-induced hyperinsulinemia which may develop late in the clinical course, even after the patient appears to be recovering. , blood glucose levels (using a "stix" method) should be checked routinely every h (grade c), two-hourly during quinine infusion (grade c) and at any time that reduced consciousness occurs (grade a). infusion of % dextrose may be necessary to correct hypoglycemia. haemoglobin, clotting, electrolytes (including calcium and sometimes magnesium) and renal function should be closely monitored. frequent parasite counts are not helpful in the early management of severe malaria; the peripheral parasite count will fluctuate according to the stage of parasite development and it is not uncommon for the parasite count to increase in the first e h of treatment: this does not indicate failure of therapy. daily parasite counts are sufficient and are recommended to ensure clearance. some patients develop shock which may be secondary to complicating bacteraemia/septicaemia ("algid malaria"). patients with signs of shock should be treated with a broad spectrum antibiotic (grade c). platelet transfusion is not indicated even with low counts unless there is active bleeding. appropriate ventilatory support or renal replacement therapy should be initiated if clinically indicated: haemofiltration appears to be superior to peritoneal dialysis. patients with impaired consciousness or coma should be managed appropriately. corticosteroids, mannitol, n-acetylcysteine and levamisole have all been shown to be ineffective as adjunctive therapies for the treatment of severe malaria. e the role of exchange transfusion in the management of severe malaria has always been controversial, with no clear evidence of benefit and potential risks, especially in individuals who may have haemodynamic instability. , artesunate has its greatest mortality advantage in those with high parasite counts. the rapid action of artesunate in reducing parasite burden means that any benefit of exchange transfusion is likely to be substantially reduced. current opinion is that exchange transfusion is now no longer indicated in severe malaria (grade c). routine blood transfusion may be indicated in those with symptomatic anaemia. there is clear evidence that risks of both falciparum and vivax malaria leading to mortality increase steadily with age over . all elderly patients should be admitted, and monitored closely. malaria in pregnancy carries a higher risk of severe disease and is also associated with miscarriages or stillbirths. pregnant women with malaria require prompt treatment and should be managed in collaboration with the obstetric team. close observation in hospital, including uterine and foetal heart monitoring for development of complications, is necessary and early delivery of a near-term infant at risk may need to be considered. the rcog has produced specific guidelines for the treatment of malaria in pregnancy. uncomplicated falciparum malaria. neither artemetherelumefantrine (riamet Ò ), atovaquoneeproguanil (malarone Ò ) or dha-ppq are licenced in pregnancy. there is no evidence of adverse effects of artemetherelumefantrine in the second and third trimester from a limited number of studies. data on safety in the first trimester are even more limited but no evidence of harm has been detected. small studies on atovaquoneeproguanil have shown no evidence of adverse effects. artemetherelumefantrine is considered the treatment of choice in the second and third trimester (grade b) (information on dha-ppq is currently more limited). quinine (seven days) in combination with clindamycin (see dose above) can be used in all box . intensive care management of severe or complicated malaria. careful management of fluid balance to optimise oxygen delivery and prevent pulmonary oedema. regular monitoring for hypoglycemia. consider broad spectrum antibiotics if evidence of shock or secondary bacterial infection. haemofiltration for renal failure or control of acidosis or fluid/electrolyte imbalance. consider medication to control seizures. three trimesters. quinine can increase the risk of uterine contraction and hypoglycemia. severe malaria. severe malaria in pregnancy is associated with high case fatality rates, pregnancy loss and hypoglycemia and pulmonary oedema are particularly common. there is little published evidence of the use or safety of intravenous artesunate in pregnant women, particularly in the first trimester. on balance of risk, artesunate is preferred to quinine on the basis of its likely higher effectiveness in reducing mortality (grade c). intravenous quinine (with clindamycin) is an alternative. oral quinine, atovaquoneeproguanil (malarone Ò ), artemetherelumefantrine and dha-ppq can all be used for the treatment of uncomplicated malaria in children (box ). there is limited experience in the use of artemisinin combination therapies in a non-endemic paediatric population, although acts are recommended as first-line treatment of uncomplicated malaria in children in malaria endemic regions. despite this, on the basis of evidence from endemic areas, the committee believes that an act should be first line therapy for children in the uk (grade a). the combination of oral quinine with seven days of clindamycin or doxycycline (for children greater than years age) remains highly effective in the uk, with very low relapse rates in children. in contrast to the views of some authors, , we believe that oral quinine is usually well-tolerated by children and should be used for the treatment of uncomplicated falciparum malaria in the uk if an act is not available. , tetracyclines should not be given to children under years of age because of risk of dental hypoplasia and permanent discolouration of teeth. severe and complicated falciparum malaria in children (box ) the main clinical presentations of severe malaria in children are cerebral malaria, severe anaemia and respiratory distress/acidosis. features of cerebral malaria include depressed conscious level, seizures, altered respiration and posturing (decorticate or decerebrate). hypoglycemia, metabolic acidosis, circulatory shock and electrolyte disturbance may also be present. prostration (the inability to stand or sit) is also an indicator of severe disease in children. management of severe or complicated malaria in children involves emergency assessment and provision of supportive care including respiratory and cardiovascular support as outlined by maitland et al. children with severe or complicated malaria should be managed in a paediatric intensive care unit or high dependency unit together with support/advice from a paediatric infectious diseases/tropical medicine specialist who has experience in managing malaria. judicious and slow volume resuscitation is important in those children presenting with shock; the feast trial showed a detrimental effect of routine fluid bolus administration. hypoglycemia is a common complication of severe malaria; serial blood glucose estimations must be performed, and hypoglycemia corrected using e % glucose in maintenance fluid. as it is often difficult to exclude or differentiate concurrent bacterial septic shock or meningitis from severe malaria, empirical broad spectrum antibiotics should be given to children with severe malaria until bacterial infection can be excluded (grade b). management of seizures should follow the evidence-based guidelines advocated by the advanced paediatric life support group. blood transfusions may be required for severe anaemia, although a cochrane review found that routine transfusion did not reduce mortality, but caused more adverse events. clear evidence from a large randomised trial now shows that although quinine remains effective, artesunate is associated with a survival advantage (relative risk reduction of . %) and a significant reduction in clinical complications (development of coma, convulsions and deterioration of coma score). in line with the who guidelines, we recommend that iv artesunate should be used preferentially over quinine as the drug of choice for treatment of severe falciparum malaria in children (grade a). recent who guidelines suggest that a higher dose is required for children under kg. intravenous quinine is still indicated if artesunate is not immediately available and treatment should not be delayed whilst awaiting artesunate therapy. treating the acute infection the treatment of non-falciparum malaria consists of treating the erythrocytic asexual forms that cause symptoms and, for infections with p. vivax and p. ovale, also ensuring eradication of liver hypnozoites to prevent relapse of infection (box ). if a mixed infection of either vivax or ovale with falciparum has been treated, there is no need for an additional drug to treat the blood forms of nonfalciparum infection, but relapse due to the liver forms will still need to be prevented. either an oral act or chloroquine ( mg/kg in total over days) can be used to treat the blood forms of all non-falciparum species (grade b). fever and parasite clearance times are faster with most act regimens than chloroquine in non-falciparum malaria; most of the evidence comes from studies in vivax. , chloroquine is highly effective against p. malariae, p. ovale and p. knowlesi and is effective in most cases of vivax malaria. chloroquine resistance leading to poor clinical outcomes of chloroquine treatment has been recognised in vivax malaria since . this is an uncommon but increasing problem, particularly in the regions of papua new guinea and indonesia. , , chloroquine can still be used for vivax infections from such regions with appropriate follow-up but an act may be preferred (grade b). act regimens should be first line therapy if falciparum malaria cannot be reliably excluded or for treatment of mixed infections that include p. falciparum. they also provide an alternative for individuals with non-falciparum malaria who cannot tolerate chloroquine. cases of severe or complicated non-falciparum malaria should be treated with parenteral artesunate or quinine as for severe falciparum malaria. artesunate appears to be highly effective for severe p. knowlesi infections. chloroquine is a safe option for treatment of nonfalciparum malaria throughout pregnancy. acts can be used in the second and third trimesters. quinine may be used in the first trimester if there is concern about resistant vivax. box . severe or complicated malaria in children. impaired consciousness or seizures. respiratory distress or acidosis (ph < . ). hypoglycemia ( . mmol/l). severe anaemia (< g/dl). prostration. parasitaemia > % red blood cells parasitized. late presentation or relapse due to hypnozoites in the liver occurs in more than % of patients with vivax malaria treated with chloroquine alone. blood schizonticides such as chloroquine, and all other drugs currently used for treating acute malaria, do not eliminate these liver stages, so a second drug is required to achieve "radical" cure. primaquine is the drug of choice for elimination of hypnozoites in ovale or vivax malaria (grade a). patients should be screened for g pd deficiency before primaquine treatment, as primaquine may cause haemolysis in g pd deficient individuals. the efficacy of primaquine in preventing relapse is highly dependent upon co-administration with chloroquine or an alternative drug to clear the red cell forms. , primaquine also has intrinsic activity against asexual blood forms of p. vivax and p. ovale, and concomitant administration with chloroquine boosts blood primaquine levels. , administration of the two drugs should therefore overlap (grade c). in centres that commonly see patients with vivax malaria, systems for rapid assessment of g pd status should be set up. the standard therapeutic dose of mg primaquine base/day for days is appropriate for the radical treatment of p. ovale. however, certain geographical strains of p. vivax have long been recognised to be less sensitive to primaquine and to require higher doses of primaquine to prevent relapse. there has also been increasing evidence of failure of standard dose primaquine from other geographical areas: clinical relapse occurs in the uk in more than % of patients with imported vivax treated with chloroquine followed by unsupervised primaquine mg daily for days. higher dose primaquine mg daily ( . mg/kg) is more effective than mg daily in south asia, the source of most uk infections. , administration of primaquine therapy for less than days is associated with higher relapse rates than day regimens. we therefore recommend that in vivax malaria, primaquine should be given at a dose of mg daily for days to prevent relapse along with treatment with chloroquine.(grade c). in pregnant or breastfeeding women, weekly suppressive chloroquine prophylaxis ( mg each week) should be given until primaquine can be given following delivery or completion of breastfeeding. expert opinion should be sought when treating patients with g pd deficiency. in those with mild to moderate g pd deficiency, alternative regimens of mg ( . mg/kg) primaquine weekly for eight weeks may be effective and safely tolerated e (grade c). in some cases, particularly those who have previously suffered severe adverse effects of primaquine, it may be prudent to withhold primaquine treatment, but to treat relapses promptly. patients and their relatives should be provided with a full explanation of the condition and specific issues related to their therapy, including warnings about possible immediate and late complications of their treatment (box ). although there are no systematic studies on the value of early follow-up of patients following treatment for imported malaria, the committee believes that it is good practice to repeat a blood film and full blood count approximately fourteen days after treatment, particularly if patients have had severe malaria or received artemisinin therapy. it also provides a further opportunity to reinforce the need for appropriate antimalarial precautions to be taken by the patient and his or her relatives next time they travel (https://www.gov.uk/government/ publications/malaria-prevention-guidelines-for-travellersfrom-the-uk). malaria reference laboratory https://www.gov.uk/ government/publications/malaria-reference-laboratorymrl-user-handbook. scottish parasite and diagnostic reference laboratory http://www.nhsggc.org.uk/about-us/professional-supportsites/scottish-microbiology-reference-laboratories/ scottish-parasite-diagnostic-reference-laboratory/ hospital for tropical diseases, uch, london. . tropical and infectious disease unit, royal liverpool university hospital. . estimating unreported malaria cases in england: a capture-recapture study malaria imported into the united kingdom imported malaria and high risk groups: observational study using uk surveillance data e falciparum malaria as a cause of 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chloroquine? chloroquine resistance in plasmodium vivax. antimicrob agents chemother primaquine therapy for malaria potentiation of the curative action of primaquine in vivax malaria by quinine and chloroquine pharmacokinetic interactions between primaquine and chloroquine. antimicrob agents chemother plasmodium vivax chesson strain treatment of plasmodium vivax malariaetime for a change? primaquine: report from cdc expert meeting on malaria chemoprophylaxis i primaquine for preventing relapse in people with plasmodium vivax malaria treated with chloroquine mitigation of the haemolytic effect of primaquine and enhancement of its activity against exoerythrocytic forms of the chesson strain of plasmodium vivax tolerability and safety of weekly primaquine against relapse of plasmodium vivax in cambodians with glucose- -phosphate dehydrogenase deficiency a randomised trial of an eight-week, once weekly primaquine regimen to prevent relapse of plasmodium vivax in northwest frontier province chemotherapy of malaria revised nd edition plc is supported by the national institute for health research university college london hospitals biomedical research centre. none declared.box . what to tell the patient.reassure patients they are not infectious to others. discussion about informing fellow travellers of potential risk of having caught malaria and need to present early if symptoms develop. inform patient that he/she will be notified to public health authorities as part of routine national policy. warn patients about possible recrudescence or relapse of malaria following treatment and to report recurrence of fever to their general practitioner. warn patients treated with artesunate about possible haemolysis and the importance of attending for follow-up blood tests. at follow up, review results of any tests needed or performed for other travel related infections including hiv infection. discuss period of exclusion from blood donation after malaria e blood donors need to inform and discuss own situation with national blood service. reinforce need for up to date advice on malaria prevention during all future travels, for patient and family, and provide information on sources of advice. key: cord- -rumqopg authors: jacob, chaim oscar title: on the genetics and immunopathogenesis of covid- date: - - journal: clin immunol doi: . /j.clim. . sha: doc_id: cord_uid: rumqopg most severe cases with covid- , especially those with pulmonary failure, are not a consequence of viral burden and/or failure of the ‘adaptive’ immune response to subdue the pathogen by utilizing an adequate ‘adaptive’ immune defense. rather it is a consequence of immunopathology, resulting from imbalanced innate immune response, which may not be linked to pathogen burden at all. in fact, it might be described as an autoinflammatory disease. the kawasaki-like disease seen in children with sars-cov- exposure might be another example of similar mechanism. replicate both in the upper and lower respiratory tract might explain why people infected have such different experiences. the virus can start in the throat or nose, producing a cough, disrupting taste and smell, and then end there. or it might work its way down to the lungs and debilitate that organ and the entire organism. it is quite likely that once sars-cov- gets down in the lungs, it's probably just as deadly as sars-cov- . in sum, the assessment expressed in the general media, but sometimes also in professional literature [ ] [ ] [ ] [ ] that the virus is entirely -new‖ and that the immune system is -naive‖ namely that it is totally inexperienced when it comes to this virus is exaggerated. a more accurate description of this virus would be a reemergence of a known foe that has a somewhat more optimal organization of its resources making it more suitable to become a pandemic pathogen. the genetic sequence diversity of sars-cov- is low. as a consequence of a massive effort to sequence viral samples obtained from humans infected all around the world, there are now thousands of viral sequences available which permits an unprecedented and detailed analysis of the virus's genetic evolution. during the early phase of the outbreak in wuhan two major lineages of sars-cov- , with different exposure histories, were categorized as l (∼ % of sequences) and s (∼ %) based on two tightly linked single nucleotide polymorphism (snp)s-a synonymous mutation in the orf ab of the genome, and a non-synonymous s l amino acid change in orf . [ ] . the s variant was evolutionarily more related to animal cov. the functional consequences of the s l mutation are not known. nevertheless, the two variants exhibited similar virulence and clinical outcome [ ] . a mutation in the spike protein-that mediates sars-cov- entry into host cells and potentially of functional importance-was described by several teams of researchers [ ] [ ] [ ] [ ] . the d g mutation at residue of spike (s) protein causes an amino acid change from aspartate to glycine. the mutation that causes the d g amino change is transmitted as part of a conserved haplotype defined by four snps that almost always track together, although they probably arise independently. beside the d g mutation the haplotype includes another nonsynonymous mutation (p l) in the nsp viral protein. while the functional consequences of the p l mutation remain unclear at present, there is strong evidence that the g variant is associated with greater infectivity and higher viral loads in the upper airways during infection [ , [ ] [ ] [ ] [ ] . the s protein must be cleaved by host proteases to enable membrane fusion, which is critical for viral entry. the g mutation creates a novel serine protease cleavage site that can facilitate its cleavage by host serine protease elastase- [ ] . also g mutation increases both spike stability and membrane incorporation [ ] . in cell culture, s-g pseudovirus infected ace -expressing cells significantly more efficiently than the s-d pseudovirus [ , ] . even though the g variant appears to be more infectious, it did not appear to be more virulent since hospital outcomes were similar with either variant [ , ] . dynamic tracking of variant frequencies sampled from covid- patients from asia, europe, north america, and australia show that the d variant was initially dominant, so most subjects infected during december through the end of february , had this variant [ ] . the earliest viral sequence that carried the g mutation with the other three snps that characterize the haplotype was sampled in northern italy on february th . during march both variants could be identified circulating in the population [ , ] . by april, the g variant was circulating almost exclusively in european and in the greater nyc area. this variant continued increasing in frequency over several months so that by june it has become the dominant variant all over the globe. it is tempting to hypothesize that the successful mitigation of the outbreak in china and several other east asian countries was due, at least in part, to the fact that they faced the less infective d variant. the european countries and nyc had to deal with the more infectious g variant. similarly, the early outbreak in washington state was caused by the d variant [ ] . the original march-april outbreak on the western coast of the united states might have been less severe than in the northeastern united states, because it originated from the washington state d variant. by june , the g became the global dominant variant, and is responsible for the much more infective july phase of the outbreak in the southern and western usa. the viral genetic data presented clearly demonstrate that variants may arise quickly, even in cov, and have profound effects on the spread and consequences of the covid- pandemic. prior research has uncovered gene variants that can alter a person's chances of contracting a viral infectious disease. the most famous example is a mutation in the ccr gene, which offers protection against hiv. while there are coding variants in human ace , with some of them in residues considered important for the binding of s-protein in cov [ ] , there is no evidence for the existence of cov s-protein-resistant ace mutation in any population [ ] . neither is there any evidence for ace variants that bind more, or less, efficiently to sars-cov- s-protein. based on publicly available data from east asia, europe and north america, a group of british epidemiologists conclude that children are half as likely to be infected by sars-cov- as adults [ ] . the reliability of these estimates are limited by lack of direct assessments of the transmission of the virus between adult to child, and child to child, compared with adult to adult. furthermore, because children tend to have less comorbidities than adults, they experience less disease symptoms, and as such are tested less than adults. hence, the number of children infected may be grossly undercounted. a publication that in this short timeframe has already been cited frequently, seeks to confirm that the incidence of sars-cov- infection is lower in children than adults due to a lower density of ace receptors in the nasal epithelium of children as compared to adults [ ] . the researchers report finding less ace rna in cells scraped from the noses of children than in those from adults. the significant difference in this study is reported to be between those under , and those aged - . in addition, there was no difference in the amount of ace rna detected according to gender, or those with or without asthma. several caveats to consider: the average relative amount of rna ranged from . for those less than years old, to . for those and older. the differences are relatively small and the error bars large. while those over are those most affected by covid- , the study did not include individuals older than years. also, the density of ace receptors may not be uniform throughout the nasal mucosa, and no evidence is presented that the entire nasal mucosa was j o u r n a l p r e -p r o o f equally sampled. lastly, it is unknown how many receptors are needed for the sars-cov- virus to successfully infect us. in any case, there is no doubt that children of any age can be infected. i am more convinced by the conclusions of dong et al. [ ] that children of all ages are susceptible to infection by sars-cov- without significant sex differences. although clinical manifestations of children's covid- cases seem generally less severe than those of adult patients, young children, particularly infants, are vulnerable to infection. as the aforementioned publication [ ] assumes, if ace is the sole receptor for viral entry, then the expectation is that high ace tissue expression equals higher infectivity and worse outcomes. however, a careful consideration of the role of ace in the renin-angiotensin system physiopathology is indicative of it playing a protective role-meaning higher ace expression is more likely to protect us from a worse outcome of viral infection. as such, ace has an important role in counterbalancing the effects of ace . angiotensin ii, a product of ace cleaving angiotensin i, can cause vasoconstriction, inflammation, and fibrosis. ace cleaves angiotensin ii to angiotensin - , which antagonizes the activities of angiotensin ii-hence, it can suppress inflammation, fibrosis, and generate vasodilation. further, a high ace /ace ratio protects the integrity of the endothelium and promotes antithrombotic activity. previous studies have found ace playing a protective role in severe lung injury in ace knockout (ko) mice [ ] . ace ko mice challenged with avian influenza h n [ ] or h n [ ] resulted in severe lung injury, despite that ace is not a receptor for avian influenza. in fact infection with avian influenza strains resulted in downregulation of ace expression in the lung and increased serum angiotensin ii, both in mice and human subjects infected by the virus [ ] . the question is whether ace expression levels are pertinent to sars-cov- infection only in the tissues relevant to viral entry and the lungs as its major target, [ , ] or, given that covid- in its severe form is a systemic disease with multi-organ disfunction [ , ] , ace expression levels may be important in multiple organs and tissues other than those of the respiratory system. relevant to this question, lungs do not have high expression levels of ace , and relatively few cell types express ace in the lung compared to other tissues [ , ] . assuming the importance of ace expression throughout the human organism, in silico analyses have been undertaken through integrating public genomics, epigenomics and transcriptomics data in multiple tissues, different populations, disease conditions, and age as well as sex considerations. intriguing in silico findings suggest that east asian populations have higher allele frequencies in expression quantitative trait loci (eqtl) variants associated with higher ace tissue expression compared to european and african populations [ , ] . furthermore, ace expression increases by estrogens and to a much lesser degree by androgens, which possibly explains the higher ace expression in females [ ] . the study suggests an inverse age-dependent ace expression in both males and females, a reduced expression in type diabetes, and inhibition of ace expression by inflammatory cytokines [ ] . these interesting suggestions (supporting a protective role of high ace expression against sars-cov fatality) need to be validated by clinical observations, in vivo, and in vitro experimentation. until such time, the functional consequences of ace expression levels to the susceptibility or response to sars-cov- remain unclear [ ] . in addition to ace , viral entry requires s-protein cleavage at the s /s and s ' sites allowing fusion of viral and cellular membranes by host proteases. the transmembrane serine protease s (tmprss ) is frequently employed for this purpose by sars-cov- and sars-cov- [ , ] , but also by mers [ ] , and human cov- e [ ] . the functional importance of tmprss in cov infections was tested in tmprss ko mice infected with sars-cov- and mers-cov. results show that lack of tmptss in the airways reduced severity of lung pathology after infection by sars-cov- and mers-cov, despite that all other host proteases were intact [ ] . apart from cov, tmprss is an important host protease for influenza viruses, by cleaving the influenza virus hemagglutinin (ha) molecule [ ] . furthermore, genetic variants with higher tmprss expression increase susceptibility to severe human h n ( ) and avian h n influenza [ ] . these findings, raise the intriguing question whether genetic variants with higher tmprss expression confer higher risk and/or severity of sars-cov- infections. a preliminary study from italy suggests that two distinct tmprss haplotypes show significant frequency differences between italian and east asian populations [ ] . specifically, the rare alleles of these haplotypes predicted to induce higher levels of tmprss , are more frequent among italians. a snp belonging to one of the haplotypes is the same one found to be associated with increased susceptibility to severe influenza [ ] . as will be discussed further, cytokines play an essential role in the pathogenesis of covid- . while environment, microbiome, genetics, and host factors, each can influence cytokine responses to pathogen stimulation, genetic variations appears to be a main component shaping cytokine responses in humans [ ] . for perspective, the microbiome does seem to have a smaller impact on cytokine production capability. it is estimated that microbiome explain only % of cytokine production [ ] . large-scale studies from the human functional genomics project [ ] have shown that different cytokines have different levels of genetic influence. this is an important concept for host defense and disease, as cytokines are fundamental in orchestrating overall immune responses, and can drive pathology when dysregulated, as is the case in covid- . pertinent to the present discussion, the il- /il- pathway especially, seems to be regulated mainly by genetic factors [ ] . my own early work, has provided evidence for the heritability of tnf production capability in mouse and man [ , ] . notwithstanding the current paucity of such studies in cov, i strongly believe this area of research promises to generate valuable information as for the pathogenesis and potential treatment of covid- . african americans infected with sars-cov- seem to be at a greater risk for severe outcome. while comorbidities and socioeconomic circumstances certainly play a critical role, cytokines may have an important role too. a preliminary study compared expression levels of cytokines and other immune modulators between caucasian americans and african americans using rnaseq data [ ] . results show il-  and il- receptor (il r ), il r , tlr and tlr were significantly higher in african americans suggesting perhaps the tendency to develop higher inflammatory cytokine responses. much more work is needed to validate these observations. genome wide association studies (gwas) allowing the unbiased clustering of genetic variation defining human diseases, require assembly of dna samples from very large number of subjects which usually take a long time to collect. given that we are just six months into the pandemic, it is quite remarkable that a medium size gwas was already completed. it is the first to document a statistically significant association between genetic variants and severe covid- [ ] . variations at two loci in the human genome were associated with an increased risk of respiratory failure in patients with covid- ; one, j o u r n a l p r e -p r o o f within the abo blood type cluster on chromosome , and the other at position p . . the frequency of the chromosome risk allele was significantly higher in those patients that needed mechanical ventilation, compared to those with less severe disease progress. importantly, this locus is home to six genes, and it is not yet possible to identify which of them is responsible for aggravating the disease course. this locus contains a cluster of chemokine receptors, xcr , ccr and cxcr (and several other cytokine receptor genes close by), which have important regulatory functions in the innate immune system. another candidate within the locus, slca is an amino acid (proline) transporter expressed at luminal membrane of small intestine and proximal tubule kidney cells and functions in absorption of proline. its expression in rodent intestine depends on the presence of ace [ ] . however, amino acid transporters have been shown to induce cytokine responses: genetic variation at the slc a amino acid transporter show strong association with pathogen induced il- production [ ] . also amino-acids or amino acid catabolites have been reported to modulate cytokine production [ , ] . so it is conceivable that slca might influence the course of covid- severity by affecting pathogen induced cytokine production, rather than viral entry through ace . in this respect, the same risk snp in the chromosome abo blood type cluster that affects covid- severity has been associated with elevated il- levels in childhood obesity in a previous gwas, thus, possibly linking this genetic allele with elevated il- levels (with or without full-blown ‗cytokine storm') described in severe covid- patients. the study [ ] is equally striking for the genes that failed to turn up. pathogen microorganisms, including several viral infections, are controlled by genetic variations at the hla complex at chromosome p [ , ] . the class i and class ii gene products of the hla are involved in antigen presentation, a mandatory process to initiate an adaptive immune response geared to restrain a pathogen. but genetic variants at the hla region did not appear to make a difference in the risk of severe covid- . thus, the so called ‗adaptive' arm of the immune system seems to be less relevant to covid- than the ‗innate' immune response. i have offered here a hypothesis for what these genetic associations might actually mean. if correct it has major mechanistic implication as for the pathogenesis. at minimum it suggests avenues for further studies. recognition of a pathogen by the innate immune system triggers the secretion of the crucially important type i/iii interferons (ifn). the result of ifn signaling is the activation of an entire cascade of events that include the release of proinflammatory cytokines which further signal to endothelial cells, which then enable chemokines to spread throughout the blood to recruit innate immune cells to the site of infection. the recruited nk cells, monocytes, and neutrophils interact with activated endothelium to leave the blood stream and migrate toward the site of infection. at the site of infection they can perform effector functions to control infections, such as release of reactive oxygen species (ros) and, where they can perform effector functions to control infections, such as release of reactive oxygen species (ros) and directly killing infected cells, as well activating a pathogen specific adaptive immune response. given the shared sequence homology, the virus-host interactions of sars-cov- is likely to be analogous to those involving other cov. however, these interactions might be similar to j o u r n a l p r e -p r o o f other non-cov viruses as well because of limited repertoire and conserved mechanisms of innate immune signaling. sars-cov- engages host pattern recognition receptors (prr), and toll-like receptors (tlr) which initiate downstream signaling pathways triggering secretion of cytokines. if present early and properly localized, ifn are considered the most effective in limiting cov infections [ ] . ifn induced proteins can interfere with viral entry and s-protein-mediated membrane fusion [ , ] . however, in a later phase of the infection, ifn can become pathologic (e.g. upregulation of ace in airway epithelium [ ] and orchestration of inflammatory response contributing to immuno-pathogenesis [ ] ). sars-cov- , similar to other cov, have developed multifaceted mechanisms to inhibit ifn induction and signaling [ ] . this is evident by an early impaired ifn signature in severe covid- patients [ , ] , while actively promoting other inflammatory pathways contributing to pathology (e.g. secretion of pro-inflammatory cytokines interleukin (il) β (il- β) and il- [ ] ). sars-cov- is particularly effective in inducing il- and il- , by inhibiting an endogenous nf-kb repressor, nkrf [ ] but probably by other mechanisms as well. while the human innate immune system resources remained unchanged, cov employs multiple innate immunity evasion mechanisms as reviewed recently [ , ] . the earlier cov infections can provide an important road map to understand covid- pathogenesis. thus, a clear indication that immunopathogenesis contributes to sars was the observation that sars-cov- viral loads were found to be decreasing while disease severity increased [ , ] . longitudinal in vivo experiments in which ferrets were infected with sars-cov- intranasally showed a robust production of cytokines that continued beyond clearance of the virus. by day seven, despite waning viral burden, the cytokine response continued to expend. remarkably, by day fourteen, while the virus was fully cleared, some cytokines and specifically il- remained elevated [ ] . in fact, il- emerges as the dominant cytokine driving the immunopathogenesis. given that old age appears to be an independent risk factor for developing severe covid- [ ] , it is noteworthy that in a groundbreaking study ter horst et al., have shown a clear and consistent increase in il- and il- ra production in old age [ ] . evidence is accumulating in covid- patients pointing to dysregulated monocyte driven dendritic cells and macrophage responses, which then drive the characteristic acute respiratory distress syndrome (ards) and cytokine release syndrome (crs) [ ] . cd dim nk cells, generally thought to contribute to antiviral host defense through cellmediated cytotoxicity, were depleted primarily in severe cases. whereas cd bright nk cells, which are considered producers of ifn-γ and tnf-α, were significantly depleted in all covid- samples tested [ ] . evidence supports recruitment of nk cells from the periphery to the lung. activation of these nk cells in the target tissue in an environment enriched for il- (and other cytokines) probably contributes to pathogenesis [ , ] , as opposed to resolving the infection. severe cases of covid- have significant increase in neutrophil levels in circulation [ , ] and in bronchoalveolar lavage fluid (balf) [ ] . together with the upregulation of chemokines, particularly those that act as chemoattractants for neutrophils and monocytes j o u r n a l p r e -p r o o f [ , [ ] [ ] [ ] , these observations support the influx of these cell types into the bronchi. these neutrophils and monocytes probably disrupt the air-blood barrier by causing collateral damage to airway epithelial cells and vascular endothelial cells while increasing cytokine production. the damage to vascular endothelial cells certainly contributes to microthrombosis. though seemingly contradictory to mechanisms of immune evasion, enhanced innate immune activation is central to the morbidity and mortality of covid- patients. immune evasion seems to characterize the first phase of infection with sars-cov- and this is associated with reduced innate antiviral immunity. however, approximately twenty percent of infected subjects develop an excessive innate immune activation approximately - days after infection, which i argue is associated only marginally, if at all, with viral load. rather, the massive inflammatory response is a consequence of host dysregulation of the immune system. in line with observations in sars-cov- [ ] , sars-cov- induces a robust cytokine response with low levels of ifn in the early phase, culminating in improper recruitment of inflammatory monocyte-macrophage and neutrophil populations into target organs, resulting in further cytokine production [ ] . a recurring theme in covid- pathogenesis is that components of the immune system that are generally thought to contribute to antiviral host defense end up promoting disease severity. typically, the complement system can efficiently recognize and eliminate viral pathogens by opsonizing viruses and virus infected cells, inducing an antiviral inflammatory state, increasing virus-specific immune responses, and neutralizing cell-free viruses [ ] . however, the activation of multiple complement pathways, dysregulated neutrophil responses, endothelial injury, and hypercoagulability appear to be interlinked with sars-cov- infection and instead serve to drive the severity of the disease [ ] . the functional importance of complement activation in cov was tested in c ko mice infected with sars-cov- [ ] . the studies showed that complement activation regulates a systemic proinflammatory response and removal of c signaling reduced lung injury and respiratory dysfunction, despite equivalent viral loads present in the lungs. this was associated with reduced lung infiltration of neutrophils and monocytes and lower cytokine and chemokine levels in both the lungs and sera [ ] . lung biopsy samples from patients with severe covid- show widespread complement activation characterized by c a generation and c -fragment deposition [ ] . the host complement activator masp , the key serine protease in the lectin pathway of complement activation, was identified as a target of the n (nucleoprotein) protein of sars-cov- , mers-cov, and sars-cov- , resulting in aberrant complement activation and aggravated inflammatory lung injury. in mice, lung injury induced by sars-cov- or mers-cov n protein was attenuated when its masp -binding motif was altered, when masp was genetically knocked out, or when the masp -n protein interaction was pharmacologically blocked [ ] . earlier i have discussed the gwas that established significant association between severe covid- and the abo blood type cluster [ ] . having blood type a was linked to an approximately forty-five percent increase in the likelihood that a patient would develop j o u r n a l p r e -p r o o f respiratory failure, while subjects with blood group o were at a thirty-five percent decreased risk for respiratory failure. a possible mechanistic explanation could be that type o patients harbor both anti-a and anti-b natural igm abs. these may help reduce the viral load of their hosts due to early activation of the classical complement pathway followed by viral clearance. such mechanism has been shown to work in vitro, using measles virus produced in cells engineered to express only a-type, b-type or o-type carbohydrate epitopes. measles virus was neutralized by human serum (that did not contain anti-measles ab), utilizing natural abs against the a and b antigens in a strictly complement-dependent manner [ ] . these observations support a role for the complement system in enabling natural abo group abs as first line innate immune defense to viral infections. although most early studies concentrated on the lungs as the target organ in sars-cov- infection, it is now clear that covid- has a wider spectrum of organ involvement. this may be a result of the broad organ tropism of sars-cov- , but is more likely due to an outof-control host immune response to the virus. indeed, evidence is accumulating in support of vascular cell dysfunction in multiple organs during sars-cov- infection [ ] . first, sars-cov- is able to directly infect engineered human blood vessel organoids [ ] . more importantly, histopathological evidence of vasculitis, sometime associated with viral particles, and accumulation of neutrophils and monocytes, and even lymphocytes, in the wall of blood vessels in multiple organs were described [ ] . in addition, endothelial apoptosis and pyroptosis might contribute to endothelial cell injury. similarly, bryce et al., found diffuse vascular endothelial inflammation with micro and macro vascular thrombosis in the venous and arterial circulation [ ] . the vascular endothelium is indispensable for the regulation of vascular tone and the maintenance of vascular homoeostasis. endothelial dysfunction is a principal determinant of microvascular dysfunction by shifting the vascular equilibrium towards more vasoconstriction with subsequent organ ischemia, inflammation with associated tissue edema, and a pro-coagulant state [ ] . vascular endothelial damage could explain why people with pre-existing conditions like hypertension, diabetes, obesity, and cardiovascular disease are at a higher risk for severe complications. all of those conditions, identified as independent covid- risk factors, cause endothelial cell dysfunction. the additional damage and inflammation in the blood vessels caused by the viral infection could push them over the edge and cause catastrophic complications [ ] . given that vascular endothelial cells express ace , one likely hypothesis suggests that sars-cov- infects endothelial cells directly which induces injury, activates complement, and sets up a perpetual inflammatory state [ ] . however, there are alternative mechanisms for activation of endothelial cells and vasculitis induction that do not necessarily require the presence of the virus itself, e.g. neutrophil extracellular traps (nets) [ ] and hypoxia [ ] . the ability of neutrophils to form nets is considered beneficial in host defense against pathogens, but as observed regarding other innate immune mechanisms, sustained net formation can trigger a cascade of damaging inflammatory reaction. indeed elevated levels of net-specific markers, myeloperoxidase dna, and citrullinated histone h , were observed in the sera of covid- patients [ ] . as such, several research groups are supporting a more central role for nets in covid- pathogenesis [ , ] . coagulation disorders in patients with covid- were initially thought to be due to systemic disseminated intravascular coagulopathy (dic). however, considerable cross-talk and mutual engagement between the complement and the coagulation cascades is being increasingly documented in covid- and seems to be responsible for a prothrombotic environment distinct from dic, leading to serious adverse outcomes [ , ] . in fact, elevated d-dimer (a fibrin degradation product indicative of hyperactive coagulation) has emerged as a reliable marker of severe covid- [ , ] . interestingly, the interconnected complement and coagulation cascades, which are being increasingly documented in covid- , has been long recognized in antiphospholipid syndrome (aps) [ ] . given that several publications report antiphospholipid antibodies in patients with covid- [ ] [ ] [ ] , despite them not fulfilling the sydney criteria for aps [ ] , it is difficult to ignore how much the clinical and immunopathological picture of aps resembles that of the autopsy findings of an exaggerated inflammatory state and thrombosis in many covid- patients [ , ] . a study by nicolai et al., provides mechanistic evidence that multisystem disease in severe covid- involves coagulopathy driven by dysregulated innate immune response [ ] . they show inflammatory microvascular thrombi containing platelets, fibrin, and a large number of neutrophils in the lung, kidney and heart. neutrophils were highly activated in severe cases compared to less severe cases and platelets showed enhanced neutrophil adhesion and net formation in multiple organs. thus, dysregulated immunothrombosis is linked to both ards and systemic hypercoagulability [ ] . in sum, an overwhelmed innate immune system is seemingly unable to assemble a balanced response of appropriate cells, cytokines and other molecules to control the infection in a timely fashion. instead, a disoriented, misguided storm of inflammatory cytokines ends up destroying that which it intended to protect. it disintegrated in moderate and severe covid- -lymphopenia with drastically reduced cd + and cd + t-cells is the most consistent finding in moderate and severe covid- patients, which also correlates with disease severity and mortality [ ] [ ] [ ] [ ] . b-cells are decreased as well [ , ] unlikely to be the case [ ] . despite the fact that lymphopenia seems to be a prominent feature of severe covid- , patients under immunosuppressive therapy are not at a higher risk for infection, and if infected seem not to be destined to have more severe progression. in fact, reviewing the mortality and morbidity reports published on sars-cov- , mers-cov, and covid- , no mention is made on immunosuppression as a risk factor for morbidity and mortality. further, no severe complications or fatality is reported to be linked to transplantation, chemotherapy, autoimmune hepatitis, ibd, or other conditions requiring immuno-suppressive treatment for patients at any age [ , ] . in fact, reviewing the mortality and morbidity reports published j o u r n a l p r e -p r o o f on sars-cov- , mers-cov, and covid- , no mention is made on immunosuppression as a risk factor for morbidity and mortality. further, no severe complications or fatality is reported to be linked to transplantation, chemotherapy, autoimmune hepatitis, ibd, or other conditions requiring immuno-suppressive treatment for patients at any age [ , ] . these observations reinforce the notion that the reduced t and b lymphocytes in covid- patients is not the direct cytopathic effect of the virus itself. the possibility that the peripheral lymphopenia observed in patients with covid- reflects recruitment of lymphocytes to the respiratory tract or adhesion to inflamed respiratory vascular endothelium has been suggested [ ] . although autopsy studies of patients' lungs and single-cell rna sequencing of balf do identify the presence of lymphocytes, the lymphocytic infiltration is modest at best, arguing against sequestration as a main cause of lymphopenia [ ] . the most likely scenario is that inflammatory cytokines are key factors behind the observed lymphopenia. indeed, serum levels of il- especially have been closely correlated with lymphopenia, while recovered patients show return of lymphocytes numbers towards the normal range with significant reduction in il- levels. a likely mechanism is via the downregulation of multiple hla class ii molecules on cd monocytes and b-cells by il- , as demonstrated by multiple studies [ , , ] . hla class i molecule downregulation was less severe and inconsistently observed. a negative correlation between serum levels of il- and the number of class ii hla on cd + monocytes supports the notion of downregulation of class ii by il- [ , ] . this decrease in hla molecules suggests that severe covid- patients might be unable to mount a normal t cell response due to reduced antigen presentation capability to t cell receptors (tcr). such t cells are then eliminated by apoptosis. autopsy studies on lymphoid organs collected from patients who succumbed to the disease revealed massive lymphocyte death, which was linked to high levels of il- as well as fas-induced apoptosis [ ] . treatment with tocilizumab, an il- receptor antagonist, restores, at least partially, the hla class ii molecules on antigen-presenting cells. in addition, it increased the number of circulatory lymphocytes, further suggesting il- is a key player in the lymphopenia development [ ] . further, nk t-cells are reduced in number and impaired in function in severe covid- in an il- dependent manner [ ] . since cd + t cells require antigen presentation by hla class i for activation via tcr, and these molecules are less reduced than class ii molecule, it is unlikely that the same mechanism of apoptosis of inactivated cd + t cells would be operating to a similar extent in cd + t cells. rather, it is possible that most cd + are eliminated due to activation/exhaustion of t cells [ ] . intense immune profiling of covid- patients show a very heterogeneous immune response. a rigorous comparison between the studies is complicated by major differences in cohort sizes, dissimilar clinical phenotypes used, utilization of different experimental strategies, and emphasis on different parameters by various investigators. the study by giamarellos-bourboulis et al. [ ] suggest that severe covid- patients developing severe respiratory failure show one of two immune phenotypes: ( ) an immune dysregulation phenotype (~ % of patients) characterized by major reduction of hla-class ii molecules on cd + monocytes in the absence of elevated ferritin. this is triggered by monocyte j o u r n a l p r e -p r o o f hyperactivation, excessive il- production, and profound lymphopenia, but without il- β elevation; and ( ) a mas phenotype (~ %) associated with elevated ferritin, with relatively less reduction of hla class ii molecules on monocytes and triggered by il- β. further studies will be needed to verify whether these immunophenotypes are generalizable. mathew et al. [ ] identified a subgroup of approximately twenty percent of severe covid- patients that lack detectable lymphocyte response to the infection, suggesting a total failure of immune activation. further, these investigators emphasize that the typical tcell/b-cell communication and cooperation, the adaptive immune system depends on, is practically nonexistent in some covid- patients. lucas et al., [ ] confirmed an overall increase in innate cell lineages, reduction in hla class ii molecules on monocytes, and early surge in cytokines with parallel reduction in t cells, observed by many other studies. in addition, according to these investigators the immune responses to pathogens can be roughly grouped into three categories characterized by different sets of immune cells and cytokine signals used: type -broadly t h responses directed against viruses and intracellular bacteria; type -directed against parasites that do not invade cells; and type -directed against fungi and bacteria that can survive outside the cells. type immune response expected in sars-cov- infection was seen in mild to moderate cases, while in severe cases the immune system seemed to invest too many resources in non-appropriate type and type immune signaling. this immune disintegration, the investigators have dubbed as -misfiring,‖ seems to extend to the realm of t and b lymphocytes [ ] . other investigators describe the catastrophic cov disease as a lack of switch from an innate immune response to an adaptive immune response [ ] . these depictions of the immune response in severe covid- are symbolically captured in fig. . the antigen specificity of sars-cov- t cells have just started to be characterized in covid- patients [ , ] and their potential protective role awaits additional research. however, it is already clear that patients who recovered show specificity for multiple sars-cov- proteins, not only spike protein. interestingly, cross-reactive memory cd + and cd + t cells are also found in ~ % of subjects who have never been exposed to sars-cov- / [ , ] . while sars-cov- / unexposed donors can recognize both structural and nonstructural viral proteins, nonstructural orf- -nsp / -specific t cells are often dominant. in contrast, sars-cov- / recovered individual preferentially recognize structural proteins [ ] . at present no satisfactory explanation for this phenomenon has been offered. also unclear is how these preexisting memory t cells, which are presumably generated in response to human common cold cov, affect immunity or pathology upon sars-cov- infection. heterologous viral t cell immunity and immunopathogenesis-an important consideration that has not yet received sufficient consideration in the current viral pandemic, is highly relevant to the observations of cross-reactive t-cell responses in non-exposed subjects. in fact, only laboratory animals kept in pathogen free conditions are naïve. no human more than a few weeks old is immunologically naïve [ ] . the history of previous exposure, not only to related, but also to unrelated microorganisms, can greatly alter the host's immune response to a viral infection and can change the course of disease [ ] . in fact, it is very likely that this phenomenon is a common feature of human viral infections. t cells have high levels of j o u r n a l p r e -p r o o f cross-reactivity because the tcr first scans the peptide-hla complex by binding to the hla, and then it molds itself around the peptide. actually, the tcr contacts only - amino acids in the peptide, so the total energy of the tcr-peptide-hla interaction is heavily influenced by the hla rather than the peptide. the consequence is a highly promiscuous t cell which allows a large variation in peptide sequences without inhibiting the hla-peptide-tcr interaction [ ] . hosts that have never experienced a particular virus, might, nevertheless, have memory t cell pools that show specificity for it by virtue of crossreactivity that can shape the repertoire of the primary response. the observation that the same virus can cause widely different pathological manifestations in humans might be due (at least in part) to an established adaptive immunity toward related or unrelated viruses, which results in enhanced protective immunity in some, reduced protective immunity in others, or altered immunopathology, including enhanced disease severity in some hosts [ ] . the role of humoral responses in the pathogenesis of covid- remains unclear. as in sars-cov- infection, most subjects infected by sars-cov- seroconvert within - days after infection and this process is associated with increase in plasma cells, whereas naïve b cells decrease significantly [ ] . while recovered patients generate sars-cov- specific neutralizing abs and spike-binding abs concurrently, their titers are highly variable in different patients [ ] . about one third of recovered patients generate very low titers of sars-cov- -specific neutralizing abs [ , ] , and some patients (possibly up to %) who recover, do not have detectable neutralizing antibodies at all [ , , ] . these observations bring up the question of how the virus was cleared-as it eventually was in all those patients studied-without strong ab responses. we can speculate that t-cell mediated immune responses, or non-specific responses by innate immune cells were responsible for viral clearance. however, since a pathogen that kills off the host that it needs to survive is also threatening its very own existence, the possibility of cov self-clearing should be considered, especially when alternative hosts are plentiful. in a remarkable study [ ] the levels of total igg and igg neutralizing antibodies (as measured using a spike protein pseudotyped virus) were quantified in symptomatic and asymptomatic patients eight weeks after release from the hospital (roughly three months after start of infection). the levels of igg and neutralizing antibodies were significantly decreased in the majority of patients in both groups. the decrease in neutralizing abs was more pronounced in asymptomatic (~ %) as compared to symptomatic (~ %) patients. taken together, the finding that ~ % of people infected with sars-cov- do not make anti-viral abs, added to the observation that neutralizing abs begin to drop noticeably during convalescence-suggests that infection with sars-cov- does not establish long-lasting serological immunity, at least not for those who are asymptomatic or mildly ill (more than % of all those infected by sars-cov- ). even more problematic-several studies show significantly higher igg and iga ab responses. this does not correlate significantly with protection, but rather with severity of disease [ ] [ ] [ ] [ ] similar to what was seen in sars [ ] . at minimum, these studies suggest that a robust ab response is insufficient to protect from severe disease [ ] . while it is frequently assumed that anti-sars-cov- abs might be either beneficial or irrelevant, there is also the possibility that such abs might actually be detrimental. first, abs can cause immunopathology by binding viral fragments followed by activation of the complement cascade by the ab complex. the consequences of complement activation in the pathogenesis of covid- was discussed above (section ), and extensively reviewed by others [ ] . second, ab responses to cov may contribute to pathology via ab-dependent enhancement (ade). ade is mediated by non-neutralizing virus-specific igg engagement of fc-receptors (fcr) expressed on immune cells, particularly monocytes and macrophages, leading to inflammatory activation of these cells. anti-s-igg passive immunization of sars-cov- -infected rhesus monkeys significantly enhanced the viral induced acute lung injury with massive accumulation of monocytes and macrophages in the lung in an fcγr dependent fashion [ ] . further, serum containing anti-s-igg from patients with sars-cov- enhances the infection of sars-cov- in human monocyte-derived macrophages in vitro [ ] . a monoclonal ab isolated from a patient with mers, targeting the s-protein of mers-cov showed fcr dependent adh [ ] . high dose iv immunoglobin (ivig) treatment which has shown some efficacy in cov including covid- [ ] , may diminish adh by blocking fcr mediated activities of monocytes and macrophages. direct evidence for ade was not documented in covid- patients so far. however, as argued [ ] , ade should be given full consideration in the safety evaluation of emerging candidate vaccines for sars-cov- . finally, it should be emphasized that at present, neither anti-spike neutralizing abs nor anti-spike t-cell responses have been established as corollaries of protection. patients afflicted with a chronic autoimmune disease have an increased risk of infections including viral infections [ , ] . likewise, acute viral infections may also exacerbate pre-existing autoimmune disease, and immunosuppressive therapies may render patients with autoimmune disease more vulnerable to viral infections. despite these compounded considerations, patients with systemic or organ specific autoimmune disease are not at increased risk for infection with sars-cov- . in fact, of the hundreds of reports published, none mention autoimmune conditions as either independent risk factors for disease or as indicative of a more severe outcome if infected. the data actually suggests that the rate of infection with the virus and their clinical course is not any different from that of the general population [ ] [ ] [ ] . those autoimmune afflicted patients that have develop severe covid- , are likely to have other comorbidities that are independent risk factors for severe disease. while virus infections can cause flares in otherwise stable autoimmune disease, the data suggests that sars-cov- infection is not associated with increased incidents of autoimmune flares. i have previously (section ) discussed and presented evidence that patients under immunosuppressive therapies, including those afflicted with autoimmune conditions are not at increased risk for infection or for more severe outcome [ , ] . regarding sle, the prototypic systemic autoimmune disease, a group of investigators suggested that inherent epigenetic dysregulation causing hypomethylation and overexpression of ace , the functional receptor for sars-cov- , might facilitate viral j o u r n a l p r e -p r o o f entry, viremia, and increased likelihood of cytokine storm in such patients [ ] . the aforementioned role of ace expression, as discussed above, suggests that higher expression of ace might actually benefit the host more than it benefits the virus. in any case, the clinical experience and published data do not support these predictions. moreover, the accumulating scientific information does not support the notion that severe covid- is a direct cytopathic viral disease, but rather a disease in which multi-organ insult occurs by the host's own immune system [ ] . conditions in which the immune system attacks its own tissues are usually associated with development of autoabs. indeed, gagiannis et al., studied prospectively a group of patients for the possible role of autoimmunity in covid- patients [ ] . autoab titers ≥ : were detected in / covid- patients who required intensive care treatment, and in / patients with milder clinical course. based on serological, radiological, and histopathological similarities between covid- -associated ards and acute exacerbation of connective tissue disease induced interstitial lung disease, these authors suggest that sars-cov- infection might trigger or simulate a form of organ specific autoimmunity [ ] . similarly, zhou et al., report on autoabs in severe covid- patients. the prevalence of anti- kda ssa/ro ab, anti- kda ssa/ro ab, and antinuclear ab (ana) was %, %, and %, respectively [ ] . ana was reported in over % of consecutive covid- patients [ ] . several studies document different apl abs in covid- patients mostly associated with thrombotic phenomena [ - , , ] . whether apl in covid- is transient, as has been documented in many other viral infections, or develops into persistent and pathogenic, is very difficult to judge from the results published so far. pregnancy related morbidity with fetal losses have not been reported in connection with sars-cov- infections. however, since an infection often precedes the clinical onset of aps, it is certainly justified to follow up a positive apl test in a covid- patient with a repeated test approximately twelve weeks later to further evaluate the possibility of post-infectious aps. a kawasaki-like disease seen in children is the closest link between sars-cov- and the appearance of an autoimmune and/or autoinflammatory condition. investigators from italy's pandemic epicenter in bergamo were the first to focus attention on this disorder [ ] . d'antiga and his colleagues, quantified the time course and incidence of kawasaki-like disease in children before and after the covid- pandemic, documenting a thirty-fold increase in incidence of kawasaki-like disease after the beginning of the pandemic [ ] . kawasaki disease is an acute and usually self-limiting vasculitis of medium and small sized arteries with specific predilection for the coronary arteries that affects previously healthy young children typically under the age of five years. in the acute phase of the disease, patients with kawasaki disease might have hemodynamic instability, a condition known as kawasaki disease shock syndrome. same patients with kawasaki disease fulfil the criteria of macrophage activation syndrome (mas). an association between kawasaki disease and various viral infections have been suspected, including viruses of the coronavirus family, however a specific infectious trigger has yet to be identified. sars-cov- should be now added to the list of implicated viruses. the most accepted pathogenetic hypothesis supports j o u r n a l p r e -p r o o f an aberrant response of the immune system to one or more unidentified pathogens in genetically predisposed subjects. following the report from bergamo further reports of similar cases from many countries have been published [ ] [ ] [ ] [ ] [ ] . with approximately , kawasaki-like cases reported, these studies provide a consistent clinical picture: the disease appears - weeks after an infection with sars-cov- and most patients have serological evidence of infection; patients are on average older than those with classical kawasaki disease; patients experience respiratory and gastrointestinal involvement; signs of hemodynamic instability; greater incidence of myocardial injury; and more intense inflammatory response due to dysregulated immune response. the incidence of coronary aneurism is lower than in kawasaki disease, but this may be a consequence of relatively short follow up. most patients so far have responded well to the same therapies used for classical kawasaki disease. all these results and considerations support the notion that the immune response to sars-cov- is responsible for this kawasaki-like disease [ , ] in susceptible patients. u.k. pediatricians and their national health service defined and named the ‗new', disease -pediatric inflammatory multisystem syndrome temporally associated with severe acute respiratory syndrome coronavirus (sars-cov- ),‖ or pims-ts [ ] . the cdc in the u.s. and the who subsequently published their own differing definitions of the disorder, which they termed multisystem inflammatory syndrome in children (mis-c) [ ] [ ] [ ] [ ] . physicians and scientists in the field of biology are prone to giving names that are often more confusing than helpful. i have worked for some years on a cytokine named tumor necrosis factor (tnf) that had practically nothing to do with necrosis of tumors, and from any sensible perspective is as much an interleukin (il) as any of the approximately ils. i am not aware of any clinical, therapeutic, or long term follow up consideration that would benefit from a more nuanced definition and naming such as pims-ts or mis-c, rather than simply kawasaki-like disease. in fact some authors agree that -until more is known about long-term cardiac sequalae of mis-c, providers should consider following kawasaki disease guidelines for follow up‖ [ ] . until a much better understanding of the pathogenesis of kawasaki disease and kawasaki-like disease emerges, i do not see a reason to further confuse the literature. as i am writing this essay, the pandemic seems far from being over. it surely looks like we are not even in the end of the beginning. the long-term impact of covid- is too early to evaluate. patients who have recovered from the disease report lingering chronic fatigue, muscle weakness, loss of sense of smell, and difficulties in concentration. but this might be just the tip of the iceberg. we already know that impaired liver function continues for some time after patients have apparently recovered and the virus has cleared. it is also probable that some patients will have lasting pulmonary damage due to fibrosis as has been documented in about % of patients recovered form sars [ ] . similarly, myocardial j o u r n a l p r e -p r o o f scarring will cause cardiac impairment in certain covid- recovered patients. moreover, the long-term consequences of the massive inflammatory response affecting many tissues, and its effect on the competence of the immune system itself, are unknown. it is, however, possible that the intense inflammation in many tissues might cause cellular damage and exposure of self-antigens eliciting auto-reactive t and b cells and generating an autoimmune disease. another question that could take years to answer is whether the sars-cov- virus may lie dormant in the human body for years and then launch itself later in a different form. for example, after a chicken pox infection, the herpes virus that caused the illness reemerges after decades in form of shingles. similarly the hepatitis b virus causes the appearance of liver cancer years later. a general comment is pertinent at this point: the culmination of an interaction between an infectious agent and the human host, even when -full recovery-ensues, does not mean the organism is restored to its previous state (before the encounter), but rather the organism acquires a new equilibrium. as the french physician and philosopher george canguilhem wrote some years ago: -contrary to orthodox medical teaching, health is not some absolute state of perfect physical and mental wellbeing. it is the margin of tolerance for the inconsistencies of the environment… disease is not simply disequilibrium or discordance; it is an effort on the part of nature to effect a new equilibrium in man‖ [ ] . we have learned a tremendous amount about sars-cov- and covid- in a short amount of time. the efficient transmission of data exhibited during this time, has been surpassed only by the efficient transmission of the virus itself. although originally conceptualized as a primarily respiratory viral disease, covid- is now clearly recognized as a far more complex, multi-organ, and heterogeneous illness. as with sars-cov- and mers-cov, the important considerations for the delicate balance of the viral-host interaction that are responsible for covid- are now increasingly appreciated: ( ) fast and robust initial viral replication; ( ) early viral inhibition of ifn induction and signaling causing a delayed ifn expression which drives immunopathology; ( ) out of balance antiviral innate immune response becomes immunopathogenic; and lastly ( ) disintegration of the adaptive immune response. paul ehrlich's prediction of horror autotoxicus-at the turn of the th century-has been realized by the innate immune response to cov in the st century. the language of immunology is rife with war metaphors. for over a hundred years we have been educated to believe in the metaphor that the immune system acts as an army defending our bodies. as richard lewontin has written: -while we cannot dispense with metaphors in thinking about nature, there is a great risk of confusing the metaphor with the thing of real interest. we cease to see the world as if it were like a machine and take it to be a machine. the result is that the properties we ascribe to our object of interest and the questions we ask about it j o u r n a l p r e -p r o o f reinforce the original metaphorical image and we miss the aspects of the system that do not fit the metaphorical approximation‖ [ ] . what has sars-cov- revealed? for me, the answer is our immune response to covid- serves as proof of everything i have long thought was wrong with viewing the (adaptive) immune system as a defense organization: it reacts too slowly. it fights today's threats with the solutions of past problems. it is susceptible to exploitation. it destroys that which it intended to protect. it is large, complicated, elaborate and wasteful [ ] . if you stand back and evaluate how ineffective is the immune system as a defense organization, it is only logical to conclude that it was never intended as one. sars-cov- will eventually be contained, but not by our immune systems. rather, by the international brotherhood of the scientific community. epidemics have always played a natural part in the fabric of human history, but there has never been a time in history where so many different and powerful tools were available to accomplish this task. of all human conditions that disseminate virulent diseases, hubris emerges across centuries as a key driving force. we should embrace cesar augustus motto -festina lente‖, make haste, slowly-even or especially when you are feeling the crunch, take your time. this is not the time for us to skip corners. table . comparison of transmissibility expressed as reproduction number, r and case fatality rates of selected human viral diseases. adopted with changes from wang et al., [ ] . the disintegration of the persistence of memory ( ) (rt) by salvador dali is an oil on canvas reaction to his original work the persistence of memory ( ) . in this version, the landscape from the first painting has been engulfed by water. disintegration of objects is occurring above and below the water. the block and plane from his original (lt) have now been separated into brick-like objects that float. some of the bricks on the left side of the painting begin to disintegrate. a watch beneath the water is coming to pieces and another one that sunk beneath the layer of bricks, leaves bits of debris behind. while persistence of memory (lt) symbolizes the importance of immunological memory as cornerstone of adaptive immunity, the disintegration of the persistence of memory (rt) is a metaphor for gorbalenya, and the coronaviridae study group of the international committee on taxonomy of viruses, the species severe acute respiratory syndrome-related coronavirus: classifying -ncov and naming it sars-cov- the 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an observational cohort study clinical characteristics of children with a pediatric inflammatory multisystem syndrome temporally associated with sars-cov- multisystem inflammatory syndrome in children in new york state multisystem inflammatory syndrome in u.s. children and adolescents acute heart failure in multisystem inflammatory syndrome in children (mis-c) in the context of global sars-cov- pandemic kawasaki-like multisystem inflammatory syndrome in children during the covid- pandemic long term sequalae of sars: physical, neuropsychiatric, and quality-of-life assessment on the normal and the pathological the triple helix some savage cuts in defense i thank tamar jacob-eliasi for invaluable help with editing and proofreading the manuscript. coj is funded by nih grant r ar . key: cord- -a ft wdy authors: custovic, a.; johnston, s. l.; pavord, i.; gaga, m.; fabbri, l.; bel, e. h.; le souëf, p.; lötvall, j.; demoly, p.; akdis, c. a.; ryan, d.; mäkelä, m. j.; martinez, f.; holloway, j. w.; saglani, s.; o'byrne, p.; papi, a.; sergejeva, s.; magnan, a.; del giacco, s.; kalayci, o.; hamelmann, e.; papadopoulos, n. g. title: eaaci position statement on asthma exacerbations and severe asthma date: - - journal: allergy doi: . /all. sha: doc_id: cord_uid: a ft wdy asthma exacerbations and severe asthma are linked with high morbidity, significant mortality and high treatment costs. recurrent asthma exacerbations cause a decline in lung function and, in childhood, are linked to development of persistent asthma. this position paper, from the european academy of allergy and clinical immunology, highlights the shortcomings of current treatment guidelines for patients suffering from frequent asthma exacerbations and those with difficult‐to‐treat asthma and severe treatment‐resistant asthma. it reviews current evidence that supports a call for increased awareness of (i) the seriousness of asthma exacerbations and (ii) the need for novel treatment strategies in specific forms of severe treatment‐resistant asthma. there is strong evidence linking asthma exacerbations with viral airway infection and underlying deficiencies in innate immunity and evidence of a synergism between viral infection and allergic mechanisms in increasing risk of exacerbations. nonadherence to prescribed medication has been identified as a common clinical problem amongst adults and children with difficult‐to‐control asthma. appropriate diagnosis, assessment of adherence and other potentially modifiable factors (such as passive or active smoking, ongoing allergen exposure, psychosocial factors) have to be a priority in clinical assessment of all patients with difficult‐to‐control asthma. further studies with improved designs and new diagnostic tools are needed to properly characterize (i) the pathophysiology and risk of asthma exacerbations, and (ii) the clinical and pathophysiological heterogeneity of severe asthma. and risk of asthma exacerbations, and (ii) the clinical and pathophysiological heterogeneity of severe asthma. at a recent summit organized by the european academy of allergy and clinical immunology (eaaci), a group of experts discussed current issues of concern and unmet needs in regard to the treatment of asthma. most attendees presented evidence that the current guideline-driven treatment strategies fail in two vital aspects of asthma. there was a general consensus that the needs of patients with severe asthma or frequent asthma exacerbations are not being adequately covered by the current treatment guidelines and are of major health and socio-economic concern. such patients suffer from significantly reduced quality of life and incur disproportionately higher costs on health service resources. this report summarizes the current data that indicate the need for a more rational approach to the treatment of asthma and one that takes into account the emerging evidence for pathophysiological heterogeneity of the disease, including its more severe forms. a significant outstanding problem in the clinical management of asthma is the failure to prevent and/or efficiently treat asthma exacerbations, which are associated with significant morbidity, risk of death and high treatment cost ( ) ( ) ( ) . generally, an asthma exacerbation is considered to be an increase in a patient's asthma symptoms with increasingly impaired lung function that require increased medication, an unscheduled visit to a physician or hospitalization. a single asthma exacerbation requiring extra medication and possibly emergency treatment and hospitalization can increase the annual treatment costs by more than threefold ( ) . recurrent asthma exacerbations lead to a progressive decline in lung function ( ) , and the risk of an exacerbation doubles in children who have had one or more in the previous year ( ) . a rare but distinct phenotype of asthma exacerbation is characterized by the development of sudden severe asthma symptoms in an otherwise mild or asymptomatic asthmatic subject, which may be triggered by an allergen, drug (e.g. aspirin), food, air pollutant, occupational agent, virus infection or, in many cases, an unknown (or unidentified) trigger ( ) . it has been proposed that within the group of patients with recurrent exacerbations, detailed phenotyping in terms of clinical symptoms, lung function, inflammatory and other biomarkers is warranted ( , ) . there is a significant unmet need for identifying and characterizing the factors that increase the risk of asthma exacerbations and the therapeutic and preventive options that reduce these risks. patients with severe asthma have high morbidity and mortality, often require hospitalization and are expensive to treat. they have diverse clinical profiles, which probably reflect diverse disease mechanisms, and, for many of them, standard treatment is not sufficiently effective ( ) ( ) ( ) . severe asthma comprises a highly heterogeneous group of patients, which is defined in various ways in the published literature and in the national and international guidelines. a consensus is emerging that patients should be considered to have a 'difficultto-control asthma' if they have persistent symptoms and recurrent exacerbations despite being prescribed therapy at the highest steps of the guidelines' pharmacological management ( ) . however, it is worth emphasizing that the guidelines also make clear that 'difficult-to-control asthma' is multifactorial, and issues such as incorrect diagnosis, comorbid conditions, nonadherence to prescribed medication, psychosocial morbidity and a number of other factors discussed later in this manuscript are major causes of 'difficult' asthma ( ) . patients should be considered to have severe asthma that is resistant to currently available therapies only following a detailed analysis and appropriate management of all these background problems that are amenable to intervention. a recently published consensus statement on severe asthma broadened the concept of 'difficult asthma' to reflect the situation in less developed countries, where access to medications and appropriate care is a major issue, by defining three different patient groups including un(der)treated symptomatic patients, patients with low treatment adherence or unconventional therapies, and those remaining symptomatic despite high doses of anti-asthmatic therapies ( , ) . nonadherence to prescribed medication has been identified in a number of studies in the developed countries as the most common clinical problem amongst adults ( ) and children ( ) with asthma, including those with difficult-to-control asthma ( , ) . appropriate assessment of adherence ( ) and other potentially modifiable factors (such as passive or active smoking, ongoing allergen exposure, psychosocial factors, etc.) ( ) has to be a priority in clinical assessment of all patients with difficult-to-control asthma, before any decision about increasing the treatment is made (including possible prescription of an expensive biological therapy) ( ) . the clinical diversity and the underlying pathophysiology of severe treatment-resistant asthma need to be characterized, so that rational therapeutic targets can be identified and relevant biomarkers validated ( ) . in addition, there is a need for controller as well as noninterventional studies in this patient population. although inhaled corticosteroids may protect against asthma exacerbations due to allergen exposure, they are reported to be relatively ineffective in children with virus infectioninduced wheezing ( ) . this suggests the existence of different types of childhood asthma with different underlying pathophysiologies ( , ) . recent joint modelling of longitudinal observations on wheezing from parental reports and medical records identified a novel phenotype of persistent troublesome wheeze with high rates of severe asthma exacerbations and healthcare utilization ( ) . despite having phenotypic markers commonly considered to be indicators of good therapeutic response (atopy and eczema), these children had relatively poor response to currently available antiinflammatory treatments ( ) . an atopy-related phenotype of children with high risk of asthma exacerbations has recently been identified by bayesian inference in two birth cohort studies in which multiple skin and ige tests were collected throughout childhood ( , ) . the analysis revealed several different atopic vulnerabilities, only one of which had a much higher risk of asthma exacerbations ( , ) . however, this group of children with a significant risk of asthma exacerbations was identified only using longitudinal data, and cross-sectional biomarkers allowing their identification in clinical practice are still lacking. several attempts have been made to identify predictors of asthma exacerbations. several comorbidities associated with recurrent asthma exacerbations have been identified in adult patients ( ) , including severe nasal sinus disease, gastrooesophageal reflux disease, recurrent respiratory infections, psychological dysfunction and obstructive sleep apnoea. psychological dysfunction ( ) and treatment nonadherence are separate psychosocial factors that may significantly contribute to the risk of asthma exacerbations. a significant number of patients with high rates of asthma exacerbations, and increased airway eosinophilia, were found to have a low perception of their lung dysfunction ( ) . asthma exacerbations are frequent in asthma patients with poor treatment adherence, and poor adherence remains one of the major challenges in the treatment of severe asthma ( , ) . the basis of asthma treatment guidelines is the use of clinical symptoms or impaired lung function to guide treatment of airway inflammation. however, there is a strong rationale for using direct measurement of airway inflammation as an additional means to guide treatment in severe asthma. green et al. ( ) reported that titrating treatment to sputum eosinophil counts was more successful in reducing asthma exacerbations than treatment in accordance with guidelines. this was confirmed in a larger, multicentre study ( ) , which also reported that the frequency and severity of eosinophilic exacerbations were reduced without increasing the dose of inhaled corticosteroids (ics) (the severity but not the frequency of noneosinophilic exacerbations was also reduced). the treatment strategy of these authors was to (i) treat patients with sputum eosinophil counts of > % with the minimal ics dose required to reduce sputum eosinophil counts to normal and (ii) consider other treatment options in patients with normal sputum eosinophil counts, such as treatment of airway obstruction with long-acting beta-agonists (labas) or a leukotriene receptor antagonist, or treatment of neutrophilic inflammation with antibiotics. of interest, a treatment approach aimed at specifically reducing airway eosinophils using a monoclonal antibody directed against interleukin (il)- reduced the risk of asthma exacerbations in patients with severe asthma and persisting airway eosinophilia ( , ) . however, it is of note that even after using this targeted approach, approximately % of severe exacerbations remained unaffected by the treatment. attempts to identify clinical biomarkers for predicting the risk of asthma exacerbation have so far failed. however, some small studies have identified possible candidate biomarkers. for example, gelb et al. ( ) evaluated the use of spirometry and exhaled nitric oxide (f e no) in predicting the risk of asthma exacerbations in stable, mild-to-severe patients with asthma over months. those patients with a baseline fev < % of predicted plus an f e no > ppb had a probability of an asthma exacerbation in months of %, while those with fev > % of predicted plus a f e no < ppb had zero probability of an asthma exacerbation in the same period. the f e no parameter may be a robust measure of the degree of eosinophil airway inflammation ( , ) . numerous epidemiological studies indicate that asthma exacerbations are associated with upper respiratory viral infections, mostly with rhinoviruses, and to a lesser degree with respiratory syncytial virus or coronavirus, with frequency estimates of % and % for childhood and adult asthma, respectively ( , ) . this was confirmed by the ga len-dare systematic review ( ) that reviewed data from published epidemiological studies. the data for childhood asthma are more extensive than that for adult asthma and reveal the absence of geographical influence. the same review indicated that high rates of respiratory bacterial infections are also associated with asthma exacerbations, but the data are inconsistent. however, bisgaard et al. ( ) provided evidence that bacterial infection and viral infection were independently associated with wheezing episodes in infants (< years old), with odds ratios of . and . , respectively. although confirmatory studies need to be performed, current data strongly suggest that respiratory infections are the major cause of asthma exacerbations. case-control studies have revealed a synergism between viral infection and allergen exposure in increasing the risk of asthma exacerbation requiring hospitalization in children ( ) and in adults ( ) . atopic asthmatic patients have more severe and prolonged lower respiratory tract symptoms during rhinovirus infections than nonatopic healthy controls ( ) . this may be explained by impaired innate and acquired immunity of the airways in asthma, as indicated from the studies of experimental and clinical viral infection ( , ) . such impairments in airway immunity correlate with the in vivo severity of the infections and with the viral load ( , ) . consequently, specific and nonspecific antiviral strategies may have great potential as therapeutic and preventative strategies for asthma exacerbations, a hypothesis that is supported by preliminary clinical studies ( ) . patients with asthma also have deficient interleu-kin- and type iii interferon responses to bacterial stimuli, suggesting that inadequate host defence mechanisms and mechanisms specific to the infectious agents are the underlying factors for increased susceptibility ( , ) . rhinovirus and enterovirus infections are more likely to cause an asthma exacerbation than most other viruses in patients of any age, with the exception of rsv in infants ( ) , while virus load and virus co-infection rates correlate with symptom severity ( ) . case-control studies indicate that asthma subjects are at greater risk of invasive pneumococcal infections than patients without asthma, with reported ors ranging from . to . ( - ). for example, the population-based case-control study by klemets and colleagues ( ) assessed the risk of invasive pneumococcal (streptococcus pneumoniae) infection (ipi) amongst adult patients with asthma. using a national population-based laboratory surveillance, , cases of ipi were selected in patients aged - years (thus largely excluding copd patients) along with noninfected controls per ipi case, matched for age, sex and health district. asthma cases were categorized as high (≥ hospitalization within the previous months) and low risk (with prescription drug entitlement and no hospitalization within the previous months). overall, . % of cases had asthma ( . and . % with high-and low-risk asthma, respectively) vs . % of controls ( . and . % with high-and low-risk asthma, respectively), indicating that adults of working age with asthma are at substantially increased risk of ipi. impaired immune responses may be associated with exacerbations in asthmatic children with reduced lung function, but not in those with normal lung function ( ) . specifically, at the time of an asthma exacerbation, a lower expression of th response genes was observed in children with reduced lung function than in children with normal lung function. other studies have shown that genetic factors such as vitamin d ( ) or particulate matter (pm ) ( ) can modify the effect of environmental exposure on exacerbation frequency. the advent of genomewide association studies in populations with severe asthma ( ) and asthma exacerbations ( ) may aid in better prediction of exacerbation phenotypes and the subclassification of patients into subphenotypes that may reflect the differing aetiopathogenesis and response to treatment and so allowing better targeting of treatment. attempts to characterize severe asthma in childhood in childhood asthma, guidelines that outline stepwise treatment escalation with increased disease severity or impaired symptom control fail to adequately deal with nonresponders ( ) ( ) ( ) . in particular, the stepwise treatment appears not always to be appropriate for children younger than years, particularly those with mainly virus-driven asthma, who often do not respond to inhaled corticosteroids ( ) . severe asthma in children is complex, with the asthma phenotype changing during development and requiring continual reassessment ( , , ) . this makes cross-sectional analyses of childhood asthma less useful, as any apparent phenotypic stability within a population will obscure the significant individual instability of disease expression. one of the characteristics of severe treatment-resistant asthma in childhood is the large size of skin test wheal to inhalant and food allergens ( ) . furthermore, in this patient group, the results of skin tests and ige measurements for individual allergens are not always concordant; as a consequence, both tests should be carried out and quantified ( ) . similar quantitative relationship between skin tests (and sige levels) has also been reported in relation to severity of airway hyper-reactivity in adults ( ) . rhinovirus infection in infants induces wheeze and, in a longitudinal study, was shown to be significantly associated with the development of childhood asthma ( ) . in another -year prospective study, teenage asthma was strongly associated with infant wheezing requiring hospitalization ( ) . this study identified eczema and allergen-specific ige as early asthma-predictive factors and the risk of developing teenage asthma to be increased fivefold after respiratory syncytial virus-induced wheezing in infants and > -fold after rhinovirusinduced wheezing. rhinoviruses are frequently found in the lower airways in infants with recurrent respiratory symptoms, with the majority of these rhinovirus-infected infants exhibiting increased airway resistance ( ) . in infants with wheezing requiring hospitalization, sole rhinovirus infection, but not sole infection with any other common airway viruses, was associated with atopy ( ) . a recent study has demonstrated that a cardinal feature of bronchial epithelial cells from children with severe treatment-resistant asthma is impaired interferon-b and ifn-k induction by rhinovirus ( ) . although this patient group was highly atopic, no relationship was observed between atopy, allergy or th -mediated inflammation with impaired interferon ( ) . a longitudinal study of an unselected birth cohort, which monitored lung function from the age of - years ( ), reported that persistent wheezing, starting early in life, was associated with a decline in lung function in adult life and increased risk of exacerbations. a decline in lung function has also been associated with severe asthma exacerbations in adult asthma ( ) . a german birth cohort of more than children showed that the risk of development of persistent asthma at age years was significantly increased by early allergen sensitization in combination with exposure to high levels of perennial allergens early in life ( ) . in a recent study, predictors of subsequent troublesome symptoms amongst -year-old children with wheezing were large skin test responses to allergens and history of previous exacerbations and eczema ( ) . attempts to identify phenotypes of adult asthma include the use of unsupervised cluster analyses ( , ) aiming to group patients who share key features of asthma and airway inflammatory disease. haldar and colleagues ( ) grouped patients according to clinical symptoms and evidence of airway inflammation (based on sputum eosinophil count) and found that symptom-led treatment of airway inflammation was appropriate in patients with mild and moderate asthma, but failed in more severe forms. they identified two discordant groups of patients who were refractory to standard treatment approaches and represented about % of all patients. one discordant group of patients were characterized as an obese, symptom-predominant, noneosinophilic phenotype, while the other group had few asthma symptoms, but a high degree of airway inflammation. using symptom-driven asthma treatment in these two discordant groups would lead to overtreatment of the former and undertreatment of the latter group. one difficulty with attempts to use clustering techniques to define disease mechanisms is defining cause and effect. in other words, 'are the pathophysiological features a consequence or an underlying cause of the disease?' a further confounding factor is the influence of treatment or lack of it (nonadherence). defining groups of patients with chronic diseases according to different personality traits ( ) has identified personality traits that govern treatment adherence. there was a significant overlap between the phenotypes identified by haldar et al. ( ) and those identified by moore et al. ( ) . other similar initiatives included the eu-sponsored unbiased biomarkers for the prediction of respiratory disease outcomes (u-biopred) consortium that has published a consensus-based systematic algorithm approach to differentiate between 'problematic', 'difficult' and 'severe refractory' asthma in the evaluation of patients with chronic severe asthma symptoms for use in clinical research and specialized care ( ) . the published practal consensus report on 'endotypes' ( ) attempts to assign patients with asthma to groups sharing specific pathophysiological features, with the aim of identifying a basis for rational treatment of heterogeneous groups. patients with severe asthma are found in each of these groups. three recognizable clinical phenotypes of severe asthma emerge from these various analytical studies (see table ): (i) a severe atopic form, (ii) severe 'intrinsic' asthma (the most malignant severe asthma phenotype ( )) and (iii) severe asthma with obesity. however, it has to be emphasized that distinct pathophysiological mechanisms underpinning these different clinical phenotypes of severe asthma have not as yet been identified. one potential problem of the cross-sectional approach to unbiased clustering is that the analysis does not include the important dimension of time, which may be essential to take into account potentially crucial longitudinal changes. furthermore, while unsupervised learning may be a useful tool to generate new hypotheses, using these techniques to find an association with predefined outcomes such as severe asthma can be misleading if asthma severity is derived using the same variables that are used for clustering. there is mounting evidence to demonstrate a close association between sensitisation to fungi and asthma severity ( ) , and the term 'severe asthma with fungal sensitization' (safs) has been proposed for patients with persistent severe asthma and fungal sensitization ( ) . proof-of-concept pilot studies have suggested an improvement in asthma after antifungal treatment in this patient group ( ) . several studies have shown that the standard guideline treatment of persistent asthma with ics provides a variable response, with - % of asthmatic subjects showing little improvement in fev and/or bronchial hyper-responsiveness ( ) ( ) ( ) . in the gaining optimal asthma control ('goal') study ( ) , bateman and colleagues used year of increasing doses of combined ics and laba treatment in patients with persistent asthma and reported that about % of patients, with varying degrees of disease severity, did not achieve 'well-controlled asthma'. asthma exacerbation risk may be further reduced using the same medication (i.e. an inhaled combination of a rapid-acting beta- agonist and corticosteroid ( )) as a controller and as a reliever of the disease. the price trial ( ) aimed to identify predictors of short-term ( weeks) response to ics. although several baseline biomarkers and asthma symptoms correlated with short-term improvements, only greater bronchodilator (short-acting b agonist) reversibility showed a strong (p < . ) correlation. differentiating between responders (> % fev improvement) and nonresponders (< % fev improvement) revealed that asthma control in responders was maintained only with continued ics, while, in nonresponders, no improvement was observed with or without ics. interestingly, tiotropium, a long-acting antimuscarinic agent, still approved only for copd, has been recently shown to further improve lung function in patients with severe uncontrolled asthma ( ) . of the four clinical phenotypes of severe asthma described in table , the severe atopic form partially responds to standard treatment. table indicates possible treatment approaches in the two groups of patients with severe asthma, which were described by haldar et al. ( ) , that are less responsive to standard treatment. these treatment options for patients with severe asthma who remain symptomatic despite adhering to standard medical care include novel anti-inflammatory drugs that have been shown in preliminary studies to be effective in treating airway inflammation in asthma and so warrant further investigation ( , ( ) ( ) ( ) ( ) , and other novel approaches such as bronchial thermoplasty ( ) . it is important to note that, for a more effective use of these novel treatment options, a better understanding of the pathophysiology and of the inflammatory mechanisms of the severe asthma subtypes are required in order that studies can delineate specific response patterns. as previously noted, antifungal treatments may be of benefit in patients with severe asthma with fungal sensitization ( , ) , but large studies are needed to support these initial findings. current treatment of asthma exacerbations is inadequate, and new approaches to treatment are needed. these may include interferon-based treatments, treatments that aim to boost deficient antiviral immune responses and/or specific antiviral treatments. there may be patients with severe asthma who could benefit from treatments that target neutrophilic inflammation without further suppressing anti-infective immunity. another question that needs to be addressed is the possible benefit in asthma exacerbations of antibacterial treatments that are widely used ( ), despite not being recommended in guidelines. the advances in molecular biology and immunology are being used to develop novel biological drugs for asthma treatment. these include therapeutic antibodies, soluble receptors, cytokines, small molecules and combinations thereof that target different effector molecules that influence the underlying immune and inflammatory processes. some biological drugs are currently in clinical trials in asthma ( ) . however, there are major difficulties in developing novel drugs to treat asthma. these include (i) the complexity of the disease (in terms of the different disease phenotypes and the underlying molecular mechanisms), (ii) the limited number of biomarkers that have been identified for disease classification, (iii) the effectiveness of the current standard treatment approaches (combined inhaled steroid and beta-adrenergic agonist is not only effective but cheap) makes any comparative improvement difficult to identify in multicentre clinical trials, (iv) low patient adherence, which is characteristic of treatments of chronic disease, and (v) preclinical animal models may be poorly predictive of clinical efficacy ( ) . importantly, future clinical trials will need to identify the patient groups that respond to novel treatment to provide evidence for the stratified, personalized approach to asthma management. in primary care, recognition of the clinical heterogeneity of asthma and the existence of different forms of severe asthma is obscured by a number of general deficiencies, including limited available time and clinical resources and lack of capacity and clinical capability. nonguideline treatment of asthma is high, and provision of treatment to patients with asthma varies greatly, for example, in the use of ics across european countries ( ) . the influence of comorbidities and of asthma exacerbations on asthma symptoms, in both children and adults, are not sufficiently recognized ( , ) . most patients with asthma in primary care have been found to have uncontrolled disease ( ) . furthermore, it is becoming increasingly clear that many patients diagnosed as asthmatic, even after full evaluation at tertiary centres, do not have asthma, but various other diagnoses ( , ) , which indicate a fundamental need to establish improved diagnosis as a basis of appropriate management ( ) . the resources needed to review patients frequently until disease control is achieved vary between healthcare systems. the finnish national asthma programme ( ) , predicated on a systematic approach to disease management and underpinned by educational and skills training in primary care, demonstrates that investment in the structure of the health system that delivers asthma care reduces morbidity significantly and at a lower overall cost. the above discussion aims to highlight the prominent issues and the attempts to acquire a better understanding of asthma exacerbations and severe asthma. however, there are several additional clinical and pathophysiological issues of importance for improving our understanding of asthma as a complex disease. these have not been highlighted here partly because there is no scientific consensus of their exact relevance. however, two examples are briefly reviewed here. airway remodelling is associated with poor clinical outcomes amongst asthmatic patients, but the pathophysiological relevance of, and the effect of treatment on, airway remodelling is unclear ( ) . airway remodelling is a feature of adult asthma and is found to a similar extent, and quite early, in children with difficult-to-treat asthma ( ). using epithelial reticular basement membrane thickening as a marker for airway remodelling, no association between airway remodelling and age, symptom duration, lung function and concurrent eosinophilic airway inflammation was found ( ) , albeit in a small number of patients. inflammatory and structural changes typical of asthma, such as airway eosinophilia and angiogenesis, have been observed not only in children with asthma but also in atopic children without asthma, raising the possibility that some of these pathological lesions may be associated with atopy even in the absence of asthmatic symptoms. reticular membrane thickening and the eosinophilic inflammation characteristic of asthma in older children and adults are not present in the wheezing infants with reversible airflow obstruction, even in the presence of atopy ( , ) . it was proposed that this lack of rbm thickening in wheezy infants was due to the apparent paucity of eosinophilic inflammation ( ) , which may have a role in driving allergic airway remodelling ( ) . based on the data from many population studies, it is generally considered that the environment has an important influence on the development of asthma and other allergic diseases. however, the proposed mechanisms, particularly the role of atopy, that underlie the effects of environment on asthma aetiology need to be revised in the light of recent findings ( ) , such as the evident increase in prevalence of severe asthma in developing countries and its parallel decline in several developed countries ( ) . the interaction of environment and asthma disease mechanisms is complex, and genetic variants that are protective in one environment may be associated with increased risk in another environment ( ) . • although clinical guidelines improve patient treatment by bringing treatments to groups of patients who best benefit from them, they fail many patients who fall outside of the 'mean' clinical characteristics on which clinical guidelines are based. labas is insufficiently effective in many patients with severe asthma. • young children, particularly those with virus-driven asthma, are often poorly managed with currently available medications. • in asthma, there is a need to focus more on the individual patient, particularly those with severe asthma. • there is a need to recognize asthma as a heterogeneous disease and to properly identify the different disease mechanisms involved in patients with severe asthma. only in this way, we will begin to understand what is 'driving' severe asthma and identify novel therapeutic targets. • in childhood asthma, expression of atopy varies over time and in different characteristic ways that suggest differences in underlying pathophysiological mechanisms in relation to exacerbation-prone asthma phenotype. • in adult asthma, several forms of severe asthma have become recognized, and different treatment approaches to these forms of severe asthma are proposed. • there is a need for a consensus definition of asthma exacerbation that could usefully guide treatment. we need to better understand the mechanisms of asthma exacerbation, develop novel treatments for exacerbations and carry out studies of existing and new treatments to better guide their use. • as in copd, asthma diagnosis and treatment decisions should include consideration of future risk of exacerbations. • current data strongly link impaired innate immune responses and consequent increased risk of infection with increased risk of asthma exacerbations. this provides a sound basis for the development of novel treatments. • there is general concern that patients with asthma are not being sufficiently alerted to the risk of asthma exacerbations and that another term be considered other than 'asthma exacerbation' when describing these events to patients (such as 'lung attack' or 'asthma attack' to equate the seriousness of such an event with a heart attack). • development of diagnostic procedures accessible within primary care to ensure correct diagnosis. • an acceptance of the need for iterative review to gain control of the disease; patients, in whom asthma fails to become controlled, need further evaluation to confirm that asthma is the correct diagnosis and what further evaluations are required. • improved tools for 'scoring' the asthma patient in terms of disease severity, and future risk needs to be developed for use in primary care ( , ) . • future studies should aim to establish whether serum ige, f e no and induced sputum eosinophil counts are valuable biomarkers of the severity of asthmatic airway inflammation. • new biomarkers are needed to predict the severity of asthma, the risk of exacerbation and the response to treatments. • there is a need for a more objective definition of childhood asthma, with broader use of lung function tests, especially in preschoolers, and so obtain more uniform criteria for diagnosis before initiating treatment to control the disease. • in both childhood and adult asthma, expression of atopy varies over time and in different characteristic ways that reveal differences in underlying pathophysiological mechanisms, including a severe asthma-prone phenotype. • observational longitudinal studies should be performed in a standardized way with patient and biological data sampling (using an asthma register) to allow a better characterization of the epidemiology, current healthcare utilization, risk factors (including genetic susceptibility) and comorbidities that are linked to exacerbations and severity. • the future task is to devise studies that differentiate between patient phenotypes, use robust clinically relevant biomarkers, include longitudinal outcomes (such as time to exacerbation and increase in airway remodelling) and identify relevant environmental factors, including the influence of current and prior medication. to facilitate the identification of new asthma endotypes, such studies should contribute to a repository (or biobank) of biological samples from the asthma population. • for characterization of the heterogeneity of severe asthma, more and improved clinical studies are needed to improve the evidence base. • randomized, controlled trials in asthma include highly selected patients, who are, almost exclusively, 'healthy asthmatics' with ß -agonist reversibility. • cohort studies may complement clinical trials. • there are still insufficient longitudinal data in adult asthma, and cohort studies in this age group need to be established. • improving primary care of the asthmatic patient is in need of urgent attention and requires radical changes. • the ideal approach to dealing with the asthmatic patient in primary care is to (i) characterize the patient, (ii) confirm diagnosis, (iii) confirm whether a new or existing patient gains control by the iterative application of a structured review ( ) and consider reducing treatment when control is achieved, while undertaking appropriate monitoring, (iv) maintain control by teaching the patient about asthma and developing the patient's self-management skills ( ) according to the model developed by glasziou ( ) , (v) attempt to understand the patient's perspective; this may be the key to improving patient compliance ( ) , (vi) consider the risk of exacerbations (which is separate from controlling everyday symptoms) and factors that may increase the risk of exacerbations ( ) and (vii) realize that nonsymptomatic patients are also at risk of exacerbations. • the complexity of the multifactorial nature of asthma and the resource limitations in primary care needs to be better characterized in order to address the wide variability of care delivered in this environment ( ) . • support mechanisms that enable readily accessible means for patient self-management and self-education, having already met with varying degrees of success ( ) ( ) ( ) ( ) , need to be further developed. • primary clinicians have themselves recognized the research agenda that is needed to direct improvement in asthma care ( ). • whether seen in primary care or secondary care, patients with uncontrolled asthma either (i) have treatment-resistant disease, (ii) are not fully compliant with treatment, (iii) are unable to appreciate deterioration in their disease, (iv) have a physician who is underestimating the degree of disease control, is undertreating or not recognizing the effect of comorbidities, or (v) do not have asthma. l. m. fabbri has received consultancy fees from: boehringer ingelheim, chiesi, glaxosmithkline, msd, nycomed, pearl therapeutics, sterna, peer voice europe, om pharma sa, kyorin pharmaceutical, boston scientific and bayer. readings, advisory board or reimbursement of expenses: astrazeneca, novartis, sigma-tau, roche, deutsches zentrum f€ ur luft und raumfahrt, german aerospace center, mundipharma int., genetech inc., elevation pharmaceutical, ferrer group, nycomed, dynamicon and laboratori guidotti. e. bel has received consultancy fees from novartis, gsk and sanofi-regeneron, and fees for speaking from gsk. p. le sou€ ef has received speaker fees from glaxo smith kline and astrazeneca and has received research funding from astrazeneca and pharmanet ag. he has received research grants from the national health and medical research council of australia and the australian research council. j. l€ otvall has over the last five years been a consultant and/or given lectures for gsk, astrazeneca, aerovant, novartis, ucb, oriel and merck, for which he has received honoraria. he has also received research grants and participated in clinical trials for novartis, gsk, astrazeneca and actelion under the full organization of the university of gothenburg. p. demoly is a consultant and a speaker for stallergenes, alk, circassia and chiesi and is a speaker for merck, astrazeneca, menarini and glaxosmithkline. c. akdis serves 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monitoring of asthma: multicentre randomised controlled trial slj is funded by the asthma uk clinical professorship (ch sj). we acknowledge the editorial support of gerard p mcgregor, of omniscience sa, who was financed by a grant from the eaaci. the authors wish to thank professor christian virchow, universit€ at rostock medizinische fakult€ at, klinik & poliklinik f€ ur innere medizin, abteilung pneumologie, ernst-heydemann-str. , rostock, germany, for reviewing the manuscript and extremely useful comments. a. custovic serves as a consultant for circassia. he received speaker fees from glaxo smith kline, thermo fisher scientific, airsonet, novartis, msd and alk. he received research grants from the uk medical research council, moulton charitable foundation national institute of health research.in the past five years s. l. johnston has had research grants from astra zeneca, centocor, glaxosmithkline, med-immune, sanofi-pasteur and synairgen. s. l. johnston holds share options in synairgen. s. l. johnston does some consultancy work for astrazeneca, centocor, glaxosmithkline, medimmune, sanofi-pasteur and synairgen.in the last five years, i. d. pavord has received speaker's honoraria for speaking at sponsored meetings from astra zeneca, boehringer inglehiem, aerocrine and gsk. he has received honoraria for attending advisory panels with almirall, astra zeneca, boehringer ingelheim, gsk, msd, schering-plough, novartis, dey and napp. he has received sponsorship to attend international scientific meetings from boehringer ingelheim, gsk, astra zeneca and napp.m. gaga received research grants from novartis, bi, cephalon, teva and gsk. the other authors of the paper declare no conflicts of interest. key: cord- -anvmj li authors: liu, xinkui; yue, xinpei; liu, furong; wei, le; chu, yuntian; bao, honghong; dong, yichao; cheng, wenjie; yang, linpeng title: analysis of clinical features and early warning signs in patients with severe covid- : a retrospective cohort study date: - - journal: plos one doi: . /journal.pone. sha: doc_id: cord_uid: anvmj li coronavirus disease (covid- ) was first identified in wuhan, china, in december . although previous studies have described the clinical aspects of covid- , few studies have focused on the early detection of severe covid- . therefore, this study aimed to identify the predictors of severe covid- and to compare clinical features between patients with severe covid- and those with less severe covid- . patients admitted to designated hospital in the henan province of china who were either discharged or died prior to february , were enrolled retrospectively. additionally, patients who underwent at least one of the following treatments were assigned to the severe group: continuous renal replacement therapy, high-flow oxygen absorption, noninvasive and invasive mechanical ventilation, or extracorporeal membrane oxygenation. the remaining patients were assigned to the non-severe group. demographic information, initial symptoms, and first visit examination results were collected from the electronic medical records and compared between the groups. multivariate logistic regression analysis was performed to determine the predictors of severe covid- . a receiver operating characteristic curve was used to identify a threshold for each predictor. altogether, patients were enrolled in our study with and patients in the severe and non-severe groups, respectively. multivariate logistic analysis indicated that patients aged ≥ years (odds ratio = . ; % ci: . , . ), with an absolute lymphocyte value of ≤ . × ( )/l (odds ratio = . ; % ci = . , . ) and a c-reactive protein level of ≥ . mg/l (odds ratio = . ; % ci = . , . ) were at a higher risk of severe illness. thus, our results could be helpful in the early detection of patients at risk for severe illness, enabling the implementation of effective interventions and likely lowering the morbidity of covid- patients. a a a a a in december , a local cluster of patients with pneumonia of unknown etiology was observed; subsequently, similar cases were observed nationally. on january , , the virus strain causing pneumonia was successfully isolated by researchers [ , ] . this virus is genetically related to the virus responsible for the severe acute respiratory syndrome-related coronavirus (sars-cov) outbreak in [ ] ; consequently, the new virus was named the severe acute respiratory syndrome coronavirus (sars-cov- ) by the international committee on taxonomy of viruses on february . on the same day, the world health organization (who) proclaimed the official name of the disease caused by the new virus as coronavirus disease (covid- ) [ ] . as covid- continued to spread and developed into an epidemic, the who declared covid- a "public health emergency of international concern" [ ] . as of march , there were , confirmed cases reported in china [ ] , of which , cases were in the henan province [ ] . a study by zhou et al. reported the risk factors for mortality of covid- inpatients in wuhan [ ] , but the thresholds for each risk factor was not given. to our knowledge, there have been no studies to predict patients with severe covid- from the results of their first visit. the objective of this study was to determine the risk factors for severe covid- based on the test results from the patient's first visit, and describe the clinical features in patients with severe covid- . our study population consisted of confirmed covid- patients who were either discharged or died prior to february , . our study was launched on march , . participants with the following missing information were excluded from the study: age, date of first symptom, contact date, or examination data. all demographic and clinical information was collected retrospectively from the electronic medical records (emr). patients who underwent at least one of the following procedures were assigned to the severe group: continuous renal replacement therapy (crrt), high flow oxygen absorption, noninvasive and invasive mechanical ventilation, and extracorporeal membrane oxygenation (ecmo). the remaining participants were assigned to the non-severe group. data obtained from the emr included age, sex, contact history, latest contact date, primary symptoms and date of occurrence, comorbidities, and the results of routine blood tests as well as liver function, kidney function, c-reactive protein (crp), and procalcitonin tests from the initial visit. confidentiality of information was maintained by removing personal identifiable information. the medical research ethics committee of the first affiliated hospital of zhenghzou university approved this study. the following comorbidities were noted among our participants: respiratory diseases such as chronic obstructive pulmonary disease (copd), asthma, and interstitial pneumonia; metabolic diseases such as diabetes mellitus; cardiovascular disease (cvd) such as hypertension and coronary heart disease; and neurological diseases such as cerebral hemorrhage and infarction. a clustered onset was defined as two or more cases with fever or respiratory symptoms in a confined area, such as a home, office, school class, or similar setting, within the previous two weeks. the incubation period was defined as the time between the date of occurrence of primary symptoms and latest contact date with known infected individuals. data from the emr were entered using epidata and confirmed twice. every case was entered independently by two different data entry clerks and consistency was verified by a supervisor. three members of our research group were responsible for reviewing each data entry for which conflicting data had been entered. the doi link is dx.doi.org/ . / protocols.io.bfpejmje. all data were entered and managed in epidata. spss . was utilized for cleaning and analyzing the data. as the quantitative data did not follow a normal distribution, the data were expressed as medians and interquartile ranges (iqrs) after applying the mann-whitney u test for comparison between the groups. qualitative data were expressed using counts and percentages and compared using the chi-squared or fisher's exact test. thresholds were calculated using receiver operating characteristic (roc) curves. independent effects were shown using multivariate logistic regression. based on the univariate analysis results and clinical experience, five variables were chosen for the multivariable analysis including age, comorbidity, lymphocyte count, crp levels, and procalcitonin levels. the significance level was set at . . in the henan province, there were a total of confirmed covid- patients with a final outcome of discharge or death prior to february . after excluding cases with missing values, participants were assigned to then on-severe group and participants were assigned to the severe group. the median age of the study participants was . (iqr = . - . ) years, ranging from to years. there were male patients, accounting for . % of all participants and female patients, accounting for . % of all participants. of the participants, ( . %) had a history of traveling or residing in either wuhan or the surrounding areas, or other communities with reported covid- cases, while participants ( . %) had contact with individuals who were experiencing fever or respiratory symptoms and had recently been to wuhan or its surrounding areas, or other communities with reported cases. furthermore, ( . %) patients had a clustered onset, ( . %) had a history of contact with a covid- patient, and ( . %) had an unknown history of exposure. the median incubation period for the entire group was . (iqr = . - . ) days, ranging from days to day. an incubation period of less than days was observed in . % of the patients. further, ( . %) participants had underlying diseases, with cardiovascular disease being the most common underlying disease, followed by metabolic and respiratory disease. the main symptoms of the participants included fever, cough, and fatigue. fever was the most common initial symptom, manifesting in patients ( . %) on admission ( table ) . the median age in the non-severe group ( years) was significantly higher than in the severe group ( years). underlying diseases manifested in ( . %) patients in the severe group, which was higher than in the non-severe group, and the difference was statistically significant (p< . ). the proportion of patients with cardiovascular disease was also greater in the severe group than in the non-severe group, and the difference was statistically significant (p < . ). with regard to symptoms, the proportion of patients with dyspnea was significantly higher in the severe group than in the non-severe group (p < . ). the proportions of patients with fever, cough, asthma, and chest distress were greater in the severe group; however, these differences were not significant. other symptoms demonstrated no quantifiable difference between the severe and non-severe groups ( table ) . the majority of factors examined at first admission indicated a non-significant difference between the two groups; however, direct bilirubin, lactate dehydrogenase, crp, and procalcitonin levels were significantly higher in the severe group than in the non-severe group (p < . ). additionally, the absolute value of lymphocyte count was also significantly lower in the severe group than in the non-severe group (p < . ) as shown in table . as demonstrated by the roc curves using a single predictor, age, the absolute lymphocyte value, crp, and procalcitonin are valuable predictors for detecting severe conditions in patients; the area under the curve (auc) for these predictors were . , . , . , and . , respectively. further, the thresholds for these factors were for age, . × ^ /l for lymphocyte, . mg/l for crp, and . μg/l for procalcitonin (figs - and table ). the results of the multivariate logistic regression indicate that age, lymphocyte, and crp are independent predictors for an increased risk of severe covid- . patients who are at least years old, with an absolute lymphocyte value less than . × ^ /l, and a crp greater than . mg/l are at greater risk of developing severe covid- with corresponding odds ratios of . , . , and . , respectively (table ). an analysis of the roc curves with multiple predictors is shown in fig . these results have a high degree of accuracy as implied by an auc of . ( %ci = . , . ). our study indicates that age, the absolute lymphocyte count at initial visit, and crp may be used as predictors during the early stage of diagnosis in patients who are at risk of developing severe covid- . in many severe cases, patients have cvd as a comorbidity, dyspnea, and higher concentrations of direct bilirubin, lactate dehydrogenase, and procalcitonin. in our study, the median age of the participants was . years (iqr = . - . ) ranging from to years of age. in addition, for . % of the patient, the incubation period was less than seven days, which indicates that covid- has a short incubation period and may impact both children and adults. the same result shave been reported in previous studies [ ] . the median age in the severe group was . years (iqr = . - . ), which is higher than the median age in the non-severe group, (median = . ; iqr = . - . ). the threshold for age was years, as indicated in the roc curve analysis. the threshold age is a predictor for an increased risk of severe covid- . in addition, our multivariate logistic regression analysis implied that patients who are older than years are at higher risk of developing severe covid- (or: . ). the trends indicated in our study are similar to those indicated in previous studies [ ] . moreover, studies show that immune responses in older adults are slower, less coordinated, and less efficient, rendering older adults more susceptible to emerging infections [ ] . a study by shahid et al. indicates that the probability of having multiple comorbidities increased the risk of mortality from sars-cov- in older adults [ ] . our data indicate that . % of patients had an underlying disease, especially cardiovascular and metabolic disorders. the percentage of patients suffering from cvd was higher in the severe group than in the non-severe group. these results have been verified by epidemic reports from the chinese center for disease control and prevention [ ] . as in previous studies, the principal symptoms of covid- are respiratory symptoms. some patients also suffer significant cardiovascular damage from covid- . coupled with underlying cvd, these complications may increase the risk of death for some patients. covid- results from a spike protein in the virus binding with the angiotensin-converting enzyme (ace ) which is highly expressed in the heart and lung tissue. a contributing factor to severe covid- in patient with cvd may be the increased concentration of ace in these patients. as the middle east respiratory syndrome-related coronavirus (mers-cov) and severe acute respiratory syndrome coronavirus (sars-cov) may result in acute myocarditis and heart failure, acute coronary syndrome patients infected with the novel coronavirus are at an increased risk of cardiac insufficiency that might lead to severe covid- or death [ ] . the most common symptoms of covid- in our study were fever ( . %), cough ( . %), and fatigue ( . %). some patients also experienced dizziness ( . %), a runny nose ( . %), nasal obstruction ( . %), and diarrhea ( . %). these numbers are similar to those described in previous studies [ , ] . our study indicates that in . % of the patients, the initial symptom was fever. in addition, . % of the patients had fever during the inpatient period; of those, all patients with severe covid- had fever. as shown in previous studies, although only . % of the patients had fever on admission, . % of all patients had fever during hospitalization [ ] . this implies that fever did not present as an initial symptom in all covid- patients; nonetheless, fever occurred as pneumonia symptoms developed. fever that developed as a result of critical pulmonary infection was frequently observed in patients with severe covid- ; therefore, temperature monitoring should not be the only screening measure for covid- . we suggest that in addition to temperature monitoring, the patient's contact history and symptoms should be examined to identify individuals who require observation. a higher number of patients showed signs of dyspnea in the severe group than in the non-severe group in our study, indicating that dyspnea was one of the main symptoms among patients with severe covid- ; thus, a daily evaluation for dyspnea should be conducted among covid- patients. wang et al. suggest that more frequent chest ct scans should be performed for patients with severe dyspnea to understand the changes and indications in pulmonary imaging [ ] . all cases in our covid- study had a decreased lymphocyte count on admission; patients in the severe group had even lower absolute lymphocyte values, as indicated by a median of . (iqr = . - . )× ^ /l. similar changes in the lymphocyte counts were observed in patients with sars [ , ] . a previous study regarding lymphocyte subsets indicated that covid- patients had reduced count of total lymphocytes, cd + t cells, cd + t cells, b cells, and natural killer cells. furthermore, severe covid- cases had lower counts than non-severe cases [ ] . a study by qin et al. suggests that covid- might damage lymphocytes, especially t lymphocytes; consequently, the immune system is impaired during the course of the disease [ ] .the coefficients from our multivariate regression analysis indicated that patients with an absolute lymphocyte value of � . × ^ /l have an increased risk of experiencing severe covid- (or = . ). this implies that a decrease in the lymphocyte count should be a key predictor in the early diagnosis of severe covid- . research on sars has shown similar results; a decrease in the lymphocyte count could be an early warning of severe disease [ , ] . this study indicates that patients with elevated concentrations of serum crp on admission are at an increased risk of experiencing severe covid- . the risk of developing severe covid- in patients with a serum crp of � . mg/l is . times than in patients with a serum crp� . mg/l. at elevated concentrations, crp, which is an acute-phase protein, is correlated with an increased risk of organ failure and death for patients admitted to the icu. further, prolonged periods of high crp concentrations are associated with adverse outcomes [ ] . previous research in wuhan indicated that increased levels of crp were indicative of a sustained inflammatory response subsequent to infection with sars-cov- . additionally, patients with severe covid- had more prominent inflammation [ ] . similar trends of elevated serum crp were also found in sars patients [ ] ; a previous study shows that a crp concentration of > . mg/l is correlated with an increased risk of death for sars patients (or = . ) [ ] . our study has several limitations. first, due to the limited ability to collect data and the relatively low morbidity and mortality of covid , the sample size in this study is small, which resulted in relatively few variables being included in the multivariate analysis; it is possible that some significant variables may have been neglected. second, as this study is a retrospective study, the data collected from the electronic medical records are limited; thus, data about additional significant factors may have not been available. for example, we were unable to analyze the possible causes of higher procalcitonin levels because we lacked data on bacterial infections. in this study, we summarized the clinical features of covid- patients and identified the early warning signs of severe covid- , which will help physicians determine which patients require further observation. although this study has some limitations, including a small sample size, few variables included in the multivariate analysis, a retrospective cohort design, and limited data collected from medical records, the results of our study indicate that older age, a decreased lymphocyte count on admission, and an increased concentration of serum crp could serve as early warning signs in patients who are at risk of developing severe covid- . consequently, we suggest that patients with these clinical characteristics be monitored closely. further studies should be conducted to confirm the results of our study. conceptualization: xinkui liu, xinpei yue. the novel coronavirus originating in wuhan, china: challenges for global health governance severe acute respiratory syndrome-related coronavirus: the species and its viruses-a statement of the coronavirus study group a pneumonia outbreak associated with a new coronavirus of probable bat origin world health organization. who director-general's statement on ihr emergency committee on novel coronavirus ( -ncov) update on the epidemic situation of covid as of : on march update on the epidemic situation of covid in henan province as of : on march clinical course and risk factors for mortality of adult inpatients with covid- in wuhan, china: a retrospective cohort study epidemiology and transmission of covid- in shenzhen china: analysis of cases and , of their close contacts neutrophil-to-lymphocyte ratio predicts severe illness patients with novel coronavirus in the early stage sars-cov- and covid- in older adults: what we may expect regarding pathogenesis, immune responses, and outcomes covid- and older adults: what we know epidemiology working group for ncip epidemic response, chinese center for disease control and prevention. the epidemiological characteristics of an outbreak of novel coronavirus diseases (covid- ) in china covid- and the cardiovascular system clinical features of patients infected with novel coronavirus in wuhan clinical characteristics of hospitalized patients with novel coronavirus-infected pneumonia in wuhan clinical characteristics of coronavirus disease in china insights on diagnosis and treatment of coronavirus disease a major outbreak of severe acute respiratory syndrome in hong kong clinical features and short-term outcomes of patients with sars in the greater toronto area characteristics of peripheral lymphocyte subset alteration in covid- pneumonia dysregulation of immune response in patients with covid- in wuhan, china measurement of subsets of blood t lymphocyte in patients with severe acute respiratory syndrome and its clinical significance changes of blood cells in patients with severe acute respiratory syndrome c-reactive protein levels correlate with mortality and organ failure in critically ill patients clinical characteristics of patients infected with sars-cov- in wuhan sars in singapore-predictors of disease severity hematological and biochemical factors predicting sars fatality in taiwan key: cord- -ivwz jxi authors: anzola, gian paolo; bartolaminelli, clara; gregorini, gina alessandra; coazzoli, chiara; gatti, francesca; mora, alessandra; charalampakis, dimitrios; palmigiano, andrea; de simone, michele; comini, alice; dellaglio, erica; cassetti, salvatore; chiesa, maurizio; spedini, francesca; d’ottavi, patrizia; savio, maria cristina title: neither aceis nor arbs are associated with respiratory distress or mortality in covid- results of a prospective study on a hospital-based cohort date: - - journal: intern emerg med doi: . /s - - - sha: doc_id: cord_uid: ivwz jxi considerable concern has emerged for the potential harm in the use of angiotensin-converting enzyme inhibitors (aceis) and angiotensin receptor inhibitors (arbs) in covid- patients, given that aceis and arbs may increase the expression of ace receptors that represent the way for coronavirus to entry into the cell and cause severe acute respiratory syndrome. assess the effect of acei/arbs on outcome in covid- patients. hospital-based prospective study. a total of patients consecutively presenting at the emergency department and found to be affected by covid- were assessed. relevant clinical and laboratory variables were recorded, focusing on the type of current anti hypertensive treatment. outcome variables were no, mild, severe respiratory distress (rd) operationally defined and death. hypertension was the single most frequent comorbidity ( / = %). distribution of antihypertensive treatment was: aceis / ( %), arbs / ( %), other than aceis or arbs / ( %). in / ( %) antihypertensive medication was unknown. the proportion of patients taking aceis, arbs or others who developed mild or severe rd was / ( %), / ( %), / ( %) and / ( %), / ( %) and / ( %), respectively, with no statistical difference between groups. despite producing a rr for severe rd of . ( % ci . – . ), hypertension was no longer significant in a logistic regression analysis that identified age, crp and creatinine as the sole independent predictors of severe rd and death. aceis and arbs do not promote a more severe outcome of covid- . there is no reason why they should be withheld in affected patients. in early december , a new pathogen, later identified as a novel enveloped rna betacoronavirus, that is currently been named severe acute respiratory syndrome coronavirus (sars-cov- ), gave rise to an outbreak of pneumonia that started from wuhan in the chinese province of hubei and spread across a large territory of the country infecting more than , subjects in less than a couple of months [ ] . despite the lack of a proven specific treatment, the infection was readily contained and limited mainly through stringent social distancing and quarantine, but before being able to close the borders, an unknown number of probably asymptomatic patients let the virus spread all over the world. italy was among the first european countries to be affected. the first case was reported by media on february as being hospitalized in codogno in the lombardia region (northern italy). henceforth, the surge of epidemics has followed an exponential rise that only recently has somehow flattened, with a cumulative prevalence of infected cases beyond , individuals, more than , cumulative deaths and a number of newly diagnosed cases that is still around a day according to the who published report as to july (https ://www.who.int/emerg encie s/disea ses/novel -coron aviru s- /situa tion-repor ts). sars-cov- has a phylogenetic similarity to sars-cov, responsible for the preceding - outbreak in china, with which it shares the propensity to attack the respiratory tract and cause a severe acute respiratory syndrome. despite being probably less lethal than sars, which had a reported case fatally rate of · % ( · - · ) in patients aged years or older [ ] , current coronavirus disease is not at all a benign disease. since the first reports from chinese authors, it has become clear that some variables such as male gender, cardiovascular disease, advanced age and hypertension may drastically worsen the prognosis [ ] [ ] [ ] . in particular hypertension has become the focus of a warm debate on the use of angiotensin-converting enzyme (ace) inhibitors (aceis) and angiotensin receptor inhibitors (arb), which have established for a long time a mainstay in the treatment of hypertension [ ] [ ] [ ] . the main reason of apprehension resides in the fact that ace receptors, which belong to the renin-angiotensin system and are widely represented in many organs, including pulmonary alveoli, represent the way for coronavirus (sars-cov- ) to entry into the cell. since some laboratory data suggest that long lasting use of aceis or arbs may upregulate ace receptors, considerable concern has emerged for the potential harm in their use as they might cause an increased susceptibility to viral penetration into the respiratory cells and give way to a more serious disease [ ] [ ] [ ] [ ] [ ] . partly to help solving this issue, we undertook a prospective study aimed at assessing the clinical characteristics, with particular emphasis on the type of antihypertensive medication, of all consecutive patients presenting at the emergency department of a community hospital in gavardo, in the neighborhood of brescia in lombardia (italy), and found to be positive for sars-cov- infection. the place of recruitment was the emergency department of the hospital of gavardo, situated in the neighborhood of brescia in lombardia and belonging to the local health authority (lha) asst garda. this lha serves a territory of about square kilometers on the western border of the lake of garda with a population of about , inhabitants. these are evenly distributed between the three community hospitals located in the cities of gavardo, desenzano and manerbio. therefore the estimated catchment area of the gavardo hospital is about , people. all consecutive patients presenting at the emergency department for symptoms or signs suggestive of sars-cov- infection were considered for the study, but only those confirmed by real-time rt-pcr in nasal or pharyngeal swab were included. on admission, a structured interview assessed demographic, anthropometric variables when possible and comorbidity. in particular a dichotomous categorization (yes/no) was employed for ischemic heart disease (including history of myocardial infarction, percutaneous transcatheter coronary angioplasty, coronary artery by-bass grafting), heart failure (present or past), stroke, atrial fibrillation, chronic renal failure, chronic liver disease, diabetes mellitus, chronic obstructive pulmonary disease (copd), history of or present neoplasm, history of or present autoimmune disease, hypertension, current use of aceis, current use of arbs, current use of antihypertensives other than aceis or arbs (others). relevant laboratory tests were recorded at presentation: these included hemoglobin (hb) in g/lt, platelet, leukocyte and lymphocyte count per microliter, serum creatinine in mg/dl, aspartate aminotransferase (ast), alanine aminotransferase (alt) in units per liter and c-reactive protein (crp) in mg/liter. in case of missing data, the electronic chart was reviewed on discharge to complete ascertainment. duration of symptoms from onset to presentation and cutaneous temperature were also recorded. dichotomous outcome variables were hospitalization, imaging evidence of lung infiltrates [ ] and being dead or alive at the end of the hospital stay. clinical severity was categorized as: no respiratory distress (no rd = sat. o > % and no shortness of breath), mild respiratory distress (mild rd = sat. o ≤ %, or shortness of breath or need for non-invasive ventilation, no need of invasive mechanical ventilation), and severe respiratory distress (severe rd = need of invasive mechanical ventilation). due to the often dramatic clinical emergency, some variables were incompletely assessed in a non-significant proportion of patients except for bmi that was available in only / ( %). the prevalence of each categorical variable and the means of continuous variables were calculated in patients with no, mild or severe rd and in deceased subjects. in discharged patients, the clinical state was assessed after a mean of ± days by telephone call. single comparisons were performed with chi-square test or fisher's exact test when appropriate on categorical variables and with independent sample t test (if normally distributed) or with mann-whitney u test on continuous variables. for multiple comparisons of continuous variables, univariate anova was used. variables that had proved significant on univariate analysis were entered multivariate binomial logistic regression analyses to identify independent predictors of mild rd, severe rd and death. statistical significance was set at p < . . spss statistical package was used. findings on death are only partially reported as they will thoroughly be dealt with in a separate paper. the study was approved by the local institutional review board. from march , , to april , , the study enrolled patients (m/f = / , mean age ± , range - ). imaging was positive for lung infiltration in ( %) patients. ( %) were hospitalized. mean duration of symptoms and temperature at presentation were ± days and . ± °c, respectively. the prevalence of no rd, mild rd or severe rd was / ( %), / ( %) and / ( %), respectively. case fatality rate (cfr) was / ( %). in the whole cohort, comorbidity was distributed as follows: ischemic heart disease / ( %), heart failure / ( %), stroke / ( %), atrial fibrillation / ( %), chronic renal failure / ( %), chronic liver disease / ( %), diabetes mellitus / ( %), copd / ( %), history of or present neoplasm / ( %), history of or present autoimmune disease / ( %), and hypertension / ( %). current use of aceis was recorded in / ( %), of arbs in / ( %) and of oth-ers in / ( %) patients. in / ( %) patients, it was impossible to establish the type of antihypertensive treatment. for comparisons between outcomes, the composite cardiovascular disease (cvd) variable was created including any among: ischemic heart disease, heart failure, stroke, and atrial fibrillation. for some continuous variables, the proportion of patients exceeding the cutoff lab was also calculated in the four outcome groups. compared with those with no rd, age, bmi, cvd, diabetes, hypertension, sat. o %, lymphocyte count, creatinine, proportion of ast ≥ , of creatinine ≥ . (lab cutoff) and crp were statistically different both in mild and in severe rd patients (tables , ). males were almost twice the number of females in both mild rd and severe rd groups. for ast and chronic renal failure, the difference from no rd was significant only in severe rd patients ( table ). the impact of significant variables on outcome is expressed as relative risk, compared to no rd, in table . some variables were able to discriminate also mild from severe rd: these were age ( ± vs. ± ), cvd ( % vs. %), creatinine ( . ± . vs. . ± . ) and crp ( ± vs. ± ). two roc curves were fitted to crp and creatinine values with severe rd as outcome of interest. the area under the curve was . for crp and . for creatinine. for crp, the threshold value of . correctly identified % of severe rd cases with . % of false positives, whereas for creatinine the threshold value of . was less efficient as it picked up only . % of cases with . % of false positives. the proportion of patients taking aceis, arbs or oth-ers who developed mild rd or severe rd was / ( %), / ( %), / ( %) and / ( %), / ( %) and / ( %), respectively, with no statistical difference between groups (fig. ) . likewise, the proportion of patients treated with aceis, arbs and other antihypertensives was roughly the same (around %) and constant within and across each class of severity ( table ) . three multivariate logistic regression analyses were performed, with mild rd, severe rd and death as dependent variables and taking as covariates those that had turned significant predictors in univariate analysis. the final model performed rather well, being able to predict . % of cases in mild rd, . % in severe rd and . % in death (tables , , ). age and crp were independent predictor of mild rd, severe rd and death, ast only of mild rd and creatinine of severe rd and death. in none of the three analyses was hypertension any longer significant. since the very beginning of the outbreak of covid- , it has become clear that hypertension is the most represented comorbid condition in affected patients [ ] , with a reported prevalence ranging from to . % in china [ , ] and % in italy [ ] . moreover, in univariate analyses hypertensive patients appear to be more likely to develop acute respiratory distress syndrome [ , ] , to be admitted in intensive care units [ , ] or to die [ , , ] , although this effect may become no more significant when corrected by age [ , ] . from many authorities, it has been suggested that taking aceis or arbs may amplify the expression of ace receptors that represent the way for sars-cov- to entry the respiratory cells and are widely represented in the alveolar cells, thus promoting the spreading of viral particles from upper to lower respiratory tract [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . however, the evidence for an increased activation of ace from aceis and arbs is far from being conclusive and moreover it may differ according to organ and acei type [ ] . furthermore, ace over expression has paradoxically been claimed to be potentially beneficial because it raises levels of angiotensin-( - ) which is increasingly recognized to have organ-protective properties [ , [ ] [ ] [ ] . to further complicate matters, arbs are believed to directly inhibit angiotensin ii-induced inflammation and acute injury in the lungs [ ] . therefore, there is much uncertainty as to whether aceis and arbs do really interfere with sars-cov- aggressiveness and only clinically driven conclusions may definitely settle the question. this was the main reason for undertaking the present study. in the consecutive series of proven covid- patients enrolled in one month period, table demographic, clinical and laboratory findings in no rd, mild rd and severe rd patients data shows mean (sd) for continuous or n (%) for categorical variables no no respiratory distress, mild mild respiratory distress, severe severe respiratory distress, bmi body mass index (weight/height ), acei angiotensin-converting enzyme inhibitor, arb angiotensin receptor inhibitor, copd chronic obstructive pulmonary disease, af atrial fibrillation, cvd cardio vascular disease (any among ischemic heart disease, heart failure, af, stroke), sat. o % blood oxygen saturation, other antihypertensive other than acei or arb, ast aspartate aminotransferase, alt alanine aminotransferase, cpr c-reactive protein * by stratifying patients in three levels of clinical severity, we compared the proportion of each class of drugs in each severity class and the other way around the proportion of each severity class in each class of drugs. the main finding was that there was not even the least hint that aceis or arbs behave differently from others in terms of increased frequency of worse outcomes. their relative frequency was the same, about %, among both mild rd and severe rd patients. likewise, the proportion of patients evolving toward mild or severe rd was exactly the same in the three therapeutic classes (see table and fig. ). we believe this is a clear demonstration that on clinical grounds aceis and arbs are in no way involved in enhancing the infectivity of sars-cov- and as a consequence, there is no reason why they should be withheld. our findings confirm the results of a previous study performed on a much smaller sample [ ] . as a corollary to the findings discussed above, it is worth noticing that when corrected for age and cvd, hypertension was no longer significant as a predictor of bad outcome. indeed it is difficult to understand why adequately treated hypertension should adversely affect an infectious disease with respiratory target. therefore, we would suggest that not so much hypertension but rather other factors usually correlated to hypertension such as age and established cvd offer a biologically more plausible explanation. in univariate analyses, the relative risk of mild rd and severe rd increased by a factor of - with age, male gender, bmi, hypertension, renal failure and cvd, in agreement with most earlier reports [ , , , [ ] [ ] [ ] ] . not surprisingly also markers of multiple organ failure, such as elevated creatinine and ast, or markers of hyperimmune response such as crp were positively correlated to worse outcome. indeed, crp was the single most powerful predictor of outcome, linearly increasing from no rd to mild rd to severe rd in a way that allowed the establishment of a threshold for severe rd, with a good tradeoff of % sensitivity and . % specificity. many of the individual predictors turned out to be intercorrelated and were excluded by logistic regression analysis, which retained only age, creatinine and crp as independent predictors of severe rd and death. creatinine levels, although systematically measured in earlier papers, have never been adequately emphasized as important markers [ ] . of note, in our cohort chronic renal failure was recorded in % ( / ) of patients on presentation, whereas the proportion of patients with actual creatinine levels above threshold was overall % with a distribution that proportionally increased from no rd to severe rd (table ) . reasons for this finding may depend on a selective vulnerability of kidney to coronavirus, possibly due to a large representation of ace , on vascular impairment (pre-existing or caused by vasculitis) or both [ , ] . an important missing data in the present study is d-dimer and troponin t, important markers of the coagulopathy and silent myocardial damage that are increasingly being recognized in covid- [ ] [ ] [ ] [ ] . this occurred partly for a reduced awareness of the problem at the beginning of the study and partly because our interest was focused on antihypertensive treatment. whether this flaw may have had an impact on the ascertainment of the cause of death is unknown. we did not record the certificate of death, and it is possible that a proportion of patients died of pulmonary embolism or of myocardial failure, but we believe this does not affect our results given the primary aim of the study. in conclusion, we studied prospectively a cohort of consecutive emergency department patients found to have covid- and were able to assess the relationship between acei and arb use and the severity of the disease. our findings rule out any effect of acei or arb on prognosis [ ] . although many clinical variables had an individual effect on outcome, age, creatinine and crp were the only independent predictors of severe rd and death. limitations of the study are the single centre nature, the limited time span of enrolment and the inability to collect important laboratory data. strengths are the prospective enrolment and the direct assessment "on field" of all included patients. clinical characteristics of coronavirus disease in china epidemiological determinants of spread of causal agent of severe acute respiratory syndrome in hong kong clinical course and risk factors for mortality of adult inpatients with covid- in wuhan, china: a retrospective cohort study characteristics of and important lessons from the coronavirus disease (covid- ) outbreak in china: summary of a report of cases from the chinese center for disease control and prevention risk factors associated with acute respiratory distress syndrome and death in patients with coronavirus disease pneumonia in wuhan, china renin-angiotensin system blockers and the covid- pandemic: at present there is no evidence to abandon renin-angiotensin system blockers antihypertensive treatment with acei/arb of patients with covid- complicated by hypertension renin-angiotensin-aldosterone system inhibitors in patients with covid- are patients with hypertension and diabetes mellitus at increased risk for covid- infection? antihypertensive drugs and risk of covid- radiological findings from patients with covid- pneumonia in wuhan, china: a descriptive study clinical features and treatment of covid- patients in northeast chongqing clinical characteristics of patients infected with sars-cov- in wuhan coronavirus disease in elderly patients: characteristics and prognostic factors based on -week follow-up baseline characteristics and outcomes of patients infected with sars-cov- admitted to icus of the lombardy region ace the janus-faced protein-from cardiovascular protection to severe acute respiratory syndrome-coronavirus and covid- covid- and the cardiovascular system antihypertensive drugs and risk of covid- ?-authors' reply renin-angiotensin system inhibitors improve the clinical outcomes of covid- patients with hypertension antihypertensive drugs and risk of covid- ? antihypertensive drugs and risk ofcovid- ? prevalence of underlying diseases in hospitalized patients with covid- : a systematic review and meta-analysis cardiovascular implications of fatal outcomes of patients with coronavirus disease (covid- ) the association between cardiac injury and outcomes in hospitalized patients with covid- abnormal coagulation parameters are associated with poor prognosis in patients with novel coronavirus pneumonia prevalence of asymptomatic deep vein thrombosis in patients hospitalized with sars-cov- pneumonia: a cross-sectional study antagonizing the renin-angiotensin-aldosterone system in the era of covid- key: cord- -a sz e c authors: suryadevara, v.; adusumalli, c.; adusumilli, p. k.; chalasani, s. h.; radhakrishnan, r. title: mental health status among the south indian pharmacy students during covid- pandemic quarantine period: a cross-sectional study date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: a sz e c introduction: the covid- outbreak created a major panic among all the citizens of the country owing to its severity, contagiousness within the community, lack of specific treatment and possibility of re-infection. all these factors along with the uncertain behaviour of the virus lead to state of fear and concern all throught out the nation. the current study represents the mental health survey conducted on the students of south india after the completion of one month quarantine period of the covid- outbreak. methodology: the present study is a cross-sectional, web-based online survey which consists of -item dass questionnaire. this was used to assess the emotional states of depression, anxiety, and stress. using google forms, the questionnaire was randomly distributed among the pharmacy students of selected colleges. mean with standard deviation was calculated for continuous variables and the number with percentage was calculated for categorical variables. results: a total of participants responded to the questionnaire. more than half of the responses were received from females ( %). on assessment it was found that, % of respondents reported severe to extremely severe depressive symptoms; . % of respondents reported severe to extremely severe anxiety symptoms, and % reported severe to extremely severe stress levels. conclusion: in india during the outbreak of covid- , an alarming number of students were found to have an impact on mental health due to the outbreak and were observed to have higher levels of stress, anxiety, and depression. the study findings shows the need of conducting more such studies and can be used to prepare appropriate psychological interventions to improve mental health among the young public during the pandemic. in december , a cluster of pneumonia cases of unknown cause was was reported to who which were identified in wuhan city, china. on th february , who declared the name of this disease as "covid ";later the virus was renamed by the international committee on taxonomy of viruses (ictv) as severe acute respiratory syndrome coronavirus (sars-cov ). even though the novel corona virus is genetically related to the sars-cov outbreak of but it is of different type in several aspects. [ ] the covid- outbreak created a major panic among all the citizens of the country owing to its severity, contagiousness within the community, lack of specific treatment and possibility of re-infection. all these factors along with the uncertain behaviour of the virus lead to state of fear and concern all throught out the nation. the complete dynamics of transmission are yet to be determined but the general transmission of respiratory viruses happens through droplets. [ , ] since the droplets travel approximately meterin air and quickly settles on the surfaces, person to person transmission possibly occurs between close contacts. it is advised to frequently clean the hands with soap or alcohol-based sanitizers and to follow stand hygiene measures because the contaminated hands is reported to carry the virus into the body. [ , ] the known symptoms range from mild (cough, shortness of breath, etc) to severe (pneumonia, kidney failures, and death). currently, there is no specific drug or vaccine,and the development of treatment and vaccines are under progress. [ ] since covid- is a newly identified coronavirus, studies are going on to obtain more information about this organism as most of the data related to it is unknown. hence all the above factors contribute to the agitation among the young generation in overall well being. . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) the copyright holder for this preprint this version posted may , . . currently, there is no much data on the mental health of the young public during the outbreak of covid- . this is especially relevant; uncertainty surrounding an outbreak of such a large extent. mainly the research studies during the covid- outbreak focuses on identifying the clinical features and epidemiology of infected patients [ ], the genomic characterization of the virus [ ] , and challenges for worldwide health governance [ ] . however, to our knowledge, published articles are examining the impact of covid- on the young public of india. due to the widespread of the coronavirus in the public, the government of india has declared days of lockdown in the first phase and days of lockdown in the second phase for the entire country. as of lockdown, the people have to stay at home and during the lockdown, all the schools and colleges have been locked. the lockdown means isolation and to maintain social distancing to stop the spread of the infection. sudden isolation and social distancing can significantly affect the mental health of people due to various reasons. this study represents probably the first mental health survey conducted in the students of south indiaafter the one month quarantine period of the covid- outbreak. this study aims to find the prevalence of psychiatric symptoms among students. the results obtained may supportacademic institutions and healthcare professionals in safeguarding the psychological wellbeing of the studentsduring covid- outbreak expansion in all parts of india. the present study is a cross-sectional, web-based online survey was conducted between rd april and th april (after one month period of lockdown). a -item dass questionnaire was used to assess the emotional states of depression, anxiety, and stress. it is a set of three self-report scales; each of the three dass- scales contains items, divided into subscales with similar content. a short form ( items) and a long-form ( items) which are valid and reliable measures inpatient and general population [ , ] different racial and cultural groups. this is a -item scale measured on a -point rating scale ( - ), " " denoting "applied to me very much or most of the time"and" "denoting"did not apply to me at all". all the questions were provided with multiple options with only one answer to be chosen. . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . using google forms, the questionnaire was randomly distributed among the pharmacy students of selected colleges. through emails and whatsapp, we requested people to circulate the survey link among their respective college mates. participants were asked to give consent before taking part in the survey. the study ensured the confidentiality of the participant's personal information they provided. the study does not deliver any intervention to participants; thus, there is no risk of physical harm to participating individuals. this study received an exemption from the institutional human ethics committee. mean with standard deviation was calculated for continuous variables and the number with percentage was calculated for categorical variables. a total of participants responded to the questionnaire. more than half of the responses were received from females ( %). the majority % of participants were from nuclear family and around % are from the urban area. the age of participants ranged from - years; with mean (sd) age of . (+/- . ) years. after the country's outbreak of covid- , the government of india declared public health emergency of national concern, % of respondents reported severe to extremely severe depressive symptoms; . % of respondents reported severe to extremely severe anxiety symptoms,and % reported severe to extremely severe stress levels. is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . scores for all the subsets of depression, anxiety, and stress are calculated by summing the scores obtained from the respondents for the relevant items. the dass- is based on a dimensional rather than a categorical conception of psychological disorder. the assumption on which the dass- development was based is that the differences between the depression, anxiety, and the stress experienced by normal subjects and clinical populations are essentially differences of degree which was confirmed by research data. this scale has no direct inferences for the distribution of patients to separate diagnostic categories proposed in classificatory systems such as the dsm and icd. students were also found to havean impact on the outbreak and higher levels of stress, anxiety, and depression. as the total number of people infected by covid- is rising in an alarming condition, major cities in south india have shut down all academic institutions at all levels indefinitely. the potential negative impact and uncertainity on academic development could hurt the mental health of students [ ] . as young people are more receptive to digital applications [ ] , healthcare providers could consider providing online or smartphone-based psychological interventions and psychoeducation such as cognitive behavior therapy; to reduce the risk of negative impacts associated with the epidemic. a support network could be provided through online platforms for those people who are in quarantine during the epidemic. health authorities need to provide needful information in a systematic format in simple languages (either audio or diagrammatic) to support the public with lower grades of educational background during the epidemic. . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint though life needs stress constructively up to a certain limit and may be adequate for personality development, but if these stresses become too severe which disengage the psychic equilibrium producing maladaptive patterns of behavior. inadequate interaction with the environment and family leads to stress and anxiety [ ] . the resources available are limited in this period and we have adopted the snowball sampling strategy for collecting the data. as a result, huge data was not collected; the conclusion was less generalizable to the entire population, particularly less diverse young public. one more limitation of the study is that self-reported levels of anxiety, depression, and stress may not always be aligned with the assessment done by trained health professionals. the study also did not assess the risk factors which might have contributed for alteration of the mental health status. this study provides important information about the respondents on the psychological responses of weeks after the outbreak of covid- . our study findings directly inform the need for the developing and implementing the strategies of several psychological interventions that may help minimizing anxiety, depression, and stress during the outbreak of covid- . as the covid- epidemic, which is still ongoing at the time of communicating this manuscript, all the interventions developed will be helpful in large extent to indian youth. in india during the outbreak of covid- , around % of the respondents reported extremely severe depression, and about . % of respondents reported extremely severe anxietyand about . % of respondents reported extremely severe stress. an alarming number of students were found to havean impact on mental health due to the outbreak and were observed to havehigher levels of stress, anxiety, and depression. the study findings can be used to prepare psychological interventions to improvemental health among the young public during the covid- epidemic. conflicts of interest:none to declare . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint team arrives in iran; covid- surges in korea, italy. center for infectious disease research and policy, - )-and-the-virus-that-causes-it medical journals and the -ncov outbreak genomic characterisation and epidemiology of novel coronavirus: implications for virus origins and receptor binding the psychological e_ects of quarantining a city the short-form version of the depression anxiety stress scales (dass- ): construct validity and normative data in a large non-clinical sample the utility of somatic items in the assessment of depression in patients with chronic pain: a comparison of the zung self-rating depression scale and the depression anxiety stress scales in chronic pain and clinical and community samples methodology of development and students' perceptions of a psychiatry educational smartphone application. technol. health care off receptiveness and preferences of health-related smartphone applications among vietnamese youth and young adults effect of family environment on behavioural problems and family dynamics the authors would like to thank all the study respondents for their active participation in providing the scientific data which helps in developing strategies/interventions for building a healthy nation. key: cord- -efqf e i authors: yamasaki, yukitaka; ooka, seido; tsuchida, tomoya; nakamura, yuta; hagiwara, yuta; naitou, yoshiyuki; ishibashi, yuki; ikeda, hiroki; sakurada, tsutomu; handa, hiroshi; nishine, hiroki; takita, mumon; morikawa, daiki; yoshida, hideki; fujii, shuichi; morisawa, kenichiro; takemura, hiromu; fujitani, shigeki; kunishima, hiroyuki title: the peripheral lymphocyte count as a predictor of severe covid- and the effect of treatment with ciclesonide date: - - journal: virus res doi: . /j.virusres. . sha: doc_id: cord_uid: efqf e i we investigated whether reduced lymphocyte count, could predict the development of severe covid- . we also examined whether ciclesonide could prevent the development of severe covid- among patients with the predictors. this was a retrospective cohort study. of the included patients, , , and were allocated to severe pneumonia, non-severe pneumonia, and non-pneumonia groups, respectively. the group of the low level of lymphocyte counts of the sixth day after onset was significantly intubated approximately three days later. the incidence of the severe pneumoniae requiring intubation are significantly lower in the patients treated with ciclesonide than without it ( . % vs . %, p = . ). the lymphocyte count after ciclesonide treatment in the non-severe pneumonia group was significantly higher (p = . ) than before. the lymphocyte count could be used to identify patients that may develop severe covid- . treatment with ciclesonide may prevent the development of severe covid- . since first appearing in wuhan, hubei province, china, in december , a novel coronavirus (sars-cov- ) has spread rapidly around the world. many patients with coronavirus infection disease (covid- ) are subclinical, and it has been reported that people are j o u r n a l p r e -p r o o f contagious even when asymptomatic [ , ] , which means preventing the spread of sars-cov- is challenging [ ] . in addition, some patients have been reported to deteriorate rapidly in the early stages [ ] . therefore, early detection and preventing cases from progressing to a severe stage is essential. about % of covid- cases progress to a severe stage, of which about % die. risk factors of severe pneumonia include age, comorbidities, smoking, reduced lymphocyte count, elevated ferritin levels, and elevated c-reactive protein (crp) levels [ ] [ ] [ ] [ ] [ ] [ ] . however, it is unclear which of these risk factors are predictors of progression to severe covid- . as yet, no effective treatment has been found for covid- . there have been many medications suggested, including remdesivir [ ] [ ] [ ] , lopinavir and ritonavir [ ] , and chloroquine [ ] , but their efficaciousness have yet to be verified. ciclesonide is an inhaled corticosteroid that is approved to treat asthma. it has demonstrated antiviral effects in vitro [ ] and has been reported to be effective in treating covid- [ ] . according to a report by meehyun koa et al., the infection inhibitory effect of ciclesonide was confirmed in the mers-cov strain isolated in south korea [ ] . furthermore, because ciclesonide is a local administration, there are few side effects, and administration is possible for a pregnant woman relatively safely. we believe that preventing the development of severe covid- will help to reduce the mortality rate. we investigated whether any of the factors that have been reported to correlate with severe pneumonia could predict the development of severe covid- . in addition, we examined whether ciclesonide could prevent the development of severe covid- among patients with these predictors. this was a retrospective cohort study. all the patients were hospitalized at our institution between february and april , , and had tested positive for sars-cov- using polymerase chain reaction testing of pharyngeal or nasopharyngeal swabs taken. for all patients, the date of onset was the day clinical symptoms appeared, such as fever, cough, runny nose, and dysgeusia. the presence of pneumonia was confirmed by chest computed tomography (ct). patients who underwent intubation and respiratory management were defined as severe pneumonia group. written informed consent for this study was obtained. the study was conducted with the approval of our hospital's institutional review board (approval number: ). thirteen patients with covid- , hospitalized between february and march , , before treatment with ciclesonide starts, were enrolled in this study. blood tests performed less than days from the date of onset and before intubation were examined. if multiple blood tests were performed during the evaluation period, the minimum and maximum values were examined. the leukocyte count, lymphocyte count, platelet count, crp, ferritin, d-dimer, and kl- were examined. patients were divided into three groups: severe pneumonia, non-severe pneumonia, and non-pneumonia. for the lymphocyte count, the mean+ sd was used as the cutoff value of severe covid- pneumonia. the cases at or below this cutoff value were evaluated, and patients who started ciclesonide after intubation were excluded. the treatment group received inhalations of µg ciclesonide once a day, for a daily total of µg. the relationship between ciclesonide use and severe pneumonia were examined. in addition, the lymphocyte count prior to and approximately j o u r n a l p r e -p r o o f days after starting treatment were compared. data were analyzed with the mann-whitney u , fisher's exact and wilcoxon matchedpairs signed rank tests using graphpad prism ver. . for windows, graphpad software, san diego california usa.. of the patients who were hospitalized during the observation period, was excluded due to a lack of data before intubation. of the included patients, were allocated to the severe pneumonia group, to the non-severe pneumonia group, and to the non-pneumonia group. the study design of this study was shown in figure . baseline characteristics table details the patients' demographic information. the mean age was . years, and . % were male. of the total and those with pneumonia, . % and . % had j o u r n a l p r e -p r o o f comorbidities, respectively. blood tests were on average performed . days after onset (sd . ) and days after treatment (sd . ). on average, patients developed severe covid- and underwent intubation and respiratory management days after onset (sd . ). of the patients with covid- hospitalized between february and march , , before the start of ciclesonide therapy, there were in the severe pneumonia group, in the non-severe pneumonia group, and in the non-pneumonia group ( figure ). lymphocyte counts of approximately sixth days after onset were significantly lower in the severe pneumonia group compared to both the non-severe pneumonia group and the nonpneumonia group (p = . , . , respectively) ( figure a) . the severe pneumonia group had a low mean lymphocyte count at cells/mm (sd . ). patients in the severe pneumonia group were significantly older than those in the non-severe pneumonia group (p= . ), but not significantly different from those in the non-pneumonia group (figure b ). significant differences were not observed between the severe and non-severe pneumonia groups in relation to ferritin, crp, and d-dimer. regarding sex differences, there tended to be more males in the severe and non-severe pneumonia groups. however, there was no significant difference in sex for the pneumonia cases. while . % of patients in the pneumonia groups had a comorbidity, the difference between the severe and non-severe pneumonia groups was not significant. (table ) , patients had severe pneumonia, and had non-severe pneumonia. eleven patients from with a lymphocyte count at or below the cutoff value could be treated with ciclesonide. of these, had severe covid- pneumonia, and the incidence of the severe pneumoniae requiring intubation are significantly lower in the patients treated with ciclesonide than without it ( . % vs . %, p= . ). thus ciclesonide therapy is suspected to exhibit a significant correlation with the non-severe pneumonia group. moreover, the lymphocyte count after ciclesonide therapy in the non-severe pneumonia group was significantly higher (p= . ) compared to before treatment (mean . days, sd . ) (figure b ). five patients with pneumonia were subsequently transferred to other hospitals, so their lymphocyte counts after treatment are unknown. there is currently no therapy that has been proven to be efficacious in treating covid- . as many covid- cases are subclinical, it is challenging to track infected individuals, making it hard to prevent infections from occurring [ ] [ ] [ ] . however, some patients with covid-in the present study, patients who received ciclesonide developed severe pneumonia. patients with covid- are known to deteriorate rapidly. both of these cases began treatment with ciclesonide days before intubation, which suggests that the drug may have been introduced too late. previous research has found that age, comorbidities, lymphocyte count, ferritin, crp, and d-dimer are associated with severe pneumonia. of these, only the preintubation lymphocyte count appeared to be a possible predictor with counts in the nonpneumonia and non-severe pneumonia groups being significantly different from the severe pneumonia group. regarding age, there was a significant difference between the severe and nonsevere pneumonia groups, but not with the non-pneumonia group. the year old case in the non-pneumonia group accounts for the lack of significant difference. we believe that this case did not cause pneumonia because there is no underlying disease and there are no risk factors other than age. significant differences were not observed for ferritin or d-dimer. there were cases with high ferritin and d-dimer levels in both the severe and non-severe pneumonia groups. in addition, there was a lot of missing data, making it difficult to accurately assess whether ferritin or d-dimer could be a predictor of severe covid- . while the presence of subclinical covid- cases makes controlling infections difficult, death occurs suddenly in some cases [ ] . covid- is known to be contagious days before onset, with increased viral loads in the respiratory tract. it has been suggested that cytokine storms are associated with the development of severe covid- . therefore, the early administration of antivirals could be efficacious, similar to influenza. it is important to identify patients with covid- as soon as possible and prevent it from progressing to a severe stage. the lymphocyte count could be used as an indicator for identifying patients that may develop severe covid- . our results suggest that treatment with ciclesonide may prevent the j o u r n a l p r e -p r o o f development of severe covid- in these circumstances. it is best to introduce the drug as soon as possible in patients with reduced lymphocyte counts and other predictors of severe covid- [ ] . this study had several limitations. this was a retrospective study with a small sample size. therefore, the results need to be confirmed in a larger, prospective study. the viral load determination could not be mentioned because it was not measured in all cases. we showed treatment with ciclesonide as the candidate of the factor which inhibited severe covid- in this study. we urgently need to establish a testing system that includes the antigen-antibody method, develop a vaccine, and find treatments that can prevent the development of and treat severe covid- . intubation, n(%) ( . ) a period to intubation, mean(sd), days . ( . ) n: number, sd: standard deviation asymptomatic and presymptomatic infectors: hidden sources of covid- disease presymptomatic transmission of sars-cov- -singapore the reproductive number of covid- is higher compared to sars coronavirus intensive care patients with covid- on the alert for cytokine storm: immunopathology in covid- . arthritis rheumatol hematologic, biochemical and immune biomarker abnormalities associated with severe illness and mortality in coronavirus disease (covid- ): a meta-analysis neutrophil-to-lymphocyte ratio and lymphocyte-to-c-reactive protein ratio in patients with severe coronavirus disease (covid- ): a meta-analysis neutrophil-to-lymphocyte ratio as an independent risk factor for mortality in hospitalized patients with covid- correlation between relative nasopharyngeal virus rna load and lymphocyte count disease severity in patients with covid- coronavirus susceptibility to the antiviral remdesivir (gs- ) is mediated by the viral polymerase and the proofreading exoribonuclease. mbio arguments in favour of remdesivir for treating sars-cov- infections broadspectrum antiviral gs- inhibits both epidemic and zoonotic coronaviruses a trial of lopinavir-ritonavir in adults hospitalized with severe covid- a systematic review on the efficacy and safety of chloroquine for the treatment of covid- the inhaled corticosteroid ciclesonide blocks coronavirus rna replication by targeting viral nsp therapeutic potential of ciclesonide inahalation for covid- pneumonia: report of three cases screening of fda-approved drugs using a mers-cov clinical isolate from south korea identifies potential therapeutic options for a mouse model for mers coronavirus-induced acute respiratory distress syndrome we thank all medical staff who treated covid with us.the authors have no conflicts of interest directly relevant to the content of this article.j o u r n a l p r e -p r o o f key: cord- -ug a wx authors: de pascale, gennaro; cutuli, salvatore lucio; pennisi, mariano alberto; antonelli, massimo title: the role of mannose-binding lectin in severe sepsis and septic shock date: - - journal: mediators inflamm doi: . / / sha: doc_id: cord_uid: ug a wx severe sepsis and septic shock are a primary cause of death in patients in intensive care unit (icu). investigations upon genetic susceptibility profile to systemic complications during severe infections are a field of increasing scientific interest. particularly when adaptive immune system is compromised or immature, innate immunity plays a key role in the immediate defense against invasive pathogens. mannose-binding lectin (mbl) is a serum protein that recognizes a wide range of pathogenic microorganisms and activates complement cascade via the antibody-independent pathway. more than % of humans harbor mutations in mbl gene (mbl ) resulting in reduced plasmatic levels and activity. increased risk of infection acquisition has been largely documented in mbl-deficient patients, but the real impact of this form of innate immunosuppression upon clinical outcome is not clear. in critically ill patients higher incidence and worse prognosis of severe sepsis/septic shock appear to be associated with low-producers haplotypes. however an excess of mbl activation might be also harmful due to the possibility of an unbalanced proinflammatory response and an additional host injury. strategies of replacement therapies in critically ill patients with severe infections are under investigation but still far to be applied in clinical practice. despite the diffusion of effective care bundles and the implementation of new technologies able to support organ function, severe sepsis and septic shock still represent a leading cause of intensive care unit (icu) admission with a case fatality rate of - % [ , ] . systemic inflammation, surrounding multiorgan failure and septic shock, results from a maladaptive unbalance between early antimicrobial immune reactions and uncontrolled local infection and inflammation. innate immune system is the primitive first-line organism's response to invasive pathogens, and it interacts with other homeostatic patterns, including inflammation and coagulation. early activation of immune response is mediated by soluble pattern recognition molecules that, in addition to complement proteins, cytokines, and coagulation factors, activate humoral and cellular effectors, identifying and neutralizing the invasive pathogen [ ] . mannose-binding lectin (mbl) is a soluble pattern recognition molecule which activates the lectin pathway of the complement system and the subsequent inflammatory mechanisms [ , ] . low mbl plasmatic levels, mainly due to genetic influences, have been largely described to be associated with susceptibility to invasive infections and poor outcome [ ] . on the other hand, the excessive activation of this ancient protective system may be responsible for a detrimental unbalanced inflammatory and coagulation response, as observed in inflammatory diseases, transplant rejections, and diabetic nephropathy [ ] . many authors have investigated whether mbl may influence the susceptibility to common pathogens and the development of severe infections, but, still, there is no consensus about the clinical relevance of its deficiency or the indications for replacement therapies. the purpose of this review is to summarize the results of relevant recent studies where the role of mbl in severe sepsis and septic shock has been investigated. mannose-binding lectin is a serum calcium-dependent protein, synthesized by the liver and is detectable in the sites of inflammation, particularly in epithelial-lining fluid [ ] . small amounts of this protein are also produced in other organs (kidney, thymus, tonsil, small intestine, and vagina). mbl is a collectin (collagen-like lectin) and is characterized by a highordered oligomeric structure that is essential for its function and interaction with mbl-associated serine proteases (masps) [ , ] . mbl harbors a carbohydrate recognition domain (crd) through which it binds to specific carbohydrates (i.e., mannose or n-acetylglucosamine) exposed on pathogenic agents surface, and it is therefore called a "patternrecognition molecule" [ ] . subsequently masps (mainly masp- ) are able to trigger the lectin complement pathway cleaving c and c to form c convertase. complement system may be activated by three pathways: the classic and the alternative ones are antibody dependent and belong to the adaptive immune response; the lectin one is antibody independent and, as part of the innate immune system, comes into play within the first hours from microorganisms' contact [ ] (figure ). mbl plays a central role as a firstline defense against invading pathogens by triggering complement system, directly mediating opsonophagocytosis, and possibly functioning as a toll-like receptor coreceptor [ ] . in humans there are two genes that might code for mbl, but only mbl gene is functional, located on the long arm of chromosome [ ] . mbl deficiency may be due to the presence of single nucleotide polymorphisms (snp) either in the gene-coding or in the promoter regions. the wildtype gene is called "a" (homozygous haplotype, a/a); instead the variants alleles are widely classified as " " ( /a and / ). three points mutations involve the exon , identifying allele b (codon ), c (codon ), and d (codon ). additionally, many snps may affect the promoter region and determine low mbl protein serum levels and activity (variants h/l, y/x, and p/q). even though all these mutations could be combined with exon alleles, seven haplotypes are typically found in humans genome [ , ] . these polymorphisms determine the production of unstable proteins, with shorter half-life, mainly due to the absence of the high-ordered oligomeric structure. these variants are not able to efficiently activate the complement pathway [ ] . healthy individuals (genotype a/a) generally present mbl levels above ng/ml. in the newborn this protein is detectable at concentrations of two-thirds of their mothers. normal levels are reached within a month [ ] . mbl levels are not influenced by age, circadian cycle, and physical exercise and, during inflammation, do not increase over - folds than baseline level [ ] . mbl deficiency is generally defined by plasmatic protein levels below ng/ml or by an mbl function lower than . u/ l c deposition [ ] . the detection of pulmonary mbl concentration is quite difficult. some authors have reported bronchoalveolar lavage (bal) levels ranging between and ng/ml, but these results were not corrected for dilution factors (i.e., urea or other lung proteins with known lung concentration) that could explain the large distance from the minimum concentration needed to activate complement proteins ( - ng/ml) [ ] . plasmatic levels ranging between and ng/ml are generally detected in heterozygous patients (a/o genotype); instead homozygous variant mbl alleles usually present very low concentrations (< ng/ml). similarly the haplotypes that include mutations in promoter regions are associated with significant reduction of mbl protein production and activity. however, even though the degree of mbl deficiency is strictly dependent upon patients' genotypes, in some cases low mbl plasmatic levels have been also associated with wild-type genes [ ] . this gene was already present in early invertebrates more than fifty million years ago and has been highly conserved throughout animal and human evolution. this would suggest that the correct function of mbl protein is crucial for the survival of living animal species. it is of interest to note that there is geographic distribution of different alleles: the b variant is predominant in eurasian populations, the c variant mainly among asians and america indians, and the d haplotype seems to be frequently expressed in caucasian region. this particular distribution might be linked to the initial human migrations table : mbl deficiency and susceptibility to diseases (human and animal studies). (vi) hsv out of africa and induced by some specific advantages due to mbl deficiency. for example, high mbl production has been observed to be associated with higher incidence of preterm births; instead moderately low levels could protect the organism from mycobacteria systemic infection and from the complement induced inflammatory-mediated damage of some diseases (i.e., meningococcemia and rheumatoid arthritis) [ , ] . additionally sporadic reports have not found a clear association between mbl deficiency and increased rate of infectious episodes [ ] [ ] [ ] . the wide range of clinical effects linked to mbl haplotypes has also been attributed to the role of associated mutations in other genes encoding proteins with similar functions (i.e., l-ficolin, masp , and surfactant proteins) [ , ] . however, to date, there are few data upon the clinical role of these combined deficiencies. the prevalence of mutations in one or both mbl gene alleles is relevant, ranging between % and % in analyzed populations [ , ] . during the last twenty years an increasing body of evidence has indicated that mbl deficit, due to specific haplotypes, generally increases frequency and severity of infectious episodes [ , ] (table ) . however the structure of our immune system is redundant, and this may explain why in many cases polymorphisms of mbl gene were not observed to influence susceptibility to infections [ ] . the role of this lectin is particularly relevant when adaptive immune system is immature or compromised [ ] . in a case-control study upon infants, lower mbl cord blood concentrations were associated with a higher incidence of gram-negative sepsis ( = . ) [ ] , and an observational cohort study upon pediatric icu patients identified mbl gene exon polymorphisms as a main determinant of progression from sepsis to septic shock [ ] . additionally, the incidence and outcome of severe infections appear to be influenced by the levels and activity of mannosebinding lectin. in a cohort of leukemic patients undergoing chemotherapy, severe infections (bacteremia, pneumonia or both) occurred more frequently in those individuals with lower mbl concentrations ( < . ) [ ] . in an ethnically homogeneous english population, homozygotes for mbl codon variant alleles showed a significantly higher risk of invasive infections due to streptococcus pneumoniae, "the captain of men of death" [ ] . similarly allelic variants of this gene seem to be associated with increased susceptibility to meningococcal disease [ ] . among respiratory tract infections, independently from the causal pathogen, mbl insufficiency has been observed to predispose to higher severity and poor outcome [ ] . even though legionella spp. act as an intracellular pathogen, mbl function was lower in infected cases during an australian legionnaires' disease outbreak [ ] . increased susceptibility and worse outcome in caucasian patients with acute respiratory distress syndrome (ards) were also observed in presence of mbl gene polymorphisms [ ] . regarding viruses, in chinese population, the presence of mbl gene b variant was associated with increased risk of coronavirus infection [ , ] ; instead normal mbl function seems to worsen pandemic h n and avian h n infections by potentially upregulating inflammatory response [ ] . furthermore, in a recent large retrospective study involving donor-recipient orthotopic liver transplantation pairs, patients who received mbl-deficient livers showed a threefold increased risk of clinically significant infections including cytomegalovirus-related diseases [ ] . few authors have studied the role of mbl in severe fungal infections. polymorphisms of this gene were observed in seven of ten white patients with chronic necrotizing aspergillosis compared with % of controls [ ] . in addition variations of mbl plasmatic levels seem to correlate with the occurrence of invasive candidiasis [ ] . mbl genetic, plasmatic, and functional profiles were investigated in numerous clinical settings obtaining different results. the critically ill patient, affected by severe infections with severe sepsis and septic shock, might be a field of particular interest for a better knowledge of their clinical relevance and the possible development of novel therapeutic strategies. mannose-binding lectin is not only part of the innate recognition system of invasive pathogens but effectively modulates the cytokines' production by macrophages during phagocytosis. this effect, upon interleukin (il)- , il- , and tumor necrosis factor-, was clearly shown in an "ex vivo" model of immediate immunity response to neisseria meningitidis infection [ ] . mbl deficiency may be associated with unbalanced proinflammatory responses to infective and noninfective triggers. in a cohort of critically ill pediatric patients, fidler and coworkers observed that mbl levels less than ng/ml, consistent with mbl- gene exon polymorphisms, significantly increased the risk of developing systemic inflammatory response syndrome (sirs) and progression to severe sepsis/septic shock [ ] . additionally in patients with sirs, mbl insufficiency degree was observed to correlate with severity of systemic infection, according to the genetic profile [ ] . the association between the deficiency of this protein and worse outcome during severe systemic infections (i.e., evolution to refractory septic shock) may be also related to the significant interaction between complement activation, inflammatory cytokines' "storm", and coagulation cascade. the influence of complement activation upon septic shock development was largely investigated. many studies have shown how the classical and alternative pathways are activated during septic shock and are involved in mechanisms aimed to clear endotoxin. this role has been more recently studied also for lectin complement activation due to mbl [ , ] . disseminated intravascular coagulation (dic) may worsen the course of septic shock but the occurrence of this severe complication is unpredictable. however recent data suggest that mbl deficit may be a significant risk factor for the early development of dic and organ failure during severe infections [ ] . conversely, excessive mbl expression might be harmful, since this molecule may contribute to the pathogenesis of inflammatory induced vascular damage and organ failure, as observed in patients undergoing solid organ transplantation [ ] . hence mbl, due to its pivotal role in the crosstalking among complement activation, coagulation, and systemic inflammation, may represent a key point for the understanding of the development of systemic severe infections, as interestingly investigated in animal models and clinical studies involving patients with severe sepsis/septic shock. even though many differences between animal models and humans limit the "translationalability" of preclinical data, several mouse experiments support the role of mbl deficiency in severe infections, especially after bacteria inoculation. two functional mbl genes exist in the mouse, and the generation of double knockout gene-deficient mice has increased the investigations in this field. after inoculation of × cfu staphylococcus aureus, mbl-null mice showed at hours % mortality compared with wild-type (wt) mice which survived in a percentage of %. additionally, pretreatment of mbl-null mice with rhmbl increased their survival rate of about % [ ] . in another model of mbl and/or masp / deficient mice takahashi and coworkers observed that this deficiency was associated with early occurrence of dic and liver injury after s. aureus inoculation, suggesting the role of this protein in the development of organ failure and systemic coagulation activation during severe infections [ ] . another study demonstrated that mbl is able to strongly bind to o-antigen region of lps, contributing to mice platelets activation and rapid occurrence of septic shock [ ] . susceptibility of mbl null mice to pseudomonas aeruginosa postburn infection was also investigated [ ] . all mbl-null mice, after burn and bacterial inoculation, early developed septic shock and died; instead the majority of wt animals (two-thirds) survived. these observations underline the relevance of innate immunity and mannose-binding lectin in the susceptibility and outcome of severe bacterial infections occurring in this population. regarding fungal diseases, the protective role of this lectin was also observed in murine models of invasive pulmonary aspergillosis after ex vivo mbl administration [ ] . however lectin pathway activation does not only depend on mbl function. some authors have observed how deficient mice models, without the capability to activate mblindependent lectin cascade (i.e., ficolins and other collectins), are more susceptible to develop severe systemic pneumococcal infections [ ] . although most of literature evidence obtained by animal studies supports the importance of mbl in the acquisition and outcome of severe infections, these observations, due to unresolved several limits of animal studies, may not be considered conclusive and strongly need clinical human studies to definitely identify its clinical relevance. studies. the mbl key role as part of innate immunity is the reason why haplotypes associated with its deficiency mainly influence infectious episodes involving neonates and children or immunosuppressed adults. in a population-based prospective study performed in greenland upon almost eskimo children, both heterozygous and homozygous subjects, aged to months, for variant alleles presented a twofold increased risk of acute respiratory infections, including pneumonia [ ] . additionally, capoluongo and colleagues, analyzing preterm newborns, identified two mbl gene variants as independent risk factors associated with unfavorable outcome, including higher bronchopulmonary dysplasia prevalence [ ] . turkish authors have investigated the possible relationship between cord blood mbl levels and neonatal sepsis. the results indicated that lower mbl levels during fetal inflammatory response syndrome (firs) were associated with higher risk of sepsis development independently from gestational age and birth weight [ ] . another prospective study conducted on neonates ( of them were preterm) showed how lowest mbl levels were detected in infants with septic shock, especially in case of fatal outcome ( < . ). relevant sensitivity, specificity, positive, and negative predictive values for detecting sepsis episodes were also documented [ ] . in a recent swiss investigation, mbl levels were detected in cord blood of newborns. forty-seven developed sepsis ( % within the first hours of life) and % required catecholamines because of septic shock. after excluding those infants who underwent surgery, low mbl concentrations resulted independently being associated with increased risk of early-onset gram-negative sepsis [ ] . in pediatric oncological patients, mbl deficiency was associated with susceptibility, poor outcome, and duration of febrile neutropenic episodes [ ] . in a prospective study mbl deficit was observed to increase the severity of disease during pediatric icu admission after febrile neutropenia [ ] . additionally also mbl-related proteins deficit was mediators of inflammation investigated in this setting. in a cohort of children treated with chemotherapy for cancer, masp- deficit (< ng/ml) significantly increased the risk of febrile neutropenia and bacteraemia development and prolonged cumulative duration of hospitalization and antimicrobial treatment [ ] . the importance of mbl function during the first months of life, when the efficacy of innate immunity is crucial, has induced some authors to propose its dosing as part of a biomarkers panel for the early detection of severe neonatal infections in low-resource settings [ ] . impaired innate immune mechanisms may also increase the risk of nosocomial infections in critically ill patients as observed by sutherland and colleagues. in a genetic association study, the authors identified the relationship between snp in cd , mbl and toll-like receptor- with increased prevalence of positive cultures and sepsis [ ] . in a cohort of adult septic patients, mbl deficiency resulted also independently being associated with higher sequential organ failure assessment (sofa) score at day , suggesting its role as a risk factor for the development of severe sepsis and septic shock [ ] . additionally in a multicenter prospective study involving eight adults icus in u. k., the association between mbl- exon and promoter polymorphisms with the outcome of patients affected by severe sepsis and septic shock was studied [ ] . compared with healthy subjects, mbl deficient patients were at increased risk of sepsis, with a significant higher mortality rate in presence of levels below ng/ml ( . % versus . %, = . ). during severe sepsis and septic shock, the increase of mbl plasmatic levels, as acute phase response molecule, may be different. in a report of adult critically ill patients, dean and colleagues observed that regardless of mbl- genotype those patients who were mbl deficient at study entry were not able to reach normal plasmatic levels during severe sepsis and septic shock [ ] . furthermore, a well-conducted prospective study, performed in denmark, investigated the mbl genetic and plasmatic profile in a population of critically ill icu patients with documented sirs [ ] . among enrolled patients met the criteria for severe sepsis and for septic shock. compared with noninfectious sirs, these patients shared the carriage of mbl variants alleles and low serum levels according to the severity of disease ( = . ). another recent korean study in icu patients investigated whether mbl gene polymorphisms and serum levels might influence severity and prognosis of sepsis [ ] . the authors compared septic patients with healthy controls, analyzing three snp and dosing mbl serum levels on day one. among sepsis group, homozygosis for the polymorphism at codon (a/a) resulted in a significant risk factor for severe sepsis development ( = . ). mbl serum levels ≥ . mcg/ml were associated with a lower day mortality rate in the septic shock group ( = . ). the role of mbl deficiency in critically ill patients with severe pneumonia, a still leading cause of death due to an infectious disease, has been investigated by many authors. in a large case-control study, patients affected by community-acquired pneumonia (cap) were compared with healthy control subjects and patients without relevant infectious diseases. mbl and masp haplotypes were equally distributed among those subjects. in the multivariate analysis, mbl deficiency was associated with poor outcome measures (i.e., severe sepsis, acute respiratory failure, multiorgan dysfunction syndrome, and death) [ ] . eisen and colleagues have reanalyzed data from six studies involving patients affected by severe infections [ ] . first, the authors defined a mbl cutoff value of . mcg/ml as a reliable predictor of low producing status (negative predictive value %). they confirmed that mbl deficiency significantly increased the risk of death due to severe infection, also in icu setting, especially when streptococcus pneumoniae was the invasive causative agent (odds ratio . , % confidence interval, . - . ) . the association between mbl deficiency and s. pneumoniae invasive infection outcome has been recently investigated in a spanish prospective cohort study [ ] . during the study period patients with invasive pneumococcal infection were enrolled: the rate of allelic variants was %. snp mbl (ao/oo) and septic shock were the factors independently associated with in-hospital mortality. otherwise early adequate antibiotic dose ≤ hours resulted in a significant protective determinant. mbl deficiency role was also studied in some other systemic infections due to specific organisms. resman and colleagues recently described the case of a necrotizing myositis and septic shock due to haemophilus influenzae in a patient where igg and mbl deficiency were diagnosed [ ] . in a well-conducted prospective study, the correlation between mbl gene polymorphisms and the outcome of escherichia coli pyelonephritis was investigated [ ] . although no association was found with the incidence of e. coli infections and the presence of bacteremia, those patients who shared lowexpression mbl genotypes showed a significant higher risk of septic shock development (odd ratio: . , % confidence interval: . - . ; = . ). finally, in nonbacterial severe systemic infections, invasive candidiasis (ic), especially candidemia, still remains a leading cause of death due to infections in critically ill patients. serum mbl levels were measured in patients with proven ic, hospitalized not infected patients, and healthy subjects [ ] . even though mbl concentration was significantly higher in ic patients than controls, the authors identified a marked decrease in its plasmatic levels during the first days of infection in association with mannans increase. these observations, although limited, suggest a crucial role of mbl also in the early phase of candidiasis. therapy. mbl substitution therapy in patients with recognized lectin deficit has been proposed. apart from genetic analyses, antigenic measurement is widely diffused as diagnostic test. even though mbl serum levels < ng/ml or mbl activity < u/ml may be considered a significant deficiency, there are not standard guidelines aimed to define which patient categories need to be tested (i.e., in presence of severe recurrent respiratory infections or acquired immunesuppression). recombinant human mbl use, to supplement mbl deficiency status, has been investigated in animal and phase i/ii human studies [ , ] . although its clinical efficacy has not been clearly established, still now no adverse effects were observed. sixty-five mbl infusions were given to mbl deficient chemotherapyinduced neutropenic children. the observed postadministration level was . mcg/ml (range: . - . ) which may be considered protective [ ] . a similar pharmacokinetic profile was observed in healthy mbl-deficient volunteers and two patients with staphylococcus aureus septicemia [ ] . however, beyond these preliminary observations, mbl replacement needs to be further investigated in deficient patients affected by acute severe infections, especially in presence of multiple-level immune system impairment. an increasing body of data support the role of mbl as central player of innate immunity. several gene polymorphisms have been identified in association with decreased serum levels and activity. many authors have showed the association of this molecule deficit with recurrent severe infections, particularly involving the respiratory tract and encapsulated bacteria. additionally growing evidence suggests its importance during systemic severe infections as severe sepsis and septic shock. this correlation might derive from the crosstalking among complement system, coagulation patterns, and proinflammatory cytokines. even though many patients with systemic infections, who present mbl serum levels below the functional threshold, are at higher risk to develop severe complications and poor outcomes (i.e., septic shock, multiple organ failure), in some cases low levels have appeared to be protective, probably reducing the inflammatory cytokines' storm. moreover not all published studies have identified a clear association between deficiency and increased risk of infections. replacement therapy with recombinant human protein during severe sepsis and septic shock affecting deficient patients has been proposed but it still remains an experimental treatment. hence, until new promising and robust data will be available, the strict adherence to current standard recommendations still remains the mainstay of severe 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neutropenia in pediatric cancer patients emerging biomarkers for the diagnosis of severe neonatal infections applicable to low resource settings polymorphisms in cd , mannose-binding lectin, and toll-like receptor- are associated with increased prevalence of infection in critically ill adults low mannosebinding lectin function is associated with sepsis in adult patients mannose-binding lectin polymorphisms in severe sepsis: relationship to levels, incidence, and outcome association of mannose-binding lectin- genotype and serum levels with prognosis of sepsis genetic variants of the mbl gene are associated with mortality in pneumococcal sepsis necrotizing myositis and septic shock caused by haemophilus influenzae type f in a previously healthy man diagnosed with an igg and a mannosebinding lectin deficiency association between mannose-binding lectin deficiency and septic shock following acute pyelonephritis due to escherichia coli human plasma-derived mannose-binding lectin: a phase i safety and pharmacokinetic study infusion of plasma-derived mannan-binding lectin (mbl) into mbl-deficient humans safety and pharmacokinetics of plasma-derived mannosebinding lectin (mbl) substitution in children with chemotherapy-induced neutropaenia the pharmacokinetic profile of plasma-derived mannan-binding lectin in healthy adult volunteers and patients with staphylococcus aureus septicaemia the authors declare that there is no conflict of interests. key: cord- - oy qqy authors: bai, xiang; fang, cong; zhou, yu; bai, song; liu, zaiyi; chen, qianlan; xu, yongchao; xia, tian; gong, shi; xie, xudong; song, dejia; du, ronghui; zhou, chunhua; chen, chengyang; nie, dianer; tu, dandan; zhang, changzheng; liu, xiaowu; qin, lixin; chen, weiwei title: predicting covid- malignant progression with ai techniques date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: oy qqy background: the coronavirus disease (covid- ) has become a worldwide pandemic since mid-december , which greatly challenge public medical systems. with limited medical resources, it is a natural strategy, while adopted, to access the severity of patients then determine the treatment priority. however, our work observes the fact that the condition of many mild outpatients quickly worsens in a short time, i.e. deteriorate into severe/critical cases. hence, it has been crucial to early identify those cases and give timely treatment for optimizing treatment strategy and reducing mortality. this study aims to establish an ai model to predict mild patients with potential malignant progression. methods: a total of consecutively mild covid- patients at admission who was hospitalized in wuhan pulmonary hospital from january to february , , were selected in this retrospective irb-approved study. all mild patients at admission were categorized into groups with or without malignant progression. the clinical and laboratory data at admission, the first ct, and the follow-up ct at severe/critical stage of the two groups were compared with chi-square test, fisher's exact test, and t test. both traditional logistic regression and deep learning-based methods were used to build the prediction models. the area under roc curve (auc) was used to evaluate the models. results: the deep learning-based method significantly outperformed logistic regression (auc . vs. . ). the deep learning-based method achieved a prediction auc of . by combining the clinical data and the ct data, significantly outperforming its counterpart trained with clinical data only by . . by further considering the temporal information of the ct sequence, our model achieved the best auc of . . the proposed model can be effectively used for finding out the mild patients who are easy to deteriorate into severe/critical cases, so that such patients get timely treatments while alleviating the limitations of medical resources. in mid-december , the ongoing coronavirus disease broke out in wuhan and spread rapidly in the mainland of china ( cases, updated through march , ) . so far, the infection had burst in countries outside china, evolving into a pandemic , . according to the chinese epidemic data, the mild, severe, and critical types of covid- were %, %, and % separately . more seriously, as of march, the mortality of covid- was . % ( / ) in the mainland of china, even reached . % ( / ) in wuhan city, which was much higher than that of other influenza , . in addition, the clinical course of covid- varied individually. in order to prevent malignant progression and reduce the mortality of covid- , it is vital to identify mild patients who are easy to deteriorate into severe/critical cases and give them active treatment earlier. however, most studies focused on cross-sectional description and comparison of clinical, laboratory and ct imaging findings [ ] [ ] [ ] [ ] . some studies focused on seeking risk factors for death outcome , . none of them used ai-based methods for progression prediction of mild covid- patients up to date. to solve this problem, we aimed to apply ai techniques to study multivariate heterogeneous data (clinical data and serial chest ct imaging) and to further develop an accurate and effective prediction model. specifically, we employed a deep learning-based model to effectively mine the complementary information in static clinical data and serial quantitative chest ct sequence. since deep learning-based methods had been widely adopted and had achieved great performance in cancer outcome prediction , head ct scans detection and antibiotic discovery investigation. the inclusion criteria were: ) respiratory rate < breaths per min; ) resting blood oxygen saturation > %; ) the ratio of arterial oxygen partial pressure to fraction of inspiration oxygen > mmhg; ) non-icu patients without shock, respiratory failure, mechanical ventilation, and failure of other organs. the clinical and laboratory data at the time point of admission, together with serial chest ct images of all patients were retrospectively analyzed. based on the presence or absence of the severe/critical progression during the hospitalization, all patients were categorized into two groups. the diagnostic criteria for severe/critical progression were: ) respiratory rate radiology) independently blinded to the clinical information, and the discrepancy was resolved by consulting another radiologist (wc, years' experience in radiology). lesions and imaging features were assessed in each lung segment of each patient. the number of involved segments was counted not only for each patient or each lobe but also for each imaging feature. if more than one type of imaging features present in a segment, the segment was counted for every involved feature. the imaging features assessed in this study included ) ground glass opacity (ggo); ) consolidation; ) air bronchogram; ) paving stone sign; ) fibrosis; ) nodule; and ) halo sign. the first available ct after symptoms onset, the follow-up ct, the first available ct of the severe stage were assigned as ct , ct ~ ct n , and ct severe separately. in order to compare the longitudinal variation of ct features during the period of ct and ct severe between the two patient groups, we chose ct instead of ct severe for those patients without severe/critical progression. our raw covid- dataset contained all the clinical data and the quantitative chest ct data. after excluding invalid and duplicate information, each sample contained clinical data characteristics and a quantitative ct sequence obtained at different times. since the sequence length of each sample varied from zero to seven, we adjusted the data structure of each sample to the same shape by zero-filling the uncollected or missing chest ct data. the original quantitative chest ct data contained twelve infection distribution features, eight infection sign type features, the thickness of thoracic diaphragm, and ct course. the lung was medically divided into segments, and the infection sign characteristics at each checkpoint can be formatted as a matrix. this matrix composed of infection distribution features and sign type features was flattened into a vector and then concatenated with the original quantitative chest ct data. the pipeline of the prediction model is shown in figure . the input data includes the static data and the dynamic data, where the static data is a -dimensional vector, containing the clinical data and personal information of patients. dynamic data is a series of quantitative chest ct data collected at different times. each ct data at different checkpoint consists of a × matrix and a -dimensional vector. in order to merge these two parts, we directly flattened the all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint matrix into an -dimensional vector and concatenated it with the -dimensional vector to form a -dimensional ct feature vector. according to the checkpoints, the ct data sequence with a length of seven and a dimension of was formed. for the sake of combining static and dynamic data as the input of long short term memory (lstm), a multi-layer perceptron (mlp) was applied to the static data to obtain a -dimensional feature vector, which is used as the input data of the first timestamp of the lstm, followed by the other seven ct feature vectors . the lstm model employed in this study is a single-layer network with the embedding dimension of and the hidden dimension of . the output of the lstm, a × feature sequence, was then fed into fully connected layers. a softmax layer was added at the top of the network to output the probability of the patient conversion to the severe/critical stage. a total of samples were included in the covid- dataset. the robustness of the model was evaluated by five-fold cross-validation repeating five times, and each fold was obtained by category-wise sampling. all the statistical analysis was performed using spss (version ) with statistical significance set at . . statistical optimization of the deep learning model was done through iterative training using python (version . with scipy, scikit-learn, and pytorch packages). the differences of clinical and laboratory data and imaging features between the patient with and without severe/critical progression were compared using chi-square test, fisher's exact test, independent t test and paired t test. auc, accuracy, specificity, and sensitivity were compared among different ai methods and multivariable logistic regression. two-sided % cis were used to summarize the sample variability in the estimates. specifically, the normal approximation cis was used for accuracy, sensitivity, and specificity. the ci for the auc was estimated using the bootstrap method with replications. patients with mild covid- pneumonia at admission included male and female, age ranged from to ( . ± . ) years, the interval from symptoms onset to admission ranged from to ( . ± . ) days. patients ( / , . %) malignantly progressed to severe/critical periods during the all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted march , . . hospitalization, while the remaining patients ( / , . %) did not ( figure ). the whole clinical course of all patients, including assessment at admission, the severe or critical progression, and the outcome, was plotted in figure . the age, sex, exposure, comorbidity, signs and symptoms, laboratory results measured at admission, and serial ct imaging features of patients with and without severe/critical progression were separately summarized in tables , , and . in brief, comparing to the patients without severe/critical progression, the patients with severe/critical progression showed older age, more comorbidities, higher respiratory rate, inflammatory cell factors, lower albumin and fewer counts of lymphocyte, t cell, and its subsets. the patients with severe/critical progression were more likely to involve organs other than the lung. on the first available ct, no difference was found in either the distribution of involved lung or the other ct imaging features, except paving stone sign and the presence of fibrosis. however, the patients with severe/critical progression showed significantly more lesions in all lobes, more lesions of consolidation, paving stone sign and halo sign than patients without severe/critical progression when they progressed to the period of severe/critical stage. were the risk factors for severe/critical progression. however, the presence of fibrosis at ct (or . , %ci . - . ) was the protective factor for severe/critical progression. the accuracy of the prediction is . %. we conducted comprehensive experiments to validate our hypotheses and compared the performance of various models. table summarized the performance of traditional multi-stage and deep learning-based methods. static clinical data including personal information, dynamic quantitative chest ct data or both of them were used for predictive experiments. for traditional multi-stage methods, pca was used for data dimensionality reduction, and svm or lda was used for classification. the results indicate that quantitative chest ct data without time series modeling is also beneficial for all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted march , . the above results clearly supported the significance of complementary information from different medical data and time-series information from the chest ct sequence. finally, our proposed method had a high probability of stabilizing at a high confidence interval, which is very important for clinical applications. with the worldwide outbreak of covid- , early prediction and early aggressive treatment of mild patients at high risk of malignant progression to severe/critical stage are important ways to reduce mortality. in this work, we found that the complementarity of clinical data and quantitative chest ct sequence is important for predicting patients with malignant progression. in particular, the rich series information of the chest ct sequence, which has not been considered by other studies so far, is critical for this specific task. we also demonstrated that our method can effectively fuse these two complementary data and handle time-series information in the quantitative chest ct sequence, which achieved an auc of . ( % ci although lots of clinical, laboratory, and imaging parameters varied significantly between patients with and without severe/critical progression, seven predictive all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted march , . unlike the traditional predictive model using a hand-crafted feature extractor and shallow classifiers, our deep learning-based method using a multilayer perceptron combined with an lstm to this predictive task, which attempts to learn high-level hierarchical features from mass data, and expands the search space of the features for specific tasks. moreover, this method jointly optimizes the feature extraction network and classifier through an end-to-end manner. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. our study has several limitations. first, samples available for malignant progression prediction were limited. the diverse data in the large scale dataset will allow deep learning-based methods to gain a more comprehensive understanding of what causes the malignant progression of mild patients. second, the quantitative information of ct data is not detailed enough. using the richer original features included in pixel-wise segmentation results of the ct scans, the predictive model may perform better. in conclusion, the deep learning-based method using clinical and quantitative ct data to predict malignant progression to severe/critical stage. we modeled the spatial information in the quantitative ct data and organized the static clinical data and dynamic chest ct data into a time series form. we validated the significance of complementary data and its special formatting form for this particular prediction task. compared with traditional multi-stage methods, we demonstrate that our deep learning-based method can extract spatial and temporal information efficiently and improve the prediction performance significantly. the ability to identify patients with potentially severe and critical covid- outcomes using an inexpensive, widely available, the point-of-care test has important practical implications for preventing mild patients from becoming severe, effectively improving cure rate, and reducing mortality. our future work will focus on mining richer spatial information from the ct scan sequence and using ai technologies to screen the risk factors of potential severe/critical patients. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. estimates of global seasonal influenza-associated respiratory mortality: a modelling study influenza-associated excess respiratory mortality in china, - : a population-based study clinical characteristics of hospitalized patients with novel coronavirus-infected pneumonia in wuhan, china clinical characteristics and imaging manifestations of the novel coronavirus disease (covid- ):a multi-center study in wenzhou city outbreak of novel coronavirus (covid- ): what is the role of radiologists? radiological findings from patients with covid- pneumonia in wuhan, china: a descriptive study clinical predictors of mortality due to covid- based on an analysis of data of patients from wuhan deep learning for prediction of colorectal cancer outcome: a discovery and validation study deep learning algorithms for detection of critical findings in head ct scans: a retrospective study an end-to-end trainable neural network for image-based sequence recognition and its application to scene text recognition critical care crisis and some recommendations during the covid- epidemic in china pathological findings of covid- associated with acute respiratory distress syndrome. the lancet respiratory journal of thoracic oncology : official publication of the international association for the study of lung cancer analysis of factors associated with disease outcomes in hospitalized patients with novel coronavirus disease laboratory abnormalities in patients with covid- infection the clinical and chest ct features associated with severe and critical covid- pneumonia radiologyessentials for radiologists on covid- : an update-scientific expert panel key: cord- -jzfr t p authors: mudatsir, mudatsir; fajar, jonny karunia; wulandari, laksmi; soegiarto, gatot; ilmawan, muhammad; purnamasari, yeni; mahdi, bagus aulia; jayanto, galih dwi; suhendra, suhendra; setianingsih, yennie ayu; hamdani, romi; suseno, daniel alexander; agustina, kartika; naim, hamdan yuwafi; muchlas, muchamad; alluza, hamid hunaif dhofi; rosida, nikma alfi; mayasari, mayasari; mustofa, mustofa; hartono, adam; aditya, richi; prastiwi, firman; meku, fransiskus xaverius; sitio, monika; azmy, abdullah; santoso, anita surya; nugroho, radhitio adi; gersom, camoya; rabaan, ali a.; masyeni, sri; nainu, firzan; wagner, abram l.; dhama, kuldeep; harapan, harapan title: predictors of covid- severity: a systematic review and meta-analysis date: - - journal: f res doi: . /f research. . sha: doc_id: cord_uid: jzfr t p background: the unpredictability of the progression of coronavirus disease (covid- ) may be attributed to the low precision of the tools used to predict the prognosis of this disease. objective: to identify the predictors associated with poor clinical outcomes in patients with covid- . methods: relevant articles from pubmed, embase, cochrane, and web of science were searched and extracted as of april , . data of interest were collected and evaluated for their compatibility for the meta-analysis. cumulative calculations to determine the correlation and effect estimates were performed using the z test. results: in total, papers recording , mild and , severe cases of covid- were included. based on the initial evaluation, potential risk factors were identified for the meta-analysis. several comorbidities, including chronic respiratory disease, cardiovascular disease, diabetes mellitus, and hypertension were observed more frequent among patients with severe covid- than with the mild ones. compared to the mild form, severe covid- was associated with symptoms such as dyspnea, anorexia, fatigue, increased respiratory rate, and high systolic blood pressure. lower levels of lymphocytes and hemoglobin; elevated levels of leukocytes, aspartate aminotransferase, alanine aminotransferase, blood creatinine, blood urea nitrogen, high-sensitivity troponin, creatine kinase, high-sensitivity c-reactive protein, interleukin , d-dimer, ferritin, lactate dehydrogenase, and procalcitonin; and a high erythrocyte sedimentation rate were also associated with severe covid- . conclusion: more than risk factors are associated with a higher risk of severe covid- . these may serve as useful baseline parameters in the development of prediction tools for covid- prognosis. the coronavirus disease (covid- ) pandemic, caused by severe acute respiratory syndrome coronavirus (sars-cov- ), is a global crisis across health, economic, and educational dimensions , . the disease has spread rapidly, can cause severe illness, and is characterized by a high mortality rate in certain groups. mortality is particularly high in the absence of proven effective standard management measures . one of the problems with the management of this disease is the absence of standardized methods for diagnosis and the inability to estimate prognosis based on clinical features. certain reports have shown that poor prognostic prediction has correlated with high mortality among patients with covid- , . among patients with similar clinical characteristics and with similar treatment regiments, there may be a diversity in clinical outcomes . therefore, the development and use of an accurate predictor for covid- prognosis will be beneficial for the clinical management of patients with covid- , and will help reduce the mortality rate. successful implementation of such a prediction mechanism could have a large public health impact. better understanding of clinical progression could also improve public health messaging, particularly as many individuals may consider covid- to not be severe. prognostic tools for the prediction of covid- severity in patients have been in development since january . at least nine studies proposed the use of prognostic tools for the prediction of covid- severity [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . however, a recent systematic review and critical appraisal study evaluated the accuracy of these tools using prediction model risk of bias assessment tool (probast) and reported a high risk of bias . the establishment of a prediction model for the estimation of disease prognosis may help health workers segregate patients according to prediction status. however, the high risk of bias in these prediction tools might lead to inaccurate prediction of covid- severity. a comprehensive study of the identification of risk factors that might play a significant role in determining the severity of patients with covid- is necessary. we performed a systematic review and meta-analysis to assess the risk factors associated with poor clinical outcomes among patients with covid- . to the best of our knowledge, this is the first meta-analysis to assess the comprehensive risk factors that might affect the severity of covid- in patients. the results of our study might serve as preliminary data for the compilation or improvement of the scoring system in the prediction of covid- severity. we performed a systematic review and meta-analysis to evaluate potential risk factors that might influence the severity of covid- . these risk factors include comorbidities, clinical manifestations, and laboratory findings. accordingly, we searched the relevant studies from major scientific websites and databases to collect the data of interest, and determined the association and effect estimates by calculating the combined odds ratio (or) and % confidence intervals ( % ci). the protocols for the systematic review and meta-analysis were similar to those used in previous studies - , as well as to those recommended by preferred reporting items for systematic review and meta-analysis (prisma) . studies were included in this review if they met the following inclusion criteria: ( ) assessed the clinical manifestations and laboratory findings of patients with mild to severe covid- ; ( ) provided adequate data for the calculation of or and % ci. review articles, articles with non-standard data presentation, and duplicate publications were excluded. search strategy and data extraction major scientific databases (pubmed, embase, cochrane, and web of science) were searched for articles as of april , . a comprehensive initial search was performed to identify the potential predictors, and a final search was performed to identify the relevant papers that could be included in the meta-analysis. we used the keywords adapted from medical subject headings: ["covid- " or "coronavirus disease- " or "sars-cov- "] and ["mild" or "severe" or "prognosis" or "clinical outcome"] and ["clinical manifestation" or "morbidity" or "laboratory findings"]. only studies written in english were included. if a duplicate publication was found, the article with the larger sample size was included. we also searched for relevant studies from the reference lists in the articles. during data extraction, the following information of interest was extracted: ( ) first author name; ( ) publication year; ( ) sample size of mild and severe cases, ( ) clinical manifestations, ( ) morbidities, and ( ) laboratory findings. data extraction was performed by two independent investigators (jkf and mi) using a pilot form. before inclusion in the meta-analysis, the methodological quality of the articles was assessed using the new castle-ottawa scale (nos). nos scores range from to and consider three items: selection of patients ( points), comparability of the groups ( points), and ascertainment of exposure ( points). each study was interpreted to be of low quality (for scores ≤ ), moderate quality (for scores between - ), or high quality (for scores ≥ ) . articles with moderate to high quality were included in the analysis. the study assessment was conducted by two independent investigators (mi and yp) using a pilot form. the discrepancies between the findings of the two investigators were solved by consulting with another investigator (jkf). the outcome measure of the study was the severity of covid- (mild vs. severe). the risk factors or predictors included three major groups: comorbidities, clinical manifestations, and laboratory parameters. comorbid factors such as chronic kidney disease, chronic liver disease, chronic respiratory disease, cerebrovascular accident, cardiovascular disease, diabetes mellitus, hypertension, and malignancy were compatible with the analysis. for clinical manifestations, fever, cough, dry cough, expectoration, sore throat, dyspnea, diarrhea, myalgia, nasal congestion, anorexia, abdominal pain, fatigue, dizziness, headache, fever, heart rate, respiratory rate, systolic blood pressure, and diastolic blood pressure were included in this study. among laboratory characteristics, the presence of leukocytosis, leukocytopenia, anemia, lymphocytopenia; the levels or the counts of white blood cell (wbc), hemoglobin, neutrophil, lymphocyte, monocyte, platelet, activated partial thromboplastin time (aptt), partial thromboplastin time (ptt), aspartate aminotransferase (ast), alanine aminotransferase (alt), total bilirubin, albumin, serum creatinine, blood urea nitrogen (bun), high-sensitivity (hs)-troponin i, creatine kinase, high-sensitivity c-reactive protein (hs-crp), c-reactive protein (crp) > mg/l, interleukin (il- ), glucose, d-dimer, serum ferritin, sodium, potassium, lactate dehydrogenase, and procalcitonin, cd and cd ; erythrocyte sedimentation rate (esr); elevated il- ; and elevated esr were all included. the significant risk factors that might govern the severity of covid- were determined by the calculation of a pooled or and % ci. the significance of the pooled ors was determined using the z test (p< . was considered statistically significant). prior to identification of the significant risk factors, data were evaluated for heterogeneity and potential publication bias. the heterogeneity among included studies was evaluated using the q test. if heterogeneity existed (p< . ), a random effect model was adopted; if not, a fixed effect model was adopted. egger's test and a funnel plot were used to assess the reporting or publication bias (p< . was considered statistically significant). furthermore, we performed a moderator analysis to identify the independent predictors of poor clinical outcomes among patients with covid- . the data were analyzed using review manager version . (revman cochrane, london, uk). to prevent analytical errors, statistical analysis was performed by two authors (jkf and mi). the cumulative calculation was presented in a forest plot. our searches yielded , potentially relevant studies, of which , studies were excluded after assessment of the titles and abstracts. subsequently, further review of the complete texts was performed for potential studies. in the full text review, we excluded studies because they were reviews articles (n = ), inadequacy of data for the calculation of or and % ci (n = ), and poor quality (n = ). eventually, papers were included in our meta-analysis - the paper selection process adopted in our study is summarized in figure , and the characteristics of studies included in our analysis are outlined in table . we found that eight comorbidities, clinical manifestations, and laboratory parameters were available for the metaanalysis (table and table (figure a-d) . among the clinical manifestations, dyspnea (or: . ; % ci: . , . ), anorexia (or: . ; % ci: . , . ), fatigue (or: . ; % ci: . , . ), and dizziness (or: . ; % ci: . , . ) were associated with severe covid- ( figure a -d). in addition, increased respiratory rate (or: . ; % ci: . , . ) and increased systolic blood pressure (or: . ; % ci: . , . ) were also associated with severe covid- ( figure a and b). compared to productive cough, dry cough was associated with a lower risk of severe covid- (or: . ; % ci: . , . ). among laboratory characteristics, severe covid- was associated with elevated wbc count (or: . ; % ci: . , . ), increased neutrophil count (or: . ; % ci: . , . ), lymphocytopenia (or: . ; % ci: . , . ), and decreased hemoglobin levels (or: . ; %ci: . , . ) ( figure a -d). elevated levels of ast, alt, and serum creatinine increased the risk for severe manifestations of covid- (ors . , . , and . , respectively; figure heterogeneity was detected in the data of chronic kidney disease, cerebrovascular disease, cardiovascular disease, diabetes mellitus, hypertension, and malignancy among the comorbid factors analyzed. therefore, we used the random effect model to analyze the data. the fixed effect model was used to analyze the data on chronic liver disease and chronic respiratory disease, as there was no evidence of heterogeneity. for clinical manifestations, the data on fever, cough, sore throat, dyspnea, diarrhea, anorexia, fatigue, temperature > °c, respiratory rate, and diastolic blood pressure were analyzed using the random effect model while the rest of clinical manifestation data were analyzed using the fixed effect model. among laboratory parameters, evidence of heterogeneity was found in count of wbc, neutrophil, monocyte, lymphocyte, platelet, cd , and cd ; the presence of lymphocytopenia and anemia; the levels of ast, alt, total bilirubin, albumin, aptt, ptt, serum creatinine, bun, hs-troponin i, creatine kinase, il- , hs-crp, glucose, d-dimer, sodium, potassium, lactate dehydrogenase, and procalcitonin; elevated crp; and esr. accordingly, the data were analyzed using the random effect model. the data for the remaining parameters were analyzed using the fixed effect model. we used egger's test to assess the potential publication bias. our cumulative calculation revealed that reporting or publication bias (p< . ) existed with respect to chronic liver disease, expectoration, myalgia, abdominal pain, heart rate, leukocytosis, elevated esr, and elevated il- levels. our data suggest that comorbidities, such as chronic respiratory disease, cardiovascular disease, diabetes, and hypertension, were associated with a higher risk of severe covid- , among which, hypertension was the strongest risk factor. these results are consistent with those of previous meta-analyses , that indicated that chronic respiratory disease, cardiovascular disease, diabetes, and hypertension are significantly associated with higher covid- mortality. hypertension and diabetes are also associated with higher mortality among patients with dengue fever, west nile virus infection, zika virus infection, and yellow fever . to date, no study has reported details of the primary mechanism underlying the association between severe covid- and comorbid factors. however, immune responses might be the most crucial factor underlying this association. patients with comorbidities such as cardiovascular disease, chronic respiratory disease, hypertension, and diabetes were observed to have a lower immunity status than healthy individuals - . since covid- primarily affects the respiratory tract , patients with chronic respiratory diseases might be at a higher risk of contracting severe covid- . in addition, endothelial dysfunction might also play a pivotal role . covid- is a novel disease, and the immune response of this disease is not completely understood. our data suggest that elevated leukocyte and neutrophil levels and reduced lymphocyte levels are associated with severe covid- . in other viral infections, such as influenza, elevated leukocyte and neutrophil levels serve as important predictors of disease severity . the role of leukocytes in the pathogenesis of covid- is conflicting. in most cases, viral infections have been observed to cause leukopenia . furthermore, a study also reported that leukopenia was observed at a significantly higher frequency among covid- patients than among non-covid- patients . however, in our present study, we did not compare covid- and non-covid- patients. the major factor that seemed to affect our findings was the occurrence of cytokine storm in patients. in covid- , there is an immune system overreaction, which results in a cytokine storm. in this condition, leukocytes might be over-activated, which might lead to the release of high levels of cytokines . consistent with our data, a study has confirmed that cytokine storm is significantly associated with severe covid- . the theory underlying the role of neutrophils in covid- , as reported in our study, remains unclear. the speculations might be attributed to the involvement of neutrophil extracellular traps (nets). while no study has assessed the precise role of nets in covid- pathogenesis, certain researchers speculate that sars-cov- might stimulate neutrophils to produce nets, similar to several other viral pathogens . furthermore, this might lead to neutrophil infiltration in pulmonary capillaries, organ damage, and the development of acute respiratory distress syndrome . low lymphocyte levels were observed in patients with severe covid- compared with those with mild covid- . in the context of the immunological mechanism, our results might be contradictory. lymphocyte subsets are known to play an important role in the action against bacterial, viral, fungal, and parasitic infections ; therefore, the levels of circulating lymphocytes should increase. the immunological response in covid- is unique and remains unclear. however, certain propositions might help describe our findings. first, coronaviruses infect human cells through ace receptors . since ace receptors are also expressed by lymphocytes , the coronaviruses may enter lymphocytes and induce apoptosis. second, the feedback mechanism between pro-inflammatory cytokines (such as il- ) and lymphocytes might also explain our results. a study revealed that elevation in the levels of pro-inflammatory cytokines correlated with reduction in the levels of lymphocytes . moreover, our findings also confirmed the significant elevation in the levels of il- . third, ace receptors are expressed by cells from various organs, including the thymus and spleen . as coronaviruses infect human cells through the ace receptors, the spleen and thymus might also be damaged in patients with covid- , which would lead to lower levels of lymphocyte production. fourth, lymphocyte proliferation requires a balanced metabolism, and metabolic disorders such as hyperlactic acidemia have been reported to disturb lymphocyte proliferation . hyperlactic acidemia has been observed in patients with severe covid- . the studies included in this systematic review also suggest that the levels of d-dimer were significantly higher in patients with severe covid- . coagulation in patients with covid- has been a major concern, and the lack of reliable data and meta-analyses prevents a holistic comparison. certain infectious diseases that cause abnormal coagulation have been associated with poor clinical outcomes . the theory behind this mechanism is not understood clearly. it is widely known that ace receptors are important for the infection of host cells by sars-cov- , and ace receptors are expressed in various cells in the human body, including endothelial cells . consequently, a massive inflammatory reaction may occur in endothelial cells owing to sars-cov- infection , which may lead to increased coagulation, disseminated intravascular coagulation , and increased fibrin degradation . high fibrin degradation leads to elevated levels of fibrinogen and d-dimer , which might also explain the occurrence of venous thromboembolism in critical patients of covid- . in addition, a study with a short follow-up period also reported the existence of a dynamic correlation between the d-dimer levels and the severity of covid- . furthermore, pulmonary embolism and deep vein thrombosis were also observed in patients with severe covid- , , which suggests that d-dimer might play a prominent role in governing the severity of covid- patients. we also observed that inflammatory markers, including elevated levels of crp, esr, and il- , were found both in patients with severe and mild covid- , with a significant increase detected in patients with severe covid- . other variables associated with adverse outcomes, such as ferritin, lactate dehydrogenase, and procalcitonin levels, were found to be elevated predominantly in patients with severe covid- . our findings were consistent with those of a previous meta-analysis , and indicated that high levels of crp, lactate dehydrogenase, and esr were associated with adverse outcomes in covid- . another meta-analysis had also confirmed that elevated levels of il- were observed in patients with covid- who exhibited poor clinical outcomes . therefore, the levels of crp, esr, il- , ferritin, procalcitonin, and lactate dehydrogenase can serve as potential markers for the evaluation of covid- prognosis. the high mortality rate and treatment failure in patients with covid- can be attributed to the fact that covid- affects multiple organs, including the lung, heart, kidney, and liver . our data suggest that elevated levels of urea and creatinine, and not chronic kidney disease, were associated with severe covid- , which indicates that acute inflammation might be caused by sars-cov- infection. previous meta-analyses have also reported findings consistent with our results , . moreover, anatomical studies have reported significant renal inflammation in patients with severe covid- , , . there might be two mechanisms by which sars-cov- induces renal inflammation. first, sars-cov- might directly infect renal tubular epithelial cells and podocytes through ace receptors, which facilitates the targeted infection of certain cells by the virus. consequently, acute tubular necrosis, podocytopathy, microangiopathy, and collapsing glomerulopathy might occur owing to the massive inflammation in renal tubular epithelial cells and podocytes , . second, the binding between sars-cov- and ace receptors might activate angiotensin ii and induce cytokine production, which may lead to hypercoagulopathy and microangiopathy, and eventually cause renal hypoxia , . conversely, with respect to liver function, we observed that the levels of liver enzymes were higher in patients with severe covid- . previous studies in this context have elucidated that ace receptors are highly expressed in bile duct cells; therefore, infection of these cells by coronaviruses might lead to abnormalities in the levels of liver enzymes . however, a recent anatomical study on liver biopsy specimens from patients with severe covid- revealed that moderate microvascular steatosis and mild lobular and portal activities were observed . these data suggest that it cannot be determined clearly whether the elevated levels of liver enzymes in patients with severe covid- are caused by direct infection or by drug-induced liver injury. therefore, further studies are required to elucidate the precise mechanism underlying the elevation of liver enzymes levels in patients with severe covid- . . in our study, we included all comorbidities, clinical manifestations, and laboratory characteristics. additionally, compared to previous studies, this study has a larger sample size; the data on , patients with mild and , patients with severe covid- treated across hospitals were retrieved. however, this study also has certain limitations. certain crucial factors that might play an important role in the pathogenesis of covid- , including secondary infection, treatment, and immunological status were not controlled for. our current findings should be interpreted with caution because the majority of studies included were cross-sectional, and the samples corresponding to the data analyzed originated only in china. longitudinal studies may reveal more long-term impacts of sars-cov- infection . covid- is an emergent infectious disease, and the major problem associated with it is the unknown pattern of disease development. we identified factors that are associated with severe covid- . this might improve our understanding of covid- progression and provide baseline data to compile or improve the prediction models for the estimation of covid- prognosis. underlying data all data underlying the results are available as part of the article and no additional source data are required. the socio-economic implications of the coronavirus pandemic (covid- ): a review prevalence and fatality rates of covid- : what are the reasons for the wide variations worldwide predicting the mortality due to covid- by the next month for italy, iran and south korea; a simulation study pubmed abstract | publisher full text | free full text . siordia ja: epidemiology and clinical features of covid- : a review of current literature predicting covid- malignant progression with ai techniques a tool to early predict severe -novel coronavirus pneumonia (covid- ):a multicenter study using the risk nomogram in wuhan and guangdong, china host susceptibility to severe covid- and establishment of a host risk score: findings of cases outside wuhan neutrophil extracellular traps inhibitory effect of tumor cell-derived lactic acid on human t cells a fatal case of covid- due to metabolic acidosis following dysregulate inflammatory response (cytokine storm) infection and inflammation and the coagulation system angiotensin converting enzyme- confers endothelial protection and attenuates atherosclerosis endothelial cell infection and endotheliitis in covid- covid- and haemostasis: a position paper from italian society on thrombosis and haemostasis (siset) pubmed abstract | publisher full text | free full text covid- update: covid- -associated coagulopathy pubmed abstract | publisher full text | free full text fibrin d-dimer in thrombogenic disorders the need to manage the risk of thromboembolism in covid- patients d-dimer in patients infected with covid- and suspected pulmonary embolism covid- complicated by acute pulmonary embolism and right-sided heart failure venous thrombosis among critically ill patients with coronavirus disease (covid- ) clinical, laboratory and imaging features of covid- : a systematic review and meta-analysis elevated interleukin- and severe covid- : a meta-analysis covid- and multiorgan response prevalence and impact of acute renal impairment on covid- : a systematic review and meta-analysis incidence of acute kidney injury in covid- infection: a systematic review and meta-analysis kidney biopsy findings in a critically ill covid- patient with dialysis-dependent acute kidney injury: a case against "sars-cov- nephropathy covid- ): a literature review acute kidney injury in covid- : emerging evidence of a distinct pathophysiology pubmed abstract | publisher full text | free full text covid- -associated collapsing glomerulopathy: an emerging entity hyperinflammation and derangement of renin-angiotensin-aldosterone system in covid- : a novel hypothesis for clinically suspected hypercoagulopathy and microvascular immunothrombosis interactions of coronaviruses with ace , angiotensin ii, and ras inhibitors-lessons from available evidence and insights into covid- pubmed abstract | publisher full text | free full text specific ace expression in cholangiocytes may cause liver damage after -ncov infection. biorxiv. . publisher full text pathological findings of covid- associated with acute respiratory distress syndrome the clinical manifestations and chest computed tomography findings of coronavirus disease (covid- ) patients in china: a proportion meta-analysis laboratory findings of covid- : a systematic review and meta-analysis predictive symptoms and comorbidities for severe covid- and intensive care unit admission: a systematic review and meta-analysis pubmed abstract | publisher full text | free full text covid- patients' clinical characteristics, discharge rate, and fatality rate of meta-analysis symptom duration and risk factors for delayed return to usual health among outpatients with covid- in a multistate health care systems network -united states predictors of covid- severity: a systematic review and meta-analysis". figshare publisher full text are the rationale for, and objectives of, the systematic review clearly stated? yes are sufficient details of the methods and analysis provided to allow replication by others? yes is the statistical analysis and its interpretation appropriate? yes the authors highlight the need for a scoring system for the prediction of severity. there is another reason why it is important to identify risk factors for severe disease: to guide prioritization of high risk target populations for vaccination competing interests: no competing interests were disclosed.reviewer expertise: covid- , zika and dengue i confirm that i have read this submission and believe that i have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.reviewer report september https://doi.org/ . /f research. .r © arab-zozani m. this is an open access peer review report distributed under the terms of the creative commons attribution license, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. in this meta-analysis, you investigated the predictors of covid- severity through the literature. you considered a topic of interest and provided a well-written manuscript. however, there are some things that will improve your reporting.abstract, method section, please insert detail about critical/quality appraisal of the included studies.○ abstract, method section, line , please remove " and extracted" from the text. it maybe causes a misunderstanding between this step and the data extraction step. are the rationale for, and objectives of, the systematic review clearly stated? yes are the conclusions drawn adequately supported by the results presented in the review? yescompeting interests: no competing interests were disclosed.reviewer expertise: systematic review and meta-analysis in health and medical intervention i confirm that i have read this submission and believe that i have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.the benefits of publishing with f research:your article is published within days, with no editorial bias • you can publish traditional articles, null/negative results, case reports, data notes and more • the peer review process is transparent and collaborative • your article is indexed in pubmed after passing peer review • dedicated customer support at every stage • for pre-submission enquiries, contact research@f .com key: cord- - uodkrkp authors: bonam, srinivasa reddy; kaveri, srini v.; sakuntabhai, anavaj; gilardin, laurent; bayry, jagadeesh title: adjunct immunotherapies for the management of severely ill covid- patients date: - - journal: cell reports medicine doi: . /j.xcrm. . sha: doc_id: cord_uid: uodkrkp abstract coronavirus disease (covid- ) is caused by severe acute respiratory syndrome coronavirus (sars-cov- ). it has infected millions with more than , fatal cases as of nd april . currently, there are no specific covid- therapies. most patients depend on mechanical ventilation. current covid- data clearly highlight that cytokine storm and activated immune cell migration to the lungs characterize the early immune response to covid- that causes severe lung damage and development of acute respiratory distress syndrome. in view of uncertainty associated with immunosuppressive treatments such as corticosteroids and their possible secondary effects, including risks of secondary infections, we suggest immunotherapies as an adjunct therapy in severe covid- cases. such immunotherapies based on inflammatory cytokine neutralization, immunomodulation and passive viral neutralization, not only reduce inflammation, inflammation-associated lung damage, or viral load, but could also prevent intensive care unit hospitalization and dependency on mechanical ventilation both of which are limited resources. coronavirus disease (covid- ) is caused by "severe acute respiratory syndrome coronavirus " (sars-cov- ). it has affected over . million people with more than , fatal cases as of nd april (https://coronavirus.jhu.edu/map.html). while the majority of cases result in mild symptoms such as fever and cough, about - % progress to severe pneumonia and respiratory failure requiring intensive care unit hospitalization and mechanical ventilation. the death rate in intensive care unit (icu) admitted patients could reach up to %. age and other health problems such as chronic pulmonary or cardiac disease conditions are the risk factors that determine the mortality rate. [ ] [ ] [ ] [ ] a systematic review and meta-analysis confirmed that old age and/or associated comorbidity are the common risk factors that predict death in all three major respiratory diseases caused by coronaviruses including covid- , severe acute respiratory syndrome (sars) and middle east respiratory syndrome (mers). obesity is another risk factor for the development of severe sars-cov- . though anti-viral molecules and hydroxychloroquine are under investigation to avert severe disease, [ ] [ ] [ ] [ ] [ ] currently, there are no specific therapies for covid- , particularly when severe acute respiratory distress syndrome (ards) occurs. most covid- patients with ards required intensive care unit hospitalization and prolonged supportive mechanical ventilator management. , , ards can lead to other complications, including secondary bacterial infection and lung fibrosis. recent clinical data suggest that severe covid- is due to an overreactive immune response leading to a cytokine storm and development of acute respiratory distress syndrome. , plasma obtained from covid- patients, in particular moribund patients, demonstrated increased concentrations of various inflammation-related cytokines and chemokines implicated in the recruitment of immune cells including interleukin (il)- β, tumor necrosis factor (tnf)-α, il- , il- , il- , il- , il- , il- , granulocyte-colony stimulating factor (g-csf), granulocytemacrophage colony-stimulating factor (gm-csf), interferon (ifn)-γ, c-x-c motif chemokine (cxcl ), chemokine ligand (ccl ), ccl and ccl . , , in addition, covid- patients showed relatively increased neutrophil counts in the blood. on the other hand, monocytes, basophils and t cells, especially cd + and cd + t cells, were decreased in the peripheral blood, possibly due to their migration to the lungs. [ ] [ ] [ ] [ ] [ ] however, these t cells exhibited enhanced activated markers (cd , cd and cd ). single-cell rna sequencing of bronchoalveolar lavage (bal) fluid immune cells from a small number of mild to severely ill covid- patients suggested that balance in the lung macrophage populations is highly dysregulated in severely ill covid- patients. these data indicate that monocyte-derived macrophages (fcn + ) rather than fabp + alveolar macrophages contribute to lung inflammation and lung damage in severe cases. this study thus confirms another report based on lung postmortem biopsy of a patient that revealed the presence of bilateral diffuse alveolar damage and recruitment of monocytes. the study by liao et al. also observed an increased proportion of clonally expanded cd + t cells in the bal fluid of mild cases as compared to severe cases. another investigation that compared bal fluid immune cells in various respiratory pathologies highlighted a more prominent surge of neutrophils in covid- patients as compared to pneumonia caused by other pathogens. by metatranscriptome sequencing of bal fluid and global functional analyses of differentially expressed genes, this report also identified strong upregulation of numerous type i ifn-inducible genes. however, caution needs to be exercised while interpreting these data due to potential influence of therapies such as ifnα- b, ant-viral and/or steroids on the landscape of immune cells and immune signatures of bal fluid or lungs. nevertheless, these reports confirm the proposition that an influx of immune cells to the lungs follows sars-cov- infection (figure ). severely ill covid- patients also displayed reduced peripheral blood regulatory t cells (tregs), the immune suppressor cells critical for reducing inflammation and inflammationassociated tissue damage. another report suggested that activated gm-csf + ifnγ + pathogenic th cells that secrete gm-csf promote inflammatory cd + cd + monocyte responses with enhanced il- . gm-csf + ifnγ + th cells and inflammatory monocytes were positively correlated with the severe pulmonary syndrome characteristic of covid- patients. the simultaneous increase in il- receptor antagonist (il- ra) and il- also suggest that antiinflammatory responses, though induced, are not sufficient to reduce inflammation and eventually lead to severe lung damage. various reports have shown that higher inflammation-related biomarkers such as plasma creactive protein (crp), ferritin and il- were significantly associated with higher risks of developing ards. [ ] [ ] [ ] [ ] [ ] of note, il- levels were associated with course of the disease and death from covid- . the massive increase in plasma ferritin levels is indicative of haemophagocytic lymphohistiocytosis activation syndrome in these patients. these studies thus provide a rationale for targeting inflammatory mediators for the management of severely ill covid- patients. currently, there is no clear evidence for the use of steroids in sars-cov- infections and their use is highly debated, particularly with respect to the window of treatment, dose and management of patients in cases of bacterial co-infection. a retrospective cohort analysis of patients from wuhan suggested that methylprednisolone might benefit patients who develop ards (n= ) by reducing the death rate. a retrospective analysis of hospitalized patients with severe covid- pneumonia (n= ) indicated that an early low-dose steroid therapy ( - mg/kg/day) in patients for a short duration ( - days) reduced the oxygen requirement period and improved disease course. however, covid- patients treated with methylprednisolone were sicker and had a higher pneumonia severity index than those patients who did not receive methylprednisolone. recently, an additional retrospective matched case-control study involving paired patients in several icus in china showed that treatment with steroids for days ( - days) in addition to antiviral drugs was associated with a % -day death rate, compared to % in a propensity score matched control group. clinical evidence is also lacking for the use of steroids in other coronavirus diseases (sars-cov, mers-cov), , and steroids might even impair viral clearance mechanisms of patients and predispose them to secondary infections. double-blinded randomized clinical trials involving a large number of patients should be conducted to evaluate the use of steroids in severely ill covid- patients with reduction in intensive care unit requirement as the primary endpoint. therefore, as per who interim guidance of th march , and some local guidelines (covidprotocols.org), steroids are not recommended at present as a treatment for covid- . if steroids are used for a particular pathological condition, low dose and short duration of treatment is recommended. similarly, as per a survey from the french national agency for medicines and health products safety (ansm), non-steroidal anti-inflammatory drugs (nsaids) like ibuprofen are also not recommended due to infectious complications and kidney, cardiac and gastrointestinal adverse effects. in view of an uncertain prognosis and complications due to generalized immunosuppression from corticosteroid and nsaid treatment, immunotherapies based on immunomodulatory approaches appear promising in the treatment of covid- to regulate inflammation and antiviral immune responses. we believe that the immunotherapies discussed next could result in a quicker therapeutic response with minimal adverse effects. the current clinical data on covid- patients clearly highlight that cytokine storm and an influx of activated immune cells to the lungs characterize the early immune response to covid- that causes severe lung damage. therefore, we suggest host-directed immunotherapies as an adjunct therapy in severe cases to not only reduce inflammation and inflammation-associated lung damage, but also to prevent icu hospitalization and dependency on mechanical ventilation that are limited resources in the context of sars-cov- pandemic. in fact, several immunotherapeutic approaches that target either inflammatory mediators, passively neutralize sars-cov- , or prevent viral entry are under evaluation at various centers (table patients. , , an increase in il- leading to lung tissue damage has been observed in a majority of the covid- patients. , , , of note, similar to severely ill covid- cases, elevated serum levels of il- , tnf-α and ifn-γ have been consistently observed in cytokine release syndrome (crs) that is common in the patients receiving t cell-engaging immunotherapies (bispecific antibody constructs or chimeric antigen receptor (car) t cell therapies). targeting either il- (siltuximab, a chimeric monoclonal antibody) or il- receptor (tocilizumab, a recombinant humanized monoclonal antibody) led to rapid resolution of crs symptoms in those patients. severe crs can also lead to hypotension, pulmonary edema and cardiac dysfunction that are commonly observed in severely ill covid- patients. these data provide justification for targeting il- in severely ill covid- patients. both anti-il- receptor antibodies (tocilizumab, and a human monoclonal antibody sarilumab (kevzara®)), and anti-il- antibody siltuximab (sylvant®) are under evaluation in covid- patients at various centers (table ). an immunosuppressed patient with severe covid- related lung disease was successfully treated with tocilizumab and therapy led to a partial decrease in the pulmonary infiltrates. unpublished data based on off-label use of tocilizumab along with lopinavir and methylprednisolone in severely ill covid- patients suggested that the addition of tocilizumab not only quickly reduced their clinical symptoms (fever, oxygen requirement) and inflammation, but also normalized peripheral blood t cell counts in the majority of patients. additional targets include tnf-α (infliximab, a chimeric monoclonal antibody to tnf-α; etanercept, a recombinant fusion tnf-r that binds soluble tnf-α) and ifn-γ (emapalumab, a fully human igg monoclonal antibody against ifn-γ) ( table ) . of note, clinical trials in idiopathic pneumonia syndrome have reported favorable response to etanercept therapy. , gm-csf represents another target for severe cases of covid- . gm-csf stimulates the influx of granulocytes and monocytes from bone marrow, and hence further enhances inflammatory responses. a randomized phase b/ , double-blind, placebo-controlled clinical trial is currently recruiting patients to investigate the therapeutic efficacy of a humanized anti-gm-csf igg monoclonal antibody tj in severely ill covid- patients (nct ). in addition, another human monoclonal antibody that targets gm-csf (gimsilumab; roivant sciences ltd., altasciences co. inc.) is under consideration for treating ards in covid- patients. , likewise, baricitinib, a selective janus kinase (jak /jak ) inhibitor, inhibits gp family cytokines (such as il- , il- , il- and ifn-γ). in addition, baricitinib inhibits the ap associated protein kinase and cyclin g-associated kinases that are regulators of viral endocytosis process and hence could block viral infection of cells. similarly, anakinra (kineret®), a recombinant human il- ra and fda-approved janus kinase inhibitor, fedratinib (inrebic®) also possess the capacity to suppress inflammation (table ) . though, jak inhibitors can impair type i ifn-mediated anti-viral responses, recent data from metatranscriptome sequencing of bal fluid from eight covid- patients highlight that sars-cov- induces strong type i ifn responses with an overexpression of a large number of type i ifn-inducible genes implicated in inflammation. therefore, use of jak inhibitors might help in alleviating inflammation, but dose and window of treatment should be tailored to ensure that anti-viral response of the patient is not totally curtailed. type i ifn (β α and β α) is under consideration for severe covid- ( table ) . type i ifn signals through jak-stat (signal transducers and activators of transcription) pathway and upregulates ifn-stimulated genes that kill viruses in the infected cells. our knowledge on the role of type i ifn in the pathogenesis of sars-cov- infection is not complete. current evidence, however, reveals an overexpression of a large number of type i ifn-inducible genes with immunopathogenic potentials in covid- patients. also, results on the use of ifn-α in sars patients are inconclusive. therefore, care should be taken regarding the dosage of type i ifn and timing of treatment to avoid deleterious effects. additional immunotherapies that could be considered for covid- include immunomodulators that have broad anti-inflammatory effects such as pooled normal igg or intravenous immunoglobulin (ivig) therapy. obtained from the pooled plasma of several thousand healthy donors, ivig is one of the most widely used immunotherapies for a large number of autoimmune and inflammatory diseases. , a recent open-label trial in three patients reported benefits of ivig therapy ( . g/kg for five days) in severe sars-cov- -induced pneumonia thus providing another option for the management of covid- patients. another multicenter retrospective cohort study indicated that ivig therapy could benefit critical covid- patients, but dose ( . - . g/kg) and duration of therapy ( - days) were too variable. however, grouping of patients based on the dose of ivig (> g/day or ≤ g/day) and timing of infusion (> or ≤ days since hospitalization) implied that beneficial effects are associated with high dose and early ivig treatment. a retrospective study on severe or critically ill cases of covid- also found that adjunct therapy with ivig within hours of hospitalization reduced hospital stay and ventilator use, and improved -day mortality. other than il- and crp, inflammatory parameters following ivig immunotherapy were not evaluated in detail in these patients. based on the vast literature on the mechanisms of ivig, we suggest that ivig suppresses the activation and secretion of inflammatory cytokines from the innate immune cells, blocks the inflammatory th and th responses, exerts scavenging effects on complement cascade and possibly enhances tregs. , in addition, ivig contains antibodies to diverse pathogens and their antigens, , and hence secondary infections could also be prevented in ivig-treated patients. though these studies provide a pointer for initiating a randomized clinical trial with a large number of patients (nct ) ( table ) , certain key aspects need to be considered. first, the cost associated with ivig immunotherapy could be considered prohibitive, and second, the necessity to reserve ivig for patients whose survival essentially depends on this immunotherapy, particularly primary immunodeficient patients, must be weighed. currently, there is a worldwide shortage of ivig and the covid- pandemic will affect the collection of plasma from donors for ivig production. therefore, during this emergency period, providing ivig to those patients who depend on it should take priority over initiating an ivig trial in covid- patients. studies on covid- patients' sera confirmed the presence of seroconverted antibodies, such as igg, igm, and iga with varying kinetics and sensitivities. , findings in a monkey sars-cov- infection model confirmed that primary infection could protect the host from reinfection perhaps aided by humoral immune responses. therefore, passive immunization of sars-cov- by using convalescent plasma, intravenous hyperimmune globulin (containing high concentrations of neutralizing antibodies obtained from the pooled plasma of large number of recovered patients) or neutralizing monoclonal antibodies represent another potential therapeutic option. a meta-analysis reported a significantly reduced mortality rate in sars-cov-infected patients following infusion of convalescent plasma. these patients recorded a rapid decline in their virus load. although not conclusive, a report with mers-cov also suggested that the use of convalescent plasma could be an option for covid- . similarly, a meta-analysis of patients with spanish influenza pneumonia concluded that influenza-convalescent human blood products reduced the death risk. hyperimmune globulin treatment in severe h n influenza within five days of symptoms also reduced viral load and mortality in patients. represent an additional possibility. , it is important to validate the neutralizing activity of aforementioned plasma or antibody preparations before using with covid- patients; particularly in the case of convalescent plasma as neutralizing activity might significantly differ in mildly vs. severely ill patients. because, neutralizing monoclonal antibodies or hyperimmune globulins are pre-validated for their neutralizing activity, they do not pose such a dilemma. it is essential to note that the median seroconversion time for igm and igg was day and day respectively following onset of covid- . , at day- following onset of disease, the presence of antibodies was . % in case of igm and . % for igg. therefore, convalescent plasma and hyperimmune globulins should be collected at least three-weeks post sars-cov- infection to increase the chances of obtaining high titer neutralizing antibodies. a preliminary report from china suggested that convalescent plasma therapy leads to improvement in of covid- patients treated. an uncontrolled trial on transfusion of ml of convalescent plasma (with viral neutralizing titers more than : ) in five patients critically ill with covid- led to resolution of pulmonary lesions, and reduction in the severity of the disease and viral loads. similarly, treatment of ten severely ill covid- patients with ml of convalescent plasma containing viral neutralizing antibody titers more than : (a dilution of plasma that neutralized tcid ( % tissue-culture-infective dose) of sars-cov- ) led to reduced crp levels, undetectable viremia and improved clinical symptoms. several randomized phase and clinical trials are already planned with convalescent plasma or hyper immunoglobulin in many centers ( table ) and the plasma fractionation industry has offered support for such efforts. , meplazumab, a humanized anti-cd igg monoclonal antibody is under evaluation for covid- pneumonia. this antibody therapy was aimed at preventing cd -mediated sars-cov- viral entry and t cell chemotaxis. preliminary data in a small number of patients indicated that meplazumab improved clinical course of the disease, and normalized the peripheral lymphocyte count and crp level. tregs are classically known to suppress anti-microbial protective immunity by affecting the activation of both innate and adaptive immune cells. however, tregs are also important for preventing inflammation-associated tissue damage in acute infections. il- -based therapies to expand tregs: il- complexed to a monoclonal antibody that recognizes the cd /γc (dimeric il- receptor) epitope of il- was constructed in order to selectively activate tregs that constitutively express trimeric il- r (dimeric il- and cd ). this monoclonal antibody-il- complex displayed promising results in experimental models of several autoimmune and inflammatory diseases. recently, a human anti-il- antibody named f . has been designed that potentiates tregs by a structure-based mechanism wherein this antibody stabilizes il- in a conformation that results in the preferential stat phosphorylation of tregs. though not yet tested in humans, such approaches could boost treg number in covid- patients and reduce inflammation. on the other hand, low dose-il- therapy has been explored in the clinic for several autoimmune diseases to expand tregs. but due to the fact that covid- patients are reported to have high il- in the blood, this option might not be useful. our opinion is that neutralization of cytokines or immunomodulatory therapies are preferred over treg expansion strategies in view of the fact that due to cytokine storm, tregs might not be fully functional unless inflammatory cytokines are nullified. currently, cellular therapies have not been given much consideration for covid- , possibly due to uncertainties of such approaches, time and economic reasons. nevertheless, because of their anti-viral effector functions, several nk cell-based approaches are under consideration. little is known about the efficacy of nk cell therapy in infectious diseases. conventional treatment in combination with nk cells (twice a week, . - x nk cells/kg body weight) is under clinical evaluation (nct ) ( table ) . however, the source of nk cells for the therapy is not precise in this trial. recently, fda has given permission to celularity inc, to investigate an universal nk cell therapy cynk- in covid- patients. cynk- contains human placental cd + stem cell-derived nk cells and are culture-expanded. an innovative nk cell-based therapy with multipronged actions called nkg d-ace car-nk cell therapy is currently recruiting patients (nct ) ( table ) . nkg d-ace car-nk cells secrete il- (to ensure long-term survival of nk cells) and gm-csf neutralizing scfv (to prevent crs). in addition to clearance of virus-infected cells through ace , these nk cells can competitively inhibit sars-cov- infection of susceptible cells including alveolar epithelial cells. although the strategy looks rational, toxicity due to clearance of virus-infected lung cells, should be closely monitored. in addition to nk cell therapy, other cell-based therapies like sars-cov- -specific t cell therapies are also on the horizon. many anti-viral drugs such as protease inhibitors (camostat mesilate, lopinavir/ritonavir, and darunavir/cobicistat), nucleoside analogues (remdesivir, ribavirin), neuraminidase inhibitors lopinavir/ritonavir, and remdesivir were also found to be effective for mers-cov and sars-cov. ribavirin however, did not show benefits for mers-cov and sars-cov and was associated with severe side effects like hemolytic anemia and liver dysfunction. , the window of treatment is critical with anti-viral drugs, and they need to be used early in a disease. late initiation of therapy with anti-viral drugs, particularly after - days post appearance of clinical symptoms, might lead to a poor outcome. though the study design and results were controversial, data from a small number of patients suggested utility of hydroxychloroquine for covid- treatment. the follow-up study by the same group in a cohort of covid- patients demonstrated that a hydroxychloroquine and azithromycin combination when given during the early phase of the disease, prevents aggravation of the disease and viral persistence. further details of this study are awaited. a randomized trial also observed improved pneumonia in hydroxychloroquine-treated patients. chloroquine and hydroxychloroquine possess an unique ability to increase the ph of lysosomes and hence prevent sars-cov- viral fusion and replication. in vitro studies on chloroquine suggested anti-sars-cov- effects of these molecules. , interestingly, previous studies have demonstrated anti-inflammatory activity of chloroquine for the treatment of autoimmune diseases. but other clinical trials did not support the use of hydroxychloroquine, particularly in hospitalized covid- patients with hypoxic pneumonia or severe covid- cases. , therefore, it appears that any efficacy hydroxychloroquine might have requires that it be given during the early phase of covid- before progression to more severe disease. however, due to side effects, particularly cardiac, it is recommended to use chloroquine and hydroxychloroquine under strict medical surveillance. more evidence from randomized clinical trials is required the immediate surge in covid- positive cases prompted clinicians all around the world to explore a range of therapeutic interventions for severely ill covd- patients. due to the lack of specific therapies and time required to develop potent vaccines for sars-cov- , current investigations are focused on available anti-viral drugs. as inflammation is the hallmark of sars-cov- infections, immunotherapies that target various players of inflammation are currently being investigated. we propose that a combination of antiviral drugs and anti-inflammatory immunotherapies during early phase of the disease would function in a synergistic manner and provide maximum benefits. anti-viral drugs would reduce the viral load and hence lessen the stimuli for inflammatory responses, while immunotherapies would correct the dysregulated inflammatory response and prevent lung damage. care must be taken regarding an appropriate window of treatment and selecting the dose of anti-inflammatory immunotherapies as excessive suppression of inflammation might reduce the capacity of the patient to mount anti-viral responses. data from randomized trials are urgently needed to confirm the utility of the aforementioned immunotherapies in the management of critically ill covid- patients. the majority of the articles referred to in this manuscript are unpublished and have not yet undergone the peer-review process. they were obtained from the various preprint servers. an overreactive immune response, cytokine storm and development of acute respiratory distress syndrome characterize severe cases of covid- . bonam et al. present a perspective on diverse immunotherapeutic approaches based on the neutralization of inflammatory cytokines, immunomodulation and passive viral neutralization for treating severely ill covid- patients. • inflammation is the hallmark of sars-cov- infection • cytokine storm characterizes severely ill covid- cases • immunotherapies represent promising options for the management of severe covid- • immunotherapies either neutralize cytokines, sars-cov- or exert immunomodulation clinical features of patients infected with novel coronavirus in wuhan clinical characteristics of coronavirus disease in china a pneumonia outbreak associated with a new coronavirus of probable bat origin baseline characteristics and outcomes of patients infected with sars-cov- admitted to icus of the lombardy region incidence, clinical characteristics and prognostic factor of patients with covid- : a systematic review and meta-analysis. medrxiv high prevalence of obesity in severe acute respiratory syndrome coronavirus- (sars-cov- ) requiring invasive mechanical ventilation a trial of lopinavir-ritonavir in adults hospitalized with severe covid- chloroquine and 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critically ill patients with covid- . medrxiv sars: systematic review of treatment effects corticosteroid therapy for critically ill patients with middle east respiratory syndrome clinical management of severe acute respiratory infection when covid- is suspected. department of communications, who team brigham and women's hospital covid- clinical guidelines. brigham and women's hospital nonsteroidal anti-inflammatory drugs (nsaids) and serious infectious complications -information point therapeutic options for middle east respiratory syndrome coronavirus (mers-cov) -possible lessons from a systematic review of sars-cov therapy the cytokine storm of severe influenza and development of immunomodulatory therapy cytokine release syndrome tocilizumab, an anti-il receptor antibody, to treat covid- -related respiratory failure: a case report effective treatment of severe covid- patients with tocilizumab the impact of soluble tumor necrosis factor receptor etanercept on the treatment of idiopathic pneumonia syndrome after allogeneic hematopoietic stem cell transplantation tnf-receptor inhibitor therapy for the treatment of children with idiopathic pneumonia syndrome. a joint pediatric blood and marrow transplant consortium and children's oncology group study (asct ) altasciences completes phase i study on gimsilumab for ards in covid- . altasciences roivant announces development of anti-gm-csf monoclonal antibody to prevent and treat acute respiratory distress syndrome (ards) in patients with covid- baricitinib as potential treatment for -ncov acute respiratory disease th responses in cytokine storm of covid- : an emerging target of jak inhibitor fedratinib intravenous immunoglobulin therapy in rheumatic diseases update on the use of immunoglobulin in human disease: a review of evidence high-dose intravenous immunoglobulin as a therapeutic option for deteriorating patients with coronavirus disease clinical efficacy of intravenous immunoglobulin therapy in critical 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regulatory t cells by a structure-based mechanism revisiting il- : biology and therapeutic prospects treating human autoimmunity: current practice and future prospects fda accepts ind for nk cell therapy cynk- to treat patients with covid- . cancernetwork home of the journal oncology early treatment of covid- patients with hydroxychloroquine and azithromycin efficacy of hydroxychloroquine in patients with covid- : results of a randomized clinical trial. medrxiv remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus ( -ncov) in vitro hydroxychloroquine, a less toxic derivative of chloroquine, is effective in inhibiting sars-cov- infection in vitro no evidence of clinical efficacy of hydroxychloroquine in patients hospitalized for covid- infection with oxygen requirement: results of a study using routinely collected data to emulate a target trial. medrxiv no evidence of rapid antiviral clearance or clinical benefit with the combination of hydroxychloroquine and azithromycin in patients with severe covid- infection we thank three anonymous reviewers for their suggestions. supported by the institut national de the authors declare no competing interests key: cord- -ye dtna authors: garibaldi, b. t.; fiksel, j.; muschelli, j.; robinson, m. l.; rouhizadeh, m.; nagy, p.; gray, j. h.; malapati, h.; ghobadi-krueger, m.; niessen, t. m.; kim, b. s.; hill, p. m.; ahmed, m. s.; dobkin, e. d.; blanding, r.; abele, j.; woods, b.; harkness, k.; thiemann, d. r.; bowring, m. g.; shah, a. b.; wang, m. c.; bandeen-roche, k.; rosen, a.; zeger, s. l.; gupta, a. title: patient trajectories and risk factors for severe outcomes among persons hospitalized for covid- in the maryland/dc region date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: ye dtna background: risk factors for poor outcomes from covid- are emerging among us cohorts, but patient trajectories during hospitalization ranging from mild-moderate, severe, and death and the factors associated with these outcomes have been underexplored. methods: we performed a cohort analysis of consecutive covid- hospital admissions at johns hopkins hospitals in the baltimore/dc area between march and april , . disease severity and outcomes were classified using the who covid- disease severity ordinal scale. cox proportional-hazards regressions were performed to assess relationships between demographics, clinical features and progression to severe disease or death. results: covid- patients were hospitalized; ( . %) were discharged, ( . %) died, and ( . %) remained hospitalized. among those discharged, ( %) had mild/moderate and ( %) had severe illness. mortality was statistically significantly associated with increasing age per years (adjusted hazard ratio (ahr) . ; %ci . - . ), nursing home residence (ahr . , %ci . - . ), charlson comorbidity index ( . ; % ci . - . ), respiratory rate (ahr . ; %ci . - . ), d-dimer greater than mg/dl (ahr . ; % . - . ), and detectable troponin (ahr . ; %ci . - . ). in patients under , only male sex (ahr . ; %ci . - . ), increasing body mass index (bmi) (ahr . . - . ), charlson score (ahr . ; . - . ) and respiratory rate (ahr . ; %ci . - . ) were associated with severe illness or death. conclusions: a combination of demographic and clinical features on admission is strongly associated with progression to severe disease or death in a us cohort of covid- patients. younger patients have distinct risk factors for poor outcomes. the first case of sars-cov- in the united states was identified january th , in a returned traveler from wuhan, china. the us accounted for nearly a third of the world's cases ( , , ) and deaths ( , ) as of may , . after infection with sars-cov- , outcomes range from asymptomatic or mild illness to more severe illness and death. , age, sex, smoking, race, body mass index (bmi), and comorbidities such as hypertension and diabetes are important risk factors for severe outcomes, though to varying degrees. elevated inflammatory markers and lymphopenia are also associated with severe outcomes in covid- (the syndrome caused by sars-cov- ). [ ] [ ] [ ] while older age is one of the most important risk factors for hospitalization and death, it is increasingly recognized that younger persons may develop severe disease. the primary data source was jh-crown: the covid- pmap registry, which utilizes the hopkins precision medicine analytics platform. data in jh-crown include demographics, laboratory results, vital signs, respiratory events, medication administration, medical history, comorbid conditions, imaging, electrocardiogram results, and symptoms. our primary outcome was severe disease (including death), as defined by the who covid- disease severity scale. this is an -point ordinal scale ranging from ambulatory ( =asymptomatic, =mild limitation in activity), to hospitalized with mild-moderate disease ( =room air, =nasal cannula or facemask oxygen), hospitalized with severe disease ( =high flow nasal canula (hfnc) or non-invasive positive pressure ventilation (nippv), =intubation and mechanical ventilation, =intubation and mechanical ventilation and other signs of organ failure (hemodialysis, vasopressors, extracorporeal membrane oxygenation (ecmo)), and =death. peak covid- severity score is reported as the maximum score during the observation period for individual patients. multi-comorbidity burden was assessed using the charlson comorbidity index (cci). diagnosis of covid- was defined as detection of sars-cov- using any nucleic acid test with an emergency use authorization from the us food and drug administration. samples predominantly included nasopharyngeal swabs and less commonly oropharyngeal swabs or bronchoalveolar lavage. selection and frequency of other laboratory testing were determined by treating physicians. natural language processing was used to identify presenting symptoms as described in the supplemental appendix (table s ). we estimated the cumulative incidence functions of death using the aalen-johansen estimator (cite), with discharge and death as competing risks. to assess the association between patient characteristics and outcomes, a set of demographic and clinical variables were selected based on clinical interest and knowledge. missing values were imputed using multiple imputation by chained equations (mice) with predictive mean matching, as implemented in the mice r package (version . . ) , with rounds of multiple imputation. cox proportional-hazard models were used to relate the risk of (i) dying and (ii) developing severe disease or dying to baseline patient characteristics. four patients were discharged and then died. we censored their outcomes at time of discharge to minimize bias from lacking knowledge of deaths outside of the hopkins system. we excluded the three pediatric patients (age < years) from the models. % of deaths occurred in patients over the age of , and almost half of those patients had a "do not resuscitate/no not intubate" (dnr/dni) order within hours of admission. we used a cox proportionalhazards model to relate the risk of severe illness or death to baseline characteristics in patients under the age of in order to capture the risks of severe illness in that population. models were initially built adding variables in categorized "blocks" (e.g. "demographic") to protect against overfitting. for the composite outcome of severe disease or death, findings were equivalent to those from a model including all covariates. for other models, further variable selection was warranted when including multiple covariate blocks simultaneously. here, we fit cause-specific proportional-hazards models regularized with an elastic net penalty, as implemented in the glmnet r package (version . . ). the elastic net model was run on each of the imputed datasets, and variables with non-zero coefficients in at least half of the models were chosen for the final model, which was again run on each of the imputed datasets. demographic variables were forced to be in the models. no variable selection was done for the time to composite severe outcome or death model, as there were a sufficient number of events to allow for a larger model. standard error estimates were computed using rubin's rules (rubin, ) , as implemented in the mice r package. all analysis was done using r version . . . a total of adult and pediatric patients were admitted with confirmed sars-cov- infection from march to april (figure a) as shown in table , figure and figure , several characteristics distinguished peak illness states. increasing age (ahr . per -year increase; % ci . - . ), admission from a nursing home (ahr . ; % ci . - . ) and increasing cci (ahr . ; %ci . - . ) were independently associated with death ( table ) . a sensitivity landmark analysis excluding those who achieved an outcome within hours of admission is shown in table s . most associations were similar to the primary analysis but magnitudes of associations with respiratory and constitutional symptoms weakened. in a sub-group analysis of patients < years of age, we identified male sex (ahr . ; %ci . - . ), bmi (ahr . per -unit increase; %ci . - . ), cci (ahr . ; %ci . - . ) and respiratory rate (ahr . per increase of over ; %ci . - . ) as significantly associated with severe illness or death ( table , table s ). our study provides valuable insight into the disease trajectories of hospitalized covid- patients in the us and the risk factors associated with severe outcomes. patients who developed severe illness and survived had a median length of stay of days with % having a day stay or longer. we observed an overall mortality of %, with nearly half of all deaths occurring among nursing home residents, many of whom had dni/dnr orders on admission. although > % of patients were non-white, we did not observe statistically significant race/ethnicity associations. obesity and overall comorbidities were significantly associated with severe illness or death, particularly in persons younger than years. lastly, we found a few simple-to-measure markers such as respiratory rate, d-dimer, and troponin to be strongly associated with death. knowledge about disease trajectory and outcome is critical as providers, health systems and public health agencies plan for potentially scarce resources such as ventilators and therapeutics. , it is also important to understand disease trajectory to determine appropriate levels of care, and when discussing goals of care with patients and families. increasing age was strongly associated with death; each decade increase had a % increased hazards of mortality. similar age associations are now well described. , nursing home residents had a . fold increased hazards of death independent of age or comorbidity score, illustrating the vulnerability of this population to sars-cov- . there are approximately . million nursing home residents in the us. it is estimated that one-third of us covid- deaths are among this population. we found that % of deaths occurred in nursing home residents, similar to the % of deaths in this population that have been reported in maryland. many of our older patients, including nursing home residents, had advanced medical directives and were dnr/dni. this clearly impacted the level of intervention, measurement of severity (e.g. lack of mechanical ventilation) and time to death. the implementation of advanced directives varies substantially globally as does the age of the population. some of the global differences in sars cov- mortality are likely due to such differences. persons under years of age comprised only % of deaths ( cases) but accounted for the majority of severe illness outcomes among those who were discharged ( cases; %). more than half of those with severe illness were obese, > % were non-white, and % were male. we found that increasing bmi, cci and male sex but not race/ethnicity were strongly associated with severe disease in those < years. the ageadjusted prevalence of obesity in the adult us population is . %. obesity prevalence is higher among african americans and hispanics and linked to socioeconomic status, other comorbidities, and poor health outcomes. it is not surprising that that there is strong association between high bmi and severe covid- in the us, particularly in younger age groups who are more likely to be obese. the association between obesity and poor covid- outcomes has also been reported internationally , but the causal link remains unknown. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . mechanics of breathing may be impaired in obesity. inflammation caused by excessive fat cells might worsen the hyperinflammatory response seen in covid- . future studies of covid- interventions should include this vulnerable population. we found that a few simple-to-measure, baseline laboratory markers, namely absolute lymphocyte count (alc), albumin, d-dimer and troponin, were associated with progression to severe disease or death. lymphopenia is highly prevalent in covid- , but its impact on mortality has been inconsistent across cohorts. in our cohort lymphopenia was associated with increased illness severity or death, but not death alone. low albumin was also associated with severe disease or death, a finding that has been previously observed. both elevated d-dimer and troponin were associated with a nearly -fold increased risk of death. an elevated d-dimer is associated with increased risk of death in covid- patients independent of documented thromboembolic disease, but could also indicate an increased risk of thrombosis. an elevated troponin was also an important factor in models of severe disease or death. whether sars-cov- leads to direct or indirect cardiac toxicity, it is clear that there is a link between cardiac injury and severe outcomes in covid- . the presence of respiratory symptoms was associated with severe disease while the presence of constitutional symptoms seemed to be protective. this suggests that there are distinct phenotypes in covid- that confer differential risk. magnitudes of associations with respiratory and constitutional symptoms weakened in our landmark analysis potentially highlighting these symptoms as strong indicators of disease severity upon admission. lastly, we found that an elevated respiratory rate was associated with severe outcomes. this likely reflects that severity of illness in covid- is tightly linked to pulmonary complications including acute respiratory distress syndrome (ards) and thromboembolism. respiratory rate is included in mortality prediction scores for hospitalized patients , , and has been associated with mortality in covid- . the fact that such an easily measured parameter associates more strongly with severe outcome than several all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint inflammatory markers (e.g. crp, ferritin) suggests that inexpensive and immediately available metrics can provide valuable information about disease trajectory. there are some limitations to our study. ten percent of the patients in our cohort did not yet have an observed outcome and such incompleteness could lead to bias. however, since we adopted time-to-event approaches which handled censored survival data, our analyses remain unbiased and not affected by incompleteness. our data are derived from a single health system and may not be representative of covid- populations across the us. care practices may have differed between our hospitals. we may have under ascertained the number of covid- positive cases in our health system due to testing challenges. we may not have captured all comorbidities since some patients may not have had robust documentation in the electronic health record. post-discharge outcomes are not currently captured if they occur outside of the health system. lastly, we had to impute a considerable percentage of missing values in several laboratory tests as there is no clear standard of care for laboratory testing in covid- . in conclusion, we identified several important demographic and simple to assess factors associated with severe covid- outcomes including age, nursing home status, bmi, d-dimer, troponin, alc and respiratory rate. we also identified specific subgroups with a higher risk of disease progression including the elderly, nursing home residents, and younger patients with obesity. the data utilized for this publication were part of the jh-crown: the covid pmap registry, which is based on the contribution of many patients and clinicians. jh-crown received funding from hopkins inhealth, the johns hopkins precision medicine program. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint d. patient trajectories among those who died. this figure illustrates the trajectory of each patient who died. each horizontal line represents a single patient. the black diamonds represent the day that a code status including either a do not resuscitate (dnr) or do not intubate (dni) was entered into the medical record. fiftyfive patients ( %) had a dnr/dni order placed within hours of admission (n= deaths). all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint figure . cumulative incidence curves for death for key patient characteristics. this figure shows the cumulative incidence curves for death for key patient characteristics. in multivariate analyses increasing age (a), increasing bmi (c), charlson comorbidity index (d), increasing respiratory rate greater (e), a d-dimer greater than mg/l (g) and a detectable troponin (h) and were significantly associated with death. decreasing absolute lymphocyte count (f) was significantly associated with progression to severe disease or death in a separate multivariate analysis. male sex (b) was associated with severe disease or death in patients under the age of . all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. who maximum disease state grouped by bmi for patients under (n= ). all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint in order to identify symptoms at presentation, we first created a meta-lexicon of four symptom categories (organized into sub-categories) based on the guidelines provided by the cdc, who, and clinical findings. for each symptom category, we generated a set of synonym terms using the unified medical language system (umls) metathesaurus, and we iteratively worked with domain experts to revise the symptom categories and synonyms. table s includes the list of symptom categories and the search terms. we then selected relevant clinical note types for each patient, including h&p, critical care notes, progress notes, and ed notes, focusing specifically on the notes created within hours before and after admission. next, we pre-processed the note text and extracted only the relevant narrative parts, particularly the chief complaint and history of the present illness sections. we then used a covid- -customized version of medtagger, together with our in-house python tools to (a) identify phrases and synonyms of particular symptoms within the text narratives, (b) determine if these symptom mentions are negated, possible, or positive in their context, (c) classify symptoms into the predefined categories, and (d) map them to their corresponding umls concept unique identifiers (cuis). these nlp pipelines use a combination of machine learning models, including conditional random fields (crfs), and contextual rulebased methods, including regular expressions. finally, we selected only the positive symptom mentions in the notes and aggregated all presenting symptoms for each patient. to evaluate the performance of our nlp methods, two abstractors manually reviewed over notes from randomly selected patients. for each patient, each symptom was labeled as present or not-present (same label set as the nlp output), resulting in manually labeled symptoms with the inter-rater agreement of %. the % disagreements were individually adjudicated between the two abstractors. comparing the created gold standard to the labels generated by the nlp methods, we found that we could achieve the following results: (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. muscular/body aches body ache(s), general muscular aches and pains, generalized (acute) body aches, generalized chronic body aches, muscle aches, muscle discomfort, muscle pains, muscular aches, muscular discomfort, muscular pains, myalgia(s), myalgic, myodynia sore throat painful swallowing, difficulty swallowing, pharyngeal discomfort, pharyngeal pain, pharynx discomfort, pharynx pain, sore throat, throat discomfort, throat pain, throat soreness all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. ( %) ( %) ( %) ( %) oseltamivir ( %) ( %) ( %) ( %) all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . remdesivir a ( %) ( %) ( %) ( %) ritonavir ( %) ( %) ( %) ( %) rituximab ( %) ( %) ( %) ( %) statins ( %) ( %) ( %) ( %) tocilizumab ( %) ( %) ( %) ( %) a patients were enrolled in a randomized trial. b heparin and enoxaparin does not distinguish between prophylactic or therapeutic dosing. all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint . )* †baseline physiologic features and lab findings include the first recorded value assessed within the first hours of admission. missing values were imputed using multiple imputation prior to model creation. *p < . all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint ) a mild to moderate include patients with who ordinal score of (not on oxygen) and (on nasal cannula or face-mask oxygen). b severe includes patients with who ordinal score of (high-flow nasal cannula or non-invasive positive pressure ventilation), (intubation and mechanical ventilation), and (intubated, mechanical ventilation and other signs of organ failure including ecmo, hemodialysis or vasopressors). c data was missing for race and ethnicity data in patients, alcohol use in patients, smoking history in patients, and bmi for patients. d data was missing for admission vital signs for patients, temperature for patients, positive pressure ventilation use for patients, and sao /fio for patients. e data was missing for wbc for patient, alc for patients, hemoglobin for patient, platelet count for patients, albumin for patients, alt for patients, ast for patients, bilirubin for patients, creatinine for patients, gfr for patients, crp for patients, procalcitonin for patients, ldh for patients, d-dimer for patients, fibrinogen for patients, ferritin for patients, il- for patients, hemoglobin a c for patients, troponin for patients, and pro-bnp for patients. references clinical characteristics of hospitalized patients with novel coronavirus-infected pneumonia in wuhan, china presenting characteristics, comorbidities, and outcomes among patients hospitalized with covid- in the case-fatality rate and characteristics of patients dying in relation to covid- in italy risk factors associated with acute respiratory distress syndrome and death in patients with coronavirus disease hospitalization rates and characteristics of patients hospitalized with laboratory-confirmed coronavirus disease -covid-net, states validation of a combined comorbidity index an empirical transition matrix for non-homogeneous markov chains based on censored observations multivariate imputation by chained equations in r imputing missing covariate values for the cox model regression models and life-tables regularization paths for generalized linear models via coordinate descent how should variable selection be performed with multiply imputed data multiple imputation for nonresponse in surveys the r project for statistical computing. r foundation for statistical computing, emergency use authorization of remdesivir: the need for a transparent distribution process fair allocation of scarce medical resources in the time of covid- clinical course and risk factors for mortality of adult inpatients with covid china: a retrospective cohort study development and validation of a clinical risk score to predict the occurrence of critical illness in hospitalized patients with covid- one-third of all u.s. coronavirus deaths are nursing home residents or workers. the new york times do-not-resuscitate decisions in six european countries prevalence of obesity and severe obesity among adults: united states obesity could shift severe covid- disease to younger ages association of higher body mass index (bmi) with severe coronavirus disease (covid- ) in younger patients high prevalence of obesity in severe acute respiratory syndrome coronavirus- (sars-cov- ) requiring invasive mechanical ventilation hypoalbuminemia predicts the outcome of covid- independent of age and co-morbidity high risk of thrombosis in patients with severe sars-cov- infection: a multicenter prospective cohort study association of cardiac injury with mortality in hospitalized patients with covid- in a severity of disease classification system the sofa (sepsis-related organ failure assessment) score to describe organ dysfunction/failure. on behalf of the working group on sepsis-related problems of the european society of intensive care medicine predictors of mortality for patients with covid- pneumonia caused by sars-cov- : a prospective cohort study table s : association of clinical characteristics and severe outcomes, age < , demographics not forced into model clinical characteristic † severe outcome or death the unified medical language system (umls): integrating biomedical terminology desiderata for delivering nlp to accelerate healthcare ai advancement and a mayo clinic nlp-as-a-service implementation all rights reserved. no reuse allowed without permission. (which was not certified by peer review) is the author/funder key: cord- -ig j z authors: couetil, laurent; cardwell, jacqueline m.; leguillette, renaud; mazan, melissa; richard, eric; bienzle, dorothee; bullone, michela; gerber, vinzenz; ivester, kathleen; lavoie, jean-pierre; martin, james; moran, gabriel; niedźwiedź, artur; pusterla, nicola; swiderski, cyprianna title: equine asthma: current understanding and future directions date: - - journal: front vet sci doi: . /fvets. . sha: doc_id: cord_uid: ig j z the havemeyer workshop brought together researchers and clinicians to discuss the latest information on equine asthma and provide future research directions. current clinical and molecular asthma phenotypes and endotypes in humans were discussed and compared to asthma phenotypes in horses. the role of infectious and non-infectious causes of equine asthma, genetic factors and proposed disease pathophysiology were reviewed. diagnostic limitations were evident by the limited number of tests and biomarkers available to field practitioners. the participants emphasized the need for more accessible, standardized diagnostics that would help identify specific phenotypes and endotypes in order to create more targeted treatments or management strategies. one important outcome of the workshop was the creation of the equine asthma group that will facilitate communication between veterinary practice and research communities through published and easily accessible guidelines and foster research collaboration. the effort to clarify the phenotype and terminology used to characterize horses with chronic inflammatory airway disease started in with a workshop in east lansing, michigan ( ) . several workshops were subsequently held with similar goals in mind with the latest hosted in cabourg, france in ( ) . in the last few years, the terminology has further evolved with the term equine asthma (ea) now being recommended to describe horses with chronic respiratory signs ranging in severity from mild to severe that were previously referred as inflammatory airway disease and recurrent airway obstruction, respectively ( ) . although strong evidence supports the role of exposure to environmental dust in the pathophysiology of both mild and severe ea, the potential role of infectious agents (bacterial and viral) has not been clearly established. the goal of the havemeyer workshop on equine asthma was to bring together researchers and clinicians from different disciplines who are actively investigating airway inflammation to discuss the latest information on this topic and provide some comparative perspective from human asthma. the workshop was designed to facilitate productive discussions that would inform potential future revisions of the american college of veterinary internal medicine (acvim) consensus statement on mild-moderate ea ( ) and provide future research directions. the present report follows the format of the workshop. the manuscript is organized thematically starting with the recent advancements in the understanding of the classification and diagnosis of human and equine asthma. the second part is centered on the etiology and pathophysiology of ea. the third and final section of the manuscript summarizes the extensive discussions conducted during the workshop with the goal of prioritizing future directions of ea research. clinical asthma phenotypes have been recognized for many decades but were collapsed into a unified hypothesis of asthma as an allergic disease in when age adjusted levels of immunoglobulin e were associated with asthma. it has taken more than years to consider the heterogeneity of asthma again with an emerging emphasis on endotypes, an intrinsically more interesting approach to understanding asthma pathobiology ( ) . the term "endotype" is used to describe a subtype of disease defined by a molecular mechanism, genetic variation or by treatment response ( , ) . cluster analyses of asthma cohorts have revealed groups with different ages of onset, lung function, concordance or lack thereof between measures of airway inflammation by sputum analysis and symptoms. a recent review of asthma by a panel of experts has focused on the need to recognize asthma in its diverse forms and to identify treatable traits. this extensive review has highlighted areas for future attention ( ) . the application of the analysis of gene expression to airway epithelial cells and sputum cells from well-characterized groups of asthmatics has led to the appreciation of asthma associated with t helper cytokines and non-t asthma ( ) . the former is the more allergic subset with higher ige and peripheral and sputum eosinophilia. non-t asthma has fewer of these features and is less responsive to inhaled corticosteroids. t cells that express interleukin- have been linked to severe neutrophilic asthma. these so-called th cells have been shown in animal models to be associated with steroid-unresponsiveness. the th cytokine interferon-γ likewise has been found to be expressed in the airways of severe asthmatics. in recent years there has emerged another lymphoid cell that participates in host responses to mucosal injury. these innate lymphoid cells are lineage negative, lacking the usual lymphocyte surface markers ( ) . they express similar panels of cytokines to the t helper subsets and are labeled innate lymphoid cell (ilc) , , and . they are rapidly activated by epithelial signals such as thymic stromal lymphopoietin (tslp), interleukins and , molecules termed alarmins. the secretion of il- and il- by ilc may lead to a pattern of inflammation previously interpreted as th . innate lymphoid cells are less steroid sensitive. additionally, alarmins prime cells such as dendritic cells and therefore may have a role in adaptive immunity as well as innate immune responses. the synthesis of amphiregulin, an epidermal growth factor receptor ligand, by ilc s but also th cells, is postulated to promote mucosal integrity. one could anticipate that viral infection of epithelial cells or damage by irritants giving rise to inflammation mediated by ilcs. however, their roles have yet to be fully explored. transcriptomic analysis of sputum has revealed three patterns of inflammation and gene signatures consistent with both th and ilc driven inflammation and oxidative stress ( ) . the descriptions of molecular mechanisms of inflammation may still be considered as a deeper form of phenotyping. however, the application of novel biologics to treat asthma is now implicating certain pathways in disease and therefore is providing us with true disease endotypes. most of the progress in the identification of treatable traits has related to the t phenotype. biologics targeting ige (omalizumab), il- and therefore, the eosinophil (mepolizumab, benrazilumab, rezlizumab) and the t cytokines (dupilumab) have all demonstrated efficacy in reducing exacerbations of asthma. recent results of studies targeting the alarmin tslp and therefore both t high and low asthma have confirmed efficacy against acute attacks of asthma. oxidative stress in asthma has not been specifically addressed. a problematic form of asthma is that associated with airway remodeling and fixed airway obstruction. the association with mucus plugging and eosinophilic inflammation has been recently identified as a potential factor in long term impaired airway function ( ) . severe equine asthma is typically a neutrophilic form of asthma although expression of t cytokines has been described ( ) . there is also evidence that il- is expressed in equine asthma and its effects on neutrophil survival are steroidinsensitive ( , ) . although neutrophilic human asthma is less steroid-sensitive than the eosinophilic phenotype, severe equine asthma is responsive to steroid treatment despite the presence of neutrophilic inflammation. severe equine asthma shares the structural remodeling of the airways with human asthma and a part of the remodeling change is reversible with steroid treatment as well as withdrawal from the inciting stimulus ( ) . studies of airway remodeling in human asthma with treatment have not addressed key components of remodeling such as increased airway smooth muscle mass. revised in and discussed the use of ea to describe these conditions ( ) . the revised consensus recognized that asthmatic horses of all severities have common clinical presentations (such as chronic cough, excess mucus, poor performance) but also a wide heterogeneity in terms of triggering factors, severity, and pathologic characteristics. a phenotype is the observable physical properties of an organism, including measurable laboratory findings, which is the result of the expression of the genes in response to the environment ( ) . identifying distinct phenotypes is of interest if they facilitate the diagnosis, the prognosis or allow the implementation of targeted therapy. while currently loosely defined, the ea phenotypes discussed in the consensus statements were based on clinical presentation (severe vs. mild/moderate), triggering factors (barn/hay or pasture), endoscopy findings (mucus) and bronchoalveolar cytology. from a clinical standpoint, further dividing ea as distinct "mild" and "moderate" phenotypes may promote recognition that asthma is an underdiagnosed cause of exercise intolerance in high performance horses. horses with a cough or increased respiratory rate at rest or following exercise will commonly undergo further diagnostic procedures to confirm asthma, or "anti-asthma" treatments will be implemented. however, this is generally not the case when no clinical signs suggestive of an airway disease are present. the term "mild ea" could describe the condition affecting these horses, while "moderate ea" would be used when clinical signs of airway disease (such as cough) are present, but without the periods of labored breathing at rest seen in "severe ea" ( ) . the inflammatory airway cell phenotypes (neutrophils, mast cells, eosinophils) were recognized in the and consensus statements ( , ) . future phenotypes may include the age (early or late) of appearance of clinical signs, or specific remodeling features affecting the airways, if these new features are shown to facilitate prognostication or the implementation of specific therapy. the future development of new portable and sensitive devices for measuring the lung function of horses (forced oscillation or flow interruption techniques), or the discovery of blood biomarkers for ea would help not only to facilitate the diagnosis of mild and moderate forms of ea in clinical practice, but also to possibly identify new phenotypes for these conditions. to date, different inflammatory pathways have been proposed as contributing to ea, which may eventually lead to novel therapies ( ) . the discrepancies between results of the different studies may be an indication of different endotypes in ea, although future studies on large cohorts of horses from multiple sites would be required before specific endotypes can be recognized. multicenter tissue banking could facilitate these studies. in summary, the acvim consensus statement recognized the currently known distinctive features of ea. further defining "mild" and "moderate" ea based on the presence or absence of easily identified clinical signs may promote the investigation of the subclinical (mild) phenotype. the identification of novel phenotypes and endotypes may lead to "precision medicine" where treatments most likely to help equine patients would be selected. this approach is now implemented in humans and may eventually be applicable to horses if supported by scientific research. severe equine pasture asthma (epa) is characterized by episodes of reversible airway obstruction in horses grazing pasture during the summer in hot humid climates ( ) . affected horses demonstrate neutrophilic airway inflammation, airway hyperresponsiveness extending throughout the season of remission, and airway remodeling ( , ) . the author's experience is restricted to epa as first described in horses residing in louisiana, and diagnosed in states with subtropical climates (mississippi, alabama, and florida) ( ) . veterinarians in regions of adjoining states and distant states (oregon) describe similar signs in horses grazing pastures during hot humid conditions. epa is described in the united kingdom where it differs in its association with hot dry weather or exposure to dust from harvest/burning of crops ( ) . epa demonstrates adult onset ( ± years; range - years) without sex predilection ( ) . asthma exacerbations generally begin in summer (july), persisting until temperature and humidity decrease (october/november) ( ) . fewer horses experience asthma in the spring. a history of prior seasonal cough and/or exercise intolerance may be identified. improvement within hours to days of isolation from pasture particulates in a stall environment is a key diagnostic feature of epa in the southeastern usa ( ) ; some severe cases necessitate isolation in a climate climate-controlled environment. in the author's experience, without adequate environmental management, disease severity is progressive and responsiveness to parenteral corticosteroids decreases. though specific agent(s) that elicit epa exacerbation are not identified, the response to stall housing implicates seasonal pasture-associated particulates. costa et al. reported increases in grass but not tree pollens were significantly associated with epa exacerbation using a pollen station ∼ miles from affected horses ( ) . in this regard, intact pollen is too large to reach the respirable zone of humans in order to elicit asthma, but moist conditions that are associated with epa exacerbations can shatter pollen and disseminate respirable particles ( ) . grass pollen sensitization is classically associated with th responses, ige-mediated hypersensitivity, and eosinophilic inflammatory infiltrates. however, chronic exposure to th sensitizing antigens and to complex antigen combinations that include th sensitizing antigens each generate th responses accompanied by neutrophilic airway inflammation that typifies epa ( , ) . subtopical grasses differ substantially from grasses in temperate and continental climates ( ) . pollen from subtropical grass subfamilies is important to rhinitis and human asthma in subtropical zones of australia, asia, india, africa, and america. pollination seasons for bahia and bermuda grass (spring through september/october) align to the season of epa exacerbation ( , ) . the pollen season for johnson grass is temperature dependent, flowering from may to july, with higher temperatures moving flowering later into autumm ( ) . a role for fungal triggering in epa exacerbation is suggested by the near identical clinical picture presented by horses with epa and barn dust-associated severe asthma, wherein a role for fungal triggering is substantiated in the latter ( ) . chronic neutrophilic airway inflammation characterizing both forms of severe equine asthma also aligns to th -mediated neutrophilic inflammation in fungal asthma models ( ) . of the more than species of fungi that exist in biotropic relationships with bermuda, bahia, and johnson grasses, curvularia, helminthosporium, alternaria, puccinia, epicoccum, and fusarium are implicated in eliciting human asthma ( ) . costa et al. identified fungal spores of the genus nigrospora, and curvularia, as well as basidiospores, as temporally associated with exacerbations of pasture asthma ( ) . these findings are in agreement with reported correlations between epa exacerbation and high dew point temperature ( ) . specifically, nigrospora conidia and basidiospore release increase with increasing relative humidity, resulting in a peak in spore counts during the early morning and aligning to the association of epa exacerbations with increased dew point temperature ( , ) . in contrast, conidia of cladosporium, alternaria, epicoccum, and dreschlera spp. are released during warm, dry, windy conditions, while precipitation is required for release of many ascospores. in this way, humidity influences fungi of relevance to asthma in different locales which could influence associations of pasture asthma in the uk with hot dry conditions, rather than hot humid conditions precipitating pasture asthma in the southeastern us. as a chronic and progressive disease of undetermined etiology, epa is most effectively managed by segregation from inciting grass pastures during warm seasons. the necessity to segregate horses from pasture, particularly at a time when they are typically extensively ridden and grazed, presents a conundrum that is ultimately detrimental for most affected horses. accordingly, there is a critical need to identify the agents that trigger epa in order to improve disease management. both veterinary practitioners and researchers muse about the diagnostic armamentarium available to physicians-if only we had the chest ct, the advanced lung function testing, the biomarkers-then we would be able to have a better diagnosis. a quick search of the literature, however, shows us that our counterparts face many of the same diagnostic dilemmas that we do, albeit often with higher bills! while pulmonologists have drawn up multiple guidelines to help in the diagnosis of asthma in humans with its multiple phenotypes and endotypes, physiciandiagnosed asthma criteria often fail to be consistent with the official guidelines rendering the results of large epidemiologic studies or clinical trials fraught with the perils of resting findings on nebulous datasets. various forms of spirometry or simple pulmonary function testing are readily available in human medicine, but few non-pulmonologists avail themselves of objective data, and instead rest on reported symptoms such as difficulty breathing on exertion, cough or positive response to bronchodilation ( ) . indeed, the gina toolbox identifies "lack of access to spirometry/bronchoprovocation tests" as a barrier to implementation of gina guidelines in human asthmatics ( ) . moreover, the heterogeneity in published algorithms for diagnosis of asthma-more than in the literature at last count-make even an algorithm-based diagnosis unsure ( ) . thus, the conclusion that symptom-based diagnosis is associated with a significant risk of over-diagnosis has been reached for asthma in humans ( ) . the current push in human medicine to refine both the phenotypes and endotypes for multiple different subtypes of asthmas aims to elucidate the underlying causes and thus treatments that may be very different. we are still searching for the criteria that will help us with this in equine medicine. if there are indeed mechanistically different groups of horses within the categories of mild, moderate, and severe ea that are associated with genetic differences or cellular or molecular biomarkers, then perhaps we will gain better understanding of treatment successes and failures and will be able more logically to choose clinical therapies and predict responses. the difficult case for the clinician and the researcher alike is not the horse with severe ea-because the history and clinical exam alone can often suffice to diagnose, and there is a visible relief in respiratory embarrassment with administration of bronchodilator (although it can take some time in horses with diaphragmatic exhaustion) ( ) . the difficult horse is the one with moderate/severe asthma in remission and the horse with mild-moderate ea. as was recently pointed out, the biggest difference that we note in the clinical diagnosis of horses with mild-moderate ea vs. severe ea is the presence of an increased respiratory effort at rest, which is due to the underlying pathophysiology of bronchoconstriction, increased mucus, and bronchiolar inflammation ( ) . the need, then, is to detect the mildly or subclinically affected horse. as veterinarians, we have at hand history, clinical signs, lung function testing, radiographs, endoscopy, analysis of airway secretions, blood biomarkers and clinical pathology which can be used in a minimum database in order to classify horses into clinically useful categories that have a pathophysiologic basis that can simultaneously allow us to diagnose, treat, and translate clinical cases into field research. a tentative diagnosis of ea in its most severe form can often be made on history alone, with the key component being the recognition of episodes of reversible respiratory embarrassment precipitated by exposure to specific triggers-namely, moldy hay in the northeast of the united states, and pasture allergens and particulates in the south. history in subclinical or mild cases is seldom of such definitive use; this does not mean that it is unimportant. such questions as parentage ( ) , type of feed and how it is fed ( ) , and heat and pollen counts at the time of diagnosis ( ) may be important risk factors for equine asthma. while it has been proposed that coughing and poor performance may serve to define a phenotype of moderate vs. mild ea ( ) , these signs are not sufficiently sensitive ( ) and would misclassify a subset of horses-they alert the clinician that moderate ea is likely, but the absence of these signs does not rule out disease. the connections between ea and viral or bacterial disease are not linear, but it is becoming increasingly clear that the connection exists ( , ) , thus a thorough history should include probing for past infectious respiratory disease. one of the best described questionnaire analysis tools for classification of horses based on history is the hoarsi index ( ), developed as a means of distinguishing among normal, mild-moderate ea or severe ea phenotypes. however, clinical signs and indices are insufficiently sensitive to distinguish horses with mildmoderate ea from normal horses or horses with severe ea in remission ( ) . proposed minimum database for both practitioners in the field and for research: a common history tool should be developed that addresses the main concerns of parentage if known, current and lifetime exposures to particulates and allergens including feeds and feeding practices, barn environment, vaccinations, travel history, and recent illnesses. multiple scoring systems have been shown to be useful for distinguishing healthy horses from horses with severe ea in exacerbation, but, similar to questionnaire indices, these scoring systems do not help in the more difficult problem of distinguishing horses with mild ea from healthy or severe ea in remission ( ) . indeed, years ago, robinson et al. found that even in horses with historical severe ea, clinical score failed to reflect low-grade airway obstruction, and suggested that without easily used, field-accessible testing equipment, lower airway disease would go underdiagnosed ( ) . recently, the adapted -point scoring system has been shown to be the most useful in discriminating mild from severe cases, but it is unlikely to distinguish normal from subclinical disease ( ) , and the ideass scoring system has recently been described as a useful scoring system for moderate-to-severe equine asthma ( ) . thus, while clinical scoring is essential to a good examination and careful research, and can potentially be useful in measuring response to treatment in the individual, it is insufficient in making the phenotypic distinction between mildly affected horses and healthy horses. proposed minimum database for both practitioners in the field and for research: the -point modified clinical score appears to best stratify horses with obstruction ranging from mild to severe. an application suitable for smart phone use would enhance the adoption of a common scoring tool. in human asthma, the gold standard is the detection of variability in pulmonary function using spirometry or other methods of lung function testing ( ) . unfortunately, lung function testing remains available only to a few specialized centers, as more recently developed portable lung function testing modalities are no longer on the market ( ) . initial reports from the author's laboratory of a simple field test of respiratory resistance using the interrupter technique hold promise for increased use of lung function testing in the future. while the classic esophageal balloon/pneumotachometer method is effective in demonstrating increased maximal pleural pressure and allows for calculation of pulmonary resistance and elastance as well as dynamic compliance in severe ea, it is not sufficient for demonstrating abnormal function in mildly affected horses in which baseline lung function is rarely abnormal and histamine or other bronchoprovocation or bronchodilation must be used in order in order to detect low-grade obstruction ( ) . unfortunately, in some studies, even histamine bronchoprovocation has not been sufficient to distinguish between normal horses and horses with mild asthma ( ) , and a lack of concordance between histamine bronchoprovocation and bronchoalveolar lavage (bal) cytology has been noted in several studies ( , ) . while lung function testing and histamine bronchoprovocation have shown moderate to strong correlations with bal cytology in some studies ( , , ) , others have not ( , ) . methods of performing histamine bronchoprovocation are equally important: studies in human asthmatics have shown that it is the total dose of histamine that is most important rather than the duration of exposure. a more precise method of dosing may be important to establish. in human athletes, indirect stimuli, such as cold air, hypertonic solutions such as mannitol, exercise, and amp are all considered more accurate and useful in predicting asthma than are direct stimuli such as methacholine or histamine; this is an area that requires exploration in equine pulmonology. while hay challenge is useful for research in severe ea, it is inappropriate in a clinical case, especially in a horse that is expected to do athletic work ( ) . in moderate to severe ea, variability in airflow should be demonstrated not through bronchoprovocation but through bronchodilation using either systemic (buscopan tm ) or inhaled (albuterol, ipratropium bromide) drugs to assess reversibility; it is possible that a h period of bronchodilation is necessary for maximum effect in horses with diaphragmatic fatigue ( ) . proposed minimum database for both practitioners in the field and for research: in research, lung function should be assessed and airflow variability/changes in airway caliber should be assessed with either bronchoprovocation or bronchodilation. more research is necessary to determine if field assessment of lung function after bronchoprovocation or bronchodilation is sufficient to determine change with the -point scoring system. it is essential that a robust, easily used system for testing lung function in the field be developed. unlike in human pulmonology, examination of airway secretions is a primary method of diagnosis in ea, be it mild, moderate or severe. although a standard volume of between and ml of saline using a m long endoscope or m bal tube is recommended ( ), this practice is not always followed, and cytology should be assessed keeping in mind that the amount of fluid infused will affect the cell percentages. the relationship between bal cytology and performance is still not clear. certainly, poor performance has been associated with what have been determined to be abnormal cell types or percentages ( ) . there has been much discussion as to what is normal on bal cytology; it likely depends on a combination of technique, environment and population. even the "stringent" definition proposed by couëtil et al. ( ) of < % neutrophils, % mast cells, % eosinophils, would be considered elevated in some high-performance populations ( , ) . although an earlier study found no evidence of a clear phenotype in mast cell vs. neutrophilic inflammation with respect to pulmonary gas exchange during exercise ( ) , recently, an increase in bal mast cells or neutrophils was shown to negatively affect performance ( ) . the way that cells are counted in bal cytology is also important, especially for rare cells. in our laboratory we count a minimum of cells at x for common cells such as macrophages and lymphocytes or neutrophils in mild ea, whereas for rare cells such as mast cells we count , cells. other techniques, such as using a -field differential for mast cells, are only useful if the cell density is high ( ) . the conundrum of whether to assess airway fluid from both lungs rather than blind sampling, or to pool samples, has also occupied attention from researchers. one group found that, depending on whether the "loose" or "stringent" categorization was used, - % of horses would have been categorized as control vs. mild-moderate ea if only one lung were used ( ) . as it is the rare practitioner who has a bronchoscope in the field, it is unlikely that even pooled samples ( ) , which may be a better representation of overall lung inflammation, will be taken other than in referral centers or practices. the problem is most important for rare cells. more attention will need to be paid in future to morphology and perhaps typing of cells. the existence of neutrophil extracellular traps (netosis) in horses with severe ea presents an additional method to determine response to treatment ( ) , and recently the presence of degenerate neutrophils has been shown to raise suspicion for bacterial infection ( ) . the question of macrophage morphology as an indicator of inflammation is also an area that will profit from further investigation ( ) . recently, as well, the paucigranulocytic phenotype has been described in which horses with clear signs of severe ea have low neutrophil percentages in the bal ( ) . this is thought to be due to mucus plugging of small airways that essentially sequesters neutrophils. although a recent publication showed a rather shocking % of highperforming european horses with mild-moderate ea had fungal elements in the bal ( ) , this remains to be confirmed in other populations. proposed minimum database for both practitioners in the field and for research: for the bal, at least -mls of saline should be used, and there is a preference for counting at least cells to adequately represent rarer cells. for research purposes where rare cells are of interest (e.g., mast cells or eosinophils), sampling of both lungs appears preferable. better categorization of cells through morphological descriptions including apparent neutrophil extracellular traps and notations of fungal or birefringent elements should be done. characterization of mucus on cytology may help to elucidate the paucigranulocytic phenotype. bal in the field will usually be done blindly with a specialty tube. the debate continues to swirl around the utility of tracheal wash vs. bronchoalveolar lavage, with malikides et al. ( ) finding a % disagreement in young racehorses, while derksen et al. ( ) determining that there was no correlation between bal and tw, and others finding no relationship between tracheal neutrophil counts and racing performance ( ); thus, tracheal cytology has been considered inappropriate for diagnosis of mild ea ( ). recently, however, a comparison of tw and bal in horses, along with evidence of mucus and endoscopy, found that only . % of horses would have been classified differently if they had had the other procedure, eventually concluding that there is no gold standard-except for mast cells, which are rare in the trachea, and thus, to be found, demand that a bal be performed ( ) . proposed minimum database for both practitioners in the field and for research: tracheal wash may be most practical for some practitioners in the field and has the added benefit of allowing for bacterial culture. the inability to assess mast cells adequately continues to limit this modality. in research settings, both tracheal aspirate and bal are preferable. many clinical diagnoses are made on the basis of endoscopic visualization of mucus, with strong support from the finding that tracheal mucus quite nicely correlated with racing performance or lack thereof ( ) . the recent consensus statement considers that the demonstration through tracheobronchial endoscopy of mucus grade / in racehorses or / for sport/pleasure horses is sufficient to diagnose mild-moderate ea and in support of this recommendation, rossi et al. ( ) found that visible mucus in the trachea is indeed likely to predict inflammation. there are varying degrees of certainty about mucus in the trachea predicting inflammation ( , , , ) . nonetheless, other studies have shown that mucus is insufficient to parse out mild vs. unaffected cases ( ) . endoscopy has also been shown to be useful in detecting an increase in upper airway abnormalities in horses with mild-moderate ea, with courouce-malblanc et al. ( ) raising the chicken-and-egg question of the relationship between mild-moderate ea and dorsal displacement of the soft palate, and more recently, wysocka and klucinski ( ) found that more horses with mild-moderate ea had dynamic pharyngeal abnormalities. it may be that the answer will rest in whether any of these modalities can help to define a phenotype rather than simply further describing an already understood phenotype. proposed minimum database for both practitioners in the field and for research: upper airway endoscopy should be performed to rule out upper airway cause of obstruction as a primary cause of signs or that might confound lung function testing. assessment of tracheal mucus should be performed. endobronchial biopsies offer an excellent method of sampling larger airways, although deeper layers cannot be accessed. the brass ring-being able to distinguish normal from remission or mild ea-remains elusive, however, as correlates were evident between histopathology and impulse oscillometry and showed a difference between horses in remission at pasture and those that remained stabled and treated with glucocorticoids, but did not show any difference between horses with severe ea in remission and controls ( ) . proposed minimum database for both practitioners in the field and for research: at this time, brushings/biopsies are not considered part of a minimum database. imaging is considered an important ancillary diagnostic in humans, but radiographs have not been shown to be sensitive or specific in horses with ea ( ) . chest ct is currently not feasible in large animals. while endobronchial ultrasound shows promise for the elucidation of airway smooth muscle thickening in severe ea, the ultimate goal of being able to detect low-grade disease in erstwhile healthy horses, or to distinguish normal from severe ea in remission remains elusive ( ) . proposed minimum database for both practitioners in the field and for research: at this time, imaging is not considered part of the minimum database. equine asthma encompasses mild to severe forms of chronic airway inflammation. severe ea affects ∼ - % of horses in countries with northern, cool climate ( , ) . mild-moderate ea affects - % of pleasure horses based on tracheal wash cytology (neutrophils > %) and up to % of racehorses based on bal cytology ( , ) . horses affected with severe ea experience exacerbation of clinical signs when exposed to organic dust originating from hay and bedding, in particular molds present in poor quality hay. as a result, clinical signs tend to be worse during the winter when horses are housed indoors for extended periods of time ( ) . some horses exhibit disease flare-ups while at pasture during summer months (epa) ( ) . these horses improve clinically during winter or after being housed indoor. a small percentage of horses appear to suffer from both classic severe ea and epa. horses with severe asthma tend to be mature (> years) to old animals and a genetic predisposition has been identified in some families ( , ) . the main clinical sign characteristic of severe ea is increased respiratory effort ("dyspnea") that can rapidly improve following bronchodilator administration. although the decrease in respiratory effort following bronchodilator administration can be detected within minutes of drug administration using lung function testing, clinical improvement may not be apparent to clinicians ( ) . acute exacerbation is associated with increased pulmonary artery and right-heart vascular pressures as well as increased pulmonary artery diameter on ultrasound ( ) . blood pressure return to baseline during clinical remission however, cardiac ultrasound abnormalities such as right ventricular wall thickness remained increased ( ) . surprisingly, severe ea is rarely fatal unless complications develop such as cor pulmonale ( ) . affected horses are more likely to be euthanized because owners get discouraged with the expense associated with chronic therapy and maintaining a low-dust environment ( ) . coughing and nasal discharge are non-specific signs of respiratory disease commonly reported in horse with severe ea ( ) . horses with a history of both coughing and mucoid nasal discharge are at increased risk of developing severe ea ( ) . thoracic auscultation may reveal increased breath sounds bilaterally, extended area of auscultation, and abnormal breath sounds (i.e., crackles, wheezes). however, the thick chest wall of horses makes auscultation an insensitive indicator of pulmonary disease, with abnormal findings obtained in < % of horses with severe ea ( ) . strict management changes or medical therapy will results in rapid improvement in clinical signs however, if exposure to triggering factors is not addressed improvement will be short lived or incomplete ( , ) . this form of mild respiratory disease is mainly subclinical with horses showing non-specific signs such as intermittent coughing and poor performance ( ) . however, mild asthma should not be ruled out in horses that do not cough because coughing is reported in only % of horses with mild asthma ( ) . coughing is associated with increased bal neutrophils ( ) . poor performance and reduced willingness to perform are associated with increased tracheal mucus scores in racehorses and show-horses, respectively ( , ) . in racehorses, poor performance has been associated with increased neutrophils and mast cells in bal fluid ( ) . there is an association between nasal discharge and increased tracheal mucus in racehorses ( ) . however, the association between tracheal mucus and bal cytology has not been reported yet. the term "remodeling" defines a process resulting in a tissue that is structurally and architecturally altered compared to its healthy counterpart. in asthma, structural alterations are represented by quantitative or qualitative changes of the bronchial wall components or their surrounding tissues, whilst architectural alterations refer to the skewed relationships among such structures. airway remodeling has been studied only in horses affected by severe ea. an increased expression of metalloproteinases and their tissue inhibitors has been recently reported in a group of horses with mild respiratory signs and bal cytology compatible with mild ea ( ) . however, the possibility that the horses studied were horses with severe ea in remission of the disease was not excluded. almost all airway components undergo remodeling in severe ea, both in peripheral (diameter < mm) and central airways. the airway smooth muscle mass as well as collagen and elastic fiber deposition are increased in the lamina propria of peripheral airways during severe ea remission compared to healthy airways ( , ) . mucostasis, mucus cell hyperplasia, peribronchiolar metaplasia, and interstitial fibrosis are more frequently detected in horses with severe ea in remission compared to controls ( ) . however, histomorphometric techniques revealed no differences in the number of mucus cells per mm of lamina reticularis or in the volume of stored mucosubstance in bronchial epithelial cells ( ) . central airway remodeling during disease remission is less pronounced compared to what is observed peripherally. whether airway submucosal structures are significantly altered during severe ea remission compared to control remain to be established ( , , ) . functionally, severe ea remission is associated with a normal lung function in spite of significant structural alterations of the airways. in these conditions, the respiratory resistance correlates with the amount of collagen within the lamina propria of peripheral airways ( ) , indicating that, in the absence of bronchospasm, peripheral airway stiffness is the major determinant of respiratory resistance in asthmatic horses. the functional implications of peripheral remodeling become more important during disease exacerbations, when most of the changes are further accentuated and the mechanics of breathing are altered ( , ) . there is no doubt that the major determinant of airway obstruction during severe ea exacerbations is smooth muscle contraction and that central airways play a major role ( ) . by definition, the force produced by a muscle is proportional to its cross-sectional area. given the increased smooth muscle mass (and cross-sectional area) during severe ea exacerbations ( ) , asthmatic muscle is "stronger" and able to contract the thickened lamina propria observed in severe ea, further reducing the airway lumen. increased mucus secretions into the airway lumen also contribute to airway occlusion ( ) . these same mechanisms operate in peripheral airways, where the effects on lung function are somewhat blunted by the fact that their overall contribution to pulmonary resistance is low, due to their large cumulative cross-sectional area ( ) . at this level, the more relevant functional effects of remodeling are the loss of lung elasticity and airway-parenchymal tethering. adequate small airway patency is guaranteed by their intimal connection to the lung parenchyma by elastic and connective fibers. when the lung inflates during inspiration, small airways are stretched and passively dilate. remodeling of elastic fibers and of the extracellular matrix within and around the airways and in the alveolar septa alters this mechanism, preventing the smallest airways from remaining open ( ) . the effect is even worse during expiration, when the lungs physiologically recoil and the airway diameter physiologically narrows. with a significantly impaired expiratory airflow, part of the air that reaches the alveoli remains trapped. this leads horses with severe ea in exacerbation to breath at increasing lung volumes [functional residual capacity ( ) ] in the attempt to maintain airway patency, which causes lung hyperinflation and enlarged fields of thoracic auscultation ( ) . anecdotal evidence to date has suggested that, although bal sampling is widely accepted elsewhere as the diagnostic tool of choice for cytological assessment of equine lower airways, tracheal endoscopy and tracheal wash-based diagnostics have remained the mainstay of routine clinical lower airway investigations in british thoroughbred racehorses in training. given the emphasis on bal in research, this would present a considerable challenge to furthering evidence-based respiratory medicine in this important equine population. in a recent study we investigated british racing veterinarians' rationales for current practices, and the challenges they face in relation to diagnosing and managing racehorse airway inflammation ( ) . qualitative data were gathered through semi-structured focus group discussions designed to capture current practices and opinions relating to the diagnosis and treatment of lower airway inflammation, as well as familiarity with and views on the most recent acvim consensus statement ( ), in which the term "mild-moderate equine asthma" was recommended. four british veterinary practices, two primarily serving the flat racing community and two primarily serving the national hunt (jump racing) community, in different geographical regions of england, were purposively selected to participate. focus group discussions were conducted at the practice premises, moderated by one of the authors (tk), an experienced qualitative researcher who is not a veterinarian. discussions were audio-recorded and transcribed verbatim, and transcripts were analyzed using an inductive, thematic analysis. in total, participants contributed to the focus group discussions (number per group ranged from to ). all were veterinarians (experience ranging from recent graduate to senior partner), with the exception of one laboratory team member and one veterinary student, and five were women. discussions lasted between and min. three key themes were developed through analysis of focus group data: (i) an over-arching theme of serving the racing industry within which two further themes (ii) disregarding of the consensus and (iii) the pragmatic clinician were nested. (i) serving the racing industry: this was a key driver of clinical approaches to racehorse respiratory health, which were strongly trainer-influenced in particular. the trainer selects horses for endoscopic respiratory assessment, often because of training and racing schedules rather than any clinical signs, and the approach to investigation and treatment is strongly influenced by trainer expectations. this varies with trainer personality, experience and training methods, as well as stage of the racing season, signalment of the affected animal and general health on the yard, and is in turn driven by commercial pressures of the racing industry. (ii) disregard of the consensus: the unanimous view across all four groups was that the condition defined as mildmoderate ea by current concensus ( ) is largely not seen in british racehorses which, in the participants' considerable collective experience, are affected predominantly with excess endoscopically-visible tracheal mucus largely attributed to bacterial infections. it was also considered unfeasible to fulfill two key aspects of the consensus case definition: waiting for chronicity of clinical signs (> weeks duration), and performing bal sampling. neither of these would be acceptable to trainers, according to participants, and participants themselves were not convinced of the extra value of bal sampling. the consensus statement was therefore seen as having been developed for outsiders, by outsiders without sufficient understanding of culture and practices on british racing yards. (iii) the pragmatic clinician: participants shared a strong professional identity as pragmatic clinicians often required to base clinical decision-making on direct personal or collective experience, rather than on research-based or laboratory evidence. cytological examinations of tracheal wash samples were defended as valuable when interpreted sequentially and combined with knowledge of the history and idiosyncracies of the individual horse and yard. although this approach was generally viewed positively as flexible and individualized, participants did also express some frustration with the sometimes unsatisfactory jigsaw of diagnostic information available to them, particularly in relation to discrepancies between clinical and laboratory findings. our work has highlighted a lack of alignment between clinical practice on british racing yards and international consensus on diagnosing lower airway inflammation, which constitutes a barrier to furthering development of a contextually-relevant evidence-base for this population. equine clinicians elsewhere may find themselves in disagreement with some of the opinions expressed, or practices described, by our study participants. however, these investigations were designed to understand the experiences and rationales of clinicians in the specific context of british racing practice. the strength and consistency of views expressed support the anecdotal evidence that, in this context, tracheal endoscopy and wash sampling are widely regarded as the best available means of providing the non-invasive monitoring of respiratory health expected by trainers and used to inform training-and racing-related decisions. it would be interesting to determine whether similar approaches are being taken elsewhere, particularly in populations of yearling and year old thoroughbred racehorses in training. given the considerable resistance to bal sampling in british racing, development of new tracheal-based or other minimally-invasive diagnostics, including appropriate biomarkers and suitably sensitive, portable lung function tests, would be valuable. furthermore, our participants' views that mild-moderate ea as defined by current consensus is largely not seen in british racehorses suggest that research furthering our understanding of the etiology and pathogenesis of airway inflammation in this equine population is still required. the respiratory system is an interface between the outer environment and the inner body. lower airways have historically been seen as a sterile milieu, thanks to the anatomical configuration, local surface immunity and mucus production and clearance systems ( ) . however, with the development of high sensitivity and high throughput technologies, the microbiota of the respiratory system has been described in healthy subjects in many species, including horses ( , ) . further investigation of the relationship between infectious agents, lower respiratory tract microbiota and the development of mild ea is warranted. we and others have reported descriptive results about the microbiota of horses with mild ea ( , ), but the causality between bacterial flora and the disease is far from being understood. studies on the microbiome use dna extraction followed by high throughput amplification and sequencing of the s amplicon ( ) . the sequences are then filtered and aligned against a taxonomy database to identify and organize operational taxonomic units (otus). descriptive analysis of the phyla, otus and bacterial species are then performed, followed by statistical analysis at the community level (within and between samples; alpha and beta diversity, respectively) and at the individual level (otu diversity analysis). statistical analysis can be used to compare between groups: healthy horses vs. those with mild asthma, upper vs. lower respiratory tract ( ) . the lower airways have a decreased richness (alpha diversity, corresponding to the number and proportion of each bacterial species) when compared to the upper airways in healthy horses ( ) . however, a very large majority of the same otus are present in both the upper and the lower airways, showing an overlap and some continuity in the bacterial population between the two anatomical environments in healthy horses. furthermore, treatment with corticosteroids did not affect the composition of the bacterial flora in the upper airways ( ) . the role of the upper airways microbiota in mild ea is unknown, but two studies did not find any difference in beta diversity of the upper airways between healthy horses and those with mild ea ( , ) . the relationship between bacteria and the lower respiratory tract of the equine host seems to be dynamic. as an example, a change in the environmental respirable particulates has an effect on the lower respiratory tract flora in horses. furthermore, treatment with systemic or nebulized dexamethasone induces some changes in the microbiota of the lower respiratory tract in both healthy and mild asthma horses ( ) . systemic dexamethasone administration decreased the evenness of the flora and increased the abundance of otus. there is an agreement between studies that the lower airways microbiota between healthy and mild ea horses are clearly different ( , ) . interestingly, streptococcus is one of the otus which differed with disease status, and was the otu with the greatest increase in relative abundance in mild ea. the effect of the environment on the composition of the lower airways' microbiota is also a common finding between studies ( , ). however, a study found that treatment with corticosteroids had more effect on the composition of the bacterial flora than changes in the environment ( ) . the microbiome studies are recent in equine medicine and are limited to being descriptive. the challenge for the scientific community will be to answer the causality dilemma of the chicken or the egg regarding the role of the airway microbiota in mild ea. asthma development in humans is most probably caused by the interaction of multiple factors, including genetics, allergen exposure, microbiome and invading pathogens. human rhinovirus, human respiratory syncytial virus, human metapneumovirus, human parainfluenza virus, human enterovirus and human coronavirus are strongly associated with asthma exacerbations ( ) . the association between human rhinovirus-induced wheezing and the development of childhood asthma/wheezing has been confirmed in a recent meta-analysis ( ) . the risk for asthma by age years has been shown to increase (odds ratio . ) if children have been wheezing with rhinovirus during the first years of life ( ) . further, many prospective long-term follow-up studies have shown that human respiratory syncytial virus-induced bronchiolitis is associated with later development of asthma ( ) . however, the pathogenic role of respiratory viruses as triggers for the development and/or exacerbation in asthmatic human patients has not been fully characterized. changes in the immune response to viral infections in genetically predisposed individuals are very likely to be the main factor involved in the association between viral infection and asthma ( ) . the pathogenesis of ea remains incompletely defined. however, similar to human asthma, a multifactorial process is suspected. conditions associated with exercise, feeding and housing practices, location, seasonality, infection of the upper and lower airways and genetic influences have been linked to ea ( , , , ) and bacterial (streptococcus equi subspecies zooepidemicus, actinobacillus spp., pasteurella spp.) etiological agents have been linked to mild to moderate ea ( , ) . it remains to be determined if these agents are triggers for the development of ea or are secondary colonizers of already compromised airways. viral respiratory infections are one of the most common health problems in horses throughout the world ( table ). these infections are often self-limiting and a full recovery can be expected in most horses. young performance horses, such as racing horses, have an increased risk of respiratory viral infections. this relates to age susceptibility, commingling, stress and suboptimal biosecurity protocols ( , , ) . amongst respiratory viruses, only eiv and ervs have an affinity to the lower respiratory tract, leading to airway hyperresponsiveness. clinical signs associated with eiv are usually more severe than those seen with mild to moderate ea. further, no association has been determined between mild to moderate ea and infections with eiv, ehv- and ehv- ( , , ) . this is in sharp contrast to the detection of ervs (erav and erbv), known to cause subclinical or mild clinical disease ( , , ) . in a recent study, horses with mild to moderate ea were significantly more likely to have a positive titer as well as higher log-transformed titers to erav when compared to control horses ( ) . in another study, the detection of erbv by qpcr was significantly associated with coughing in standardbred racehorses in training ( ) . subclinical respiratory viral activity in horses with poor performance has been associated with ehv- and ehv- infection ( , ) . in a recent study, the detection of ehv- by qpcr in nasal secretions was significantly associated with mild to moderate ea ( ) . in another study, the detection of ehv- by qpcr was significantly associated with coughing and excessive tracheal mucus in standardbred racing horses ( ) . these results are in sharp contrast to two recent studies performed on swedish standardbred trotters, which were followed for months via qpcr analysis of nasal secretions and serology ( , ) . despite occurrence of poor performance and subclinical viral activity in the swedish standardbred trotters, the authors were unable to detect associations between ehv- /- and clinical respiratory disease and/or poor performance. these conflicting results reflect the ongoing challenges in establishing causality between mild to moderate ea and gamma herpesviruses, known to be ubiquitous in both healthy and clinically affected horses. in conclusion, associations between specific viruses detected via antigen or antibody detection and clinical signs of mild to moderate ea may suggest that viruses may play a role in triggering or exacerbating asthma. however, because some viruses are ubiquitous both in healthy and clinically affected horses or are often associated with subclinical disease, establishing causality is challenging and in need for further research. a growing body of research demonstrates the link between organic dust exposure and ea. introduction of horses to high dust environments not only induces profound bal fluid neutrophilia and airway obstruction in horses susceptible to severe asthma, but also significant neutrophilic airway inflammation in previously healthy horses ( , ) . outside of the experimental exposure setting, higher dust exposure has also been associated with increased risk of tracheal mucus accumulation in racing thoroughbreds ( ) . barn dust is a complex mixture, rich in potential sources of allergens as well as immunomodulators such as endotoxin and β-glucan ( , ) . in addition to individual horse factors such as age and susceptibility, this complexity may partially account for the heterogeneity of asthma phenotypes. respirable particulates, nominally < µm in diameter, have been linked to eosinophilic inflammation in young thoroughbreds entering race training ( ) and neutrophilic inflammation in actively racing thoroughbreds ( ) . increasing respirable endotoxin exposures have been shown to provide an apparent protective effect against neutrophilic inflammation at low doses ( ) , while high doses of endotoxin augment the inflammatory response to particulates ( ) , suggesting a non-linear response to inhaled endotoxin in the horse. mast cell inflammation has been found to be common in both young, untrained thoroughbreds ( ) and those that are actively racing ( ) , but unrelated to respirable dust or respirable endotoxin exposures. instead, bal mast cell proportions are related with respirable β-glucan exposures. conversely, inhalable dust exposures have not been found to affect bal inflammatory cell proportions. thus, inhalable particulates, those nominally < µm in diameter, appear to be less relevant than respirable particulates in equine respiratory health. setting exposure recommendations will require better understanding of the dose-response to inhaled non-infectious agents across wider ranges of age, breed, and discipline through study designs that include both exposure and respiratory health outcome measures and utilize appropriate statistical tools to relate them. advanced characterization of respiratory health, such as investigation of alveolar macrophage function and bal fluid cytokine profiles, coupled with extensive exposure assessment is likely to offer valuable insight into ea pathophysiology and identify new targets for intervention. miniaturization of optical particle counters has rendered realtime breathing zone exposure measurements on the horse both affordable and technically feasible. finally, the equine airway is arguably most susceptible to particle penetration during athletic exertion due to large tidal volumes and extension of the head and neck, yet the exposures that horses sustain during exercise are largely unexplored. such measures of exposure are complicated by the air speed and turbulence generated at the breathing zone during such activity and will require specialized sampling strategies. neutrophils are key actors in host defense, migrating toward sites of inflammation and infection, where they act as early responder cells toward external insults ( ). however, neutrophils can also mediate tissue damage in various noninfectious inflammatory processes. airway inflammation is one of the primary characteristics of an asthma-affected horse's response to aeroallergens with neutrophilic bronchiolitis being the main lesion ( ) . the mechanism by which airway inflammation develops in ea is a multifaceted and dynamic process. current knowledge suggests that the inflammatory component of this disease results from a combination of both the innate and adaptive immune responses ( ) . generally, airway inflammation involves activation of pathogen-specific inflammatory cells, modulation of gene transcription factors, and release of inflammatory mediators ( ) . within the airways, neutrophils likely contribute to bronchoconstriction, mucus hypersecretion, and pulmonary remodeling by release of proinflammatory mediators, including the cytokines interleukins and , neutrophil elastase, reactive oxygen species, and neutrophil extracellular traps (nets) ( ) ( ) ( ) ( ) . oxidative stress in horses with asthma is evidenced by the increase in elastase and decrease in ascorbic acid concentrations in balf associated with neutrophilia secondary to exposure to organic dust ( ) . the pathogenic role of nets has been described for many infectious and non-infectious human diseases, including respiratory cases with a massive influx of neutrophils into the airways ( ) . excessive net release is particularly deleterious in lung diseases because nets can expand easily in the pulmonary alveolar space and cause lung injury. furthermore, nets and their associated molecules can directly induce epithelial and endothelial cell death ( ) . the mechanisms that regulate neutrophil functions in tissues are complex and incompletely understood and must be regulated with exquisite precision and timing. timely apoptosis of neutrophils is central to the resolution of inflammation; dying neutrophils are known to stimulate their own efferocytosis, inducing macrophagic transition from a pro-inflammatory to an anti-inflammatory profile ( ) . thus, dysregulated apoptosis and mechanisms of inflammation may play an important role in the pathogenesis of ea. the persistence of apoptosis-resistant neutrophils in the airways of horses with asthma may also impede timely neutrophil clearance and delay the resolution of airway inflammation. the discovery and development of compounds that can help regulate ros, net formation, cytokine release and clearance of airway neutrophils would be highly beneficial in the design of therapies for ea ( ) . asthma is a highly heterogeneous condition of the lung. akin to the lining of the gastrointestinal tract, the lining of the airways is also in contact with external substances throughout life. ingested substances generally pass through the gastrointestinal tract unidirectionally, and a careful balance between processing of digested food materials, nutrient absorption and limiting immunoreactivity is maintained during homeostasis, with wellknown severe consequences of deviations in this balance. the airways function differently in that only gaseous substances normally pass into the distal alveoli and are exhaled in the reverse direction. inhaled particulates also have to be expelled in reverse direction toward the nasopharynx by largely mechanical means or taken up by alveolar macrophages for disposition with minimal inflammatory evocation ( ) . hence, a complex and selective epithelial barrier with differing functions characterizes both organs. the epithelium lining the airways has unique composition, morphology and function throughout the lung, and is intimately connected to subepithelial structures such as the basement membrane, mucous glands, smooth muscle, fibroblasts, endothelium and immune cells. the epithelium forms a barrier between inhaled components and the subepithelial constituents, and also has to balance efficient transfer of gases with controlled reactivity to non-gaseous components. while the lesions of severe ea manifest predominantly with inflammation, smooth muscle hyperplasia and fibrosis of the peripheral airways and surrounding tissues, the larger airways are exposed to the same inhaled substances and also have morphological, functional and molecular changes ( ) . research initially focused on the role of club cell secretory protein (ccsp), a member of the secretoglobin family produced by non-ciliated epithelial cells concentrated within the epithelium at the transition from bronchi to bronchioles. club cells are recognized as epithelial progenitor cells that can differentiate into ciliated and other specialized cells of the airway epithelium, participate in reduction of reactive oxygen toxicants through cytochrome enzymes, and their hydrophobic secreted protein inactivates a range of inflammatory mediators. horses with severe asthma have fewer club cells and lower concentration of ccsp in airway fluids, which may be a function of chronic inflammation resulting in reduced regenerative capacity of the airway epithelium ( ) . unique relative to other mammals, equids have two expressed ccsp genes that differ in of amino acids, and also in their interaction with hydrophobic molecules ( ) . recombinant eccsp increased neutrophil oxidative burst, phagocytosis and extracellular trap formation, lending support to the notion that loss of club cells has deleterious effects on lung health ( ) . whole transcriptomic changes in endobronchial epithelial biopsies from sites from th to th generation bronchi were investigated with next-generation sequencing. each horse served as its own control to identify changes in gene expression associated with an inhaled challenge since inter-individual variability exceeded changes attributable to the challenge. a bioinformatics pipeline including quality control measures to account for duplicates, variable sequencing depth and dispersion was implemented, results were mapped to the equine genome, and predicted proteins were procured with a combination of software and manual approaches to assign appropriate ensemble ids for analyzing interactions. an overall conservative analytic approach yielded genes differentially expressed in horses with severe asthma as a result of a challenge, with the majority up-regulated ( ) . not surprisingly, many up-regulated genes pertained to inflammatory mediators and effectors and were well-known members of protein interacting networks. however, somewhat more surprisingly, genes with altered expression also concerned more broadly epithelial cell formation and maintenance, and the circadian rhythm, suggesting that multiple cell properties are affected in exacerbated ea at the transcriptomic level. subsequent analysis of enriched gene sets in asthmatic horses further highlighted the importance of cell cycle regulation and repair pathways ( ) . transcriptomic studies of this nature yield a great deal of information, which requires subsequent confirmation regarding cell specificity, correlation with protein expression and function, and extension to a more robust number of affected and unaffected individuals. albeit, there is strong evidence to indicate that the bronchial epithelium is profoundly altered during exacerbation of severe ea, and this insight offers new venues for investigating the role of specific proteins and for potential therapeutic targets ( , ) . the entire spectrum of ea is influenced by interactions between the environment and genetics, but almost all research in this field has focused on the severe clinical phenotype. while no specific genetic risk factors have been reported for mild to moderate forms of ea, genetic susceptibility to certain bacterial lower airway infections could potentially be relevant ( ) . furthermore, mild but persistent respiratory signs such as occasional coughing and nasal discharge may represent early phenotypic indicators for an increased risk to later development of severe ea ( ) . this suggests that the genetics of milder forms of ea may be worth investigating in longitudinal studies. severe ea has been shown to be partly heritable in several breeds and has been the focus of genetic research involving family and epidemiological studies, whole-genome scans and investigation of candidate genes. reports of marked familial aggregation of severe ea date back years ( ) . parent, age, and stable environment have significant additive effects that increase the risk for developing severe ea as defined by a history of persistent frequent coughing and/or increased breathing effort ( , ) . offspring of affected sires have a more than -fold increased risk for developing severe ea ( ) . whole genome scans in high-prevalence families indicate two chromosome regions with a genome-wide significant association with severe ea ( ) . importantly, the associations differ between the families: region eca in one family and eca in another family. further association and gene expression studies indicate interleukin receptor as a candidate gene in a subset of ea-affected horses. molecular pathway analyses of genomic and proteomic data showed interactions between interleukin receptor and socs upstream of an important molecular cascade involving nuclear factor κb ( ) . so far, no causal genetic variant has been identified in interleukin . an allelic case-control genome-wide association study in the general warmblood population revealed another region on chromosome . the best-associated marker was located in the protein-coding gene txndc , which may be involved in regulating hydrogen peroxide production in the respiratory tract epithelium as well as in the expression of muc ac mucin ( ) . no genomic copy number variations were found to be associated with severe ea ( ) . integrative analyses combining gwas, differential expression (de), and expression quantitative trait loci (eqtls) were not able to uncover causative genetic variants that contribute to severe ea through gene expression regulation. however, results showed interesting similarities to human asthma with disease-associated genetic variants in clec a that also regulate gene expression of dexi ( ) . furthermore, global gene expression studies of mrna and mirna levels in these high-prevalence families have shown impaired cell cycle regulation and cd + t cell differentiation into th /th cells, respectively, in severe ea ( , ) . at present, none of these associations are useful genetic markers in the general population. most of the findings pertain to warmbloods only, or even only to certain lines and families. the fact that the chromosomal regions and the mode of inheritance do not agree between families indicates genetic heterogeneity for severe ea: depending on the genetic make-up of affected horses, different genes confer the susceptibility for the disease. it appears that the genetic basis of severe ea is robust, but remarkably complex. polygenic complexity, potentially with a larger number of genes that each may only contribute < % to the total genetic effects, may make it difficult to discover causative variants. nevertheless, the genetics of severe ea has revealed interesting links between severe ea, allergic skin diseases and susceptibility to intestinal parasites ( , ) . according to the national institutes of health, a biomarker is a characteristic that is objectively measured and evaluated as an indicator of normal biological processes, pathogenic processes or pharmacologic responses to a therapeutic intervention ( ) . in practice, biomarkers include tools and technologies that can help in understanding the prediction, cause, diagnosis, progression, and outcome of treatment of a disease. although bal cytology has been recognized as the gold standard for diagnosing respiratory diseases such as ea, currently, sensitive and specific biomarker tests useful in routine laboratory diagnostics are being sought. a simple biomarker capable of distinguishing between animals with lower airway infections and those with non-infectious airway inflammation would be helpful. although the diagnosis of severe cases of ea is relatively easy, it is difficult to diagnose cases in remission or horses with a mild form of the disease. ideally, molecular biomarkers should reflect a feature of relevant pathological processes. in addition, biomarker assessment should be easy, low-cost, technically accurate, repeatable and have an acceptable risk. therefore, a measurement from easily obtainable body fluids or tissues is preferred, such as blood, urine, exhaled breath condensates, as opposed to bal, transbronchial biopsy or lung biopsy ( ) . several biomarkers are present or altered in the airways or circulation of horses with asthma. inflammatory markers such as acute phase proteins and cytokines have been studied as markers of systemic inflammation. however, the available literature on markers of systemic inflammation in horses with severe ea is not well-characterized and controversial ( , ( ) ( ) ( ) . apart from reports on differential expression of cytokines during the course of severe ea, only a few acute phase proteins have been investigated. haptoglobin is a suitable marker of both acute and chronic systemic inflammations, whereas high concentrations of serum amyloid a indicate acute inflammation. one study found no difference in the acute phase protein levels (serum amyloid a, c-reactive protein, haptoglobin) between horses with mild ea and those with other causes of exercise intolerance ( ) . another study found elevated haptoglobin concentration in horses with mild ea ( ) . surfactant protein d is a large multimeric collagenous glycoprotein produced mainly by type ii epithelial cells in the lungs and is also detectable in the serum. serum surfactant protein d has been identified as a potential systemic biomarker for some pulmonary diseases in humans, such as idiopathic interstitial fibrosis and acute respiratory distress syndrome. elevated serum levels of surfactant protein d have been detected in horses with mild ea ( , ) . circulating immune complexes are proteins that result from an immune response against an organism or antigens of various origin. in humans, circulating immune complexes are detectable in a variety of systemic disorders such as autoimmune diseases, allergies and infectious diseases ( ) . high levels of circulating immune complexes have been reported in horses with severe ea ( ) . another study found circulating immune complexes useful for differentiating healthy vs. severe ea, and monitoring corticosteroids therapy ( ) . the main group of enzymes responsible for collagen and other protein degradation in the extracellular matrix are matrix metalloproteinases (mmps), while tissue inhibitors of metalloproteinases (timps) lead to fibrosis formation. collagen is the main structural component of connective tissue and its degradation is a very important process in development, morphogenesis, tissue remodeling, and repair. in horses with severe ea, mmps, timps, and their ratios are useful in the evaluation of the severity of respiratory disease and in identifying subclinical cases ( ) . furthermore, mmp- , mmp- , timp- , and timp- are significantly decreased after therapy with inhaled glucocorticoid therapy ( ) . exhaled breath condensate is a promising source of biomarkers of lung disease in humans. exhaled breath condensate hydrogen peroxide concentration and ph were higher in horses with mild ea, vs. controls ( ) . additionally, both hydrogen peroxide and ph had a positive association with bal neutrophil percentage, while leukotriene b- demonstrated a positive association with bal eosinophil percentage. another study characterized the metabolomic profile of tracheal wash and exhaled breath condensate in healthy horses and those with severe ea ( ) . higher concentrations of histamine and oxidant agents, such as glutamate, valine, leucine, and isoleucine, as well as lower levels of ascorbate, methylamine, dimethylamine and o-phosphocholine, were found in the group of severe ea, compared to healthy controls. many biomarkers of ea have been studied-some are already being used in clinical settings, while others require further studies. however, history, clinical evaluation, and bal still constitute the basis for diagnosis of ea. immune response has mainly been investigated in the airways of horses with severe ea and more recently mild-moderate ea, while still representing one of the futures direction for research stated in the acvim consensus statement ( ). such characterization has mostly been performed through relative mrna expression of various cytokines in bal fluid, while several publications also reported protein concentration in bal fluid for few cytokines. various methodologies for cytokine mrna expressions have been published (e.g., sybr green or taqman technology, design of primers and probes, relative quantitation, etc.). variation in methodologies may ultimately prevent objective comparisons between reports, as well as the implementation of prospective, multicenter studies. such diversity should however not be considered as a scientific weakness, and methodological homogenization among the various research groups neither represents a prerequisite nor a final goal to be reached. however, evaluation of the methodological performances of different research laboratories might represent a relevant goal. in this manner, implementation of inter-laboratory comparisons based on international standards (e.g., iso/iec and iso ) warrants further consideration. let's consider for example mrna expression of two different cytokines by pcr in balf samples. as a first and informal procedure, a simple "blind test" could be performed among up to four different teams. in this procedure, the "reference lab" will provide the three other labs with aliquots of the same sample(s). each team will evaluate mrna expression for these two cytokines based on their own procedures, and comparisons of the results obtained and agreement among the teams can be evaluated. this "blind test" might then be repeated on a regular basis, systematically alternating the "reference lab" within the group. in the end, the procedure will provide an objective evaluation of the results diversity among the teams, but clearly will not determine whether several teams are more efficient than others for these specific analyses. a second and more structured procedure would require the specific synthesis of standards (mrna for two different cytokines in this case), and the development/validation of relevant conditioning and conservation procedures. a similar group of four different labs would first evaluate their ability to detect and quantify predetermined amounts of analytical standards (evaluation of the detection, not of the sample extraction, etc.). this step is a necessary preliminary, in the absence of reference methods. a panel of at least samples (previously calibrated with standards) would then be tested, including several identical ones (for repeatability) and submitted to the group (including a "self-shipment") for testing and further statistical analyses (agreement, etc.). once the methodological performance of the lab is considered acceptable for this panel, the procedure might then be repeated with another two cytokines and so on. in the end, the whole panel of standardized samples might allow the establishment of a labeling, accessible to any voluntary laboratory involved in equine asthma. mandatory considerations about such comparisons are that there is no trap, and this does not represent an overall examination of laboratories, but simple evaluations of procedures. all labs are expected to use their methodologies, whether or not the technologies are similar within the group. among others, samples conditioning, conservation, shipment and their associated costs will represent major issues to be considered, and this should be more broadly associated with virtuous initiatives such as the equine respiratory tissue biobank. several group discussions were conducted during the havemeyer equine asthma workshop to identify future research priorities. initial rotating small-group topic explorations (pathophysiology, risk-factors, diagnostic methods and phenotype definition) facilitated by members of the workshop organizing team, were followed by a final large group "roundtable" discussion of key directions for future ea research. the discussion was informed by data gathered directly from ∼ participants (i.e., all who attended the final roundtable), who were invited to propose up to three short-or long-term, focused or "big picture, " research topics or ideas that they considered to be key future research directions. these data were submitted anonymously, during the workshop, as free-text on paper and loosely arranged into broad categories for further open discussion. following the workshop, in order to present an accessible, systematic and non-selective summary of the ideas proposed by participants, the free-text data were collated in microsoft excel for content analysis using an approach based on recommended methods for quasi-qualitative data ( , ) . the text was transcribed verbatim and coded at two levels to categorize content into (i) broad topic areas (level ) and (ii) specific subsets of these topics (level ). all instances of each level topic code were then exported into online software (worditout) to create a word cloud (figure ) , in which the relative frequencies of occurrence of each topic are represented by font size. overall, responses were received, each proposing between and research ideas, resulting in a total of research ideas, which were organized into the broad topic codes presented in the word cloud. some research ideas encompassed more than one topic and were identified with multiple codes to reflect this. frequencies of occurrence of each code ranged from n= for "diagnostics" to n = for "genetics." specific proposed areas of interest in the dominant "diagnostics" category were the development of improved, non-invasive field diagnostics through the identification of suitable biomarkers, development of portable lung function tests, improved understanding of relative values of tracheal wash in comparison with bal cytology, or relationships between the two, and identification of gold standards for all of these diagnostic modalities. another key topic was phenotype distinction ( occurrences)-in particular to clarify any distinction between mild and moderate ea, and to determine whether or not such a distinction is valuable in terms of differing pathophysiology, diagnostic indicators, therapeutics or prognosis. as with many of these proposed topics, phenotype distinction rests on the back of the category "diagnostics"-pointing out a self-identified weakness on the part of ea researchers that the goal of identifying the horse with asthma so mild that it does not present as respiratory disease per se, continues in many cases to elude us and underscores a collective pragmatism that there is little benefit in understanding the fine points if we cannot definitively identify the case in the first place. ideas relating to therapeutics ( occurrences) included investigating the efficacy of different treatments including environmental management and any evidence for the value of antibiotics, as well as the development of optimal nebulized glucocorticoids, alternatives to corticosteroids, immunological treatments, respiratory probiotics, other novel therapeutics (e.g., marcks inhibitor peptide), and individualized treatments for different endotypes and phenotypes. suggestions relating to pathophysiology ( occurrences) included furthering our understanding of the role of environmental pollutants, of when a physiological response becomes a pathological response and of factors influencing progression from mild to severe equine asthma. standardization ( occurrences) referred in particular to the need to develop or agree on standardized diagnostic approaches, including in relation to bal collection techniques, laboratory processing and cytological methods and threshold values, context-specific reference ranges, development of a central repository of protocols and improved quality control protocols. a central repository of standard protocols was suggested. academic-clinical communication ( occurrences) was recognized as an area for general improvement. related research suggestions included improving our understanding of the views and practices of field clinicians, as well as their perceptions of disease progression and treatment efficacy, particularly in regions outside the uk (to build on the kinnison and cardwell uk study) ( ) . this would inform the enhancement of multidirectional communication between academia, referral and first opinion clinical practice, development of guidelines and apps for field practice and overall improved dialogue and engagement. better use of collaborative, epidemiological and longitudinal studies was suggested for many topics and included multicenter, cross-country collaborations, more use of the existing tissue bank and the initiation of a new equine asthma group. it is recognized that the ideas for research directions generated through this roundtable discussion at the end of a day workshop are subject to biases and influences relating to the interests, priorities and perceptions of workshop participants. however, by using and describing a systematic method of representing the ideas proposed, we have aimed at least to be transparent in our reporting of this. further, longer-term, international discussion and exchange of views will be facilitated by one of the key outcomes of this workshop, which was the development of the new equine asthma group. the aim of this group is to offer a platform of information for veterinary practitioners and horse owners as well as a resource for researchers to collaborate and exchange ideas on the understanding of ea. it was suggested that this group could lead some initiatives in line with the proposed areas of interest described above. there are plans for this group to develop some guidelines for the diagnosis and treatment of equine asthma, including for example the standardization of diagnostic methods, as mentioned above. development of an equine asthma group website and other communication tools are now underway as an internationally collaborative initiative. the havemeyer equine asthma workshop has paved the way for a better understanding of this many-faceted disease by bringing together researchers and clinicians to identify both the needs of the equine industry for effective treatments and at the same time focus researchers on the gaps in knowledge and understanding that will facilitate our ability to deliver on these needs. the participants made clear the requirement for more accessible, standardized diagnostics that will enable us to understand the underlying pathophysiology and identify specific phenotypes and endotypes and thus create more targeted treatments or management strategies. by creating an 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asthmalike disease of horses markers of systemic inflammation in horses with heaves circulating immune complexes and markers of systemic inflammation in rao-affected horses acute phase proteins in racehorses with inflammatory airway disease serum surfactant protein d and haptoglobin as potential biomarkers for inflammatory airway disease in horses tests for circulating immune complexes circulating immune complexes in horses with severe equine asthma metalloproteinases and their inhibitors are influenced by inhalative glucocorticoid therapy in combination with environmental dust reduction in equine recurrent airway obstruction metabolomics of tracheal wash samples and exhaled breath condensates in healthy horses and horses affected by equine asthma is there anything else you would like to tell us" -methodological issues in the use of free-text comments from postal surveys working with words: exploring textual analysis in medical education research the authors are grateful to gene pranzo, president of the havemeyer foundation, for his support of the workshop on equine asthma. we are also thankful to boehringer ingelheim animal health, haygain, nortev, trudell medical international, and zoetis for their support of travel grants for participants and workshop activities. the workshop was made possible thanks to the generous support of the dorothy havemeyer foundation, boehringer ingelheim, haygain, nortev, trudell medical and zoetis. key: cord- -wn u u y authors: zheng, yichao; zhu, yinheng; ji, mengqi; wang, rongpin; liu, xinfeng; zhang, mudan; qin, choo hui; fang, lu; ma, shaohua title: a learning-based model to evaluate hospitalization priority in covid- pandemics date: - - journal: patterns (n y) doi: . /j.patter. . sha: doc_id: cord_uid: wn u u y summary the emergence of novel coronavirus disease (covid- ) is placing an increasing burden on the healthcare systems. although the majority of infected patients have non-severe symptoms and can be managed at home, some individuals may develop severe disease and are demanding the hospital admission. therefore, it becomes paramount to efficiently assess the severity of covid- and identify hospitalization priority with precision. in this respect, a -variable assessment model, including lymphocyte, lactate dehydrogenase (ldh), c-reactive protein (crp) and neutrophil, is established and validated using the xgboost algorithm. this model is found effective to identify severe covid- cases on admission, with a sensitivity of . %, a specificity of . %, and an accuracy of % to predict the disease progression toward rapid deterioration. it also suggests that a computation-derived formula of clinical measures is practically applicable for the healthcare administrators to distribute hospitalization resources to the most needed in epidemics and pandemics. the novel coronavirus disease (covid- ) caused by the severe acute respiratory syndrome coronavirus (sars-cov- ) infection was firstly reported in last december in china and rapidly spread across the world, affecting over million people worldwide and killing more than a half million infected patients up till now - . even worse, the global pandemic of covid- is expected to continue growing, as no effective vaccines have been officially approved for prophylaxis of this disease . though the growth in detected infections has declined in east asia and europe, the number of infections in u.s., south america and african places are witnessed with continuous growth . moreover, the suspicion on a second generation pandemic outbreak still sustains . in pandemic, a nation's healthcare system bears extraordinary burdens. however, a majority of patients infected with sars-cov- generally have non-severe disease progression and can be safely managed at home or self-quarantine, and recover under limited and basic medical care . for the infections with severe syndromes or progression toward rapid deterioration, immediate admission to hospitals for close monitoring and intensive treatment has been proven effective to reduce the complications and mortality . therefore, identifying the covid- patients at high risk for severe illness and prioritizing them for immediate admission to hospitals becomes urgently demanded, especially in nations and territories where the healthcare systems are insufficient to administrate all infections and suspicions. some studies have been reported to predict deterioration and mortality during hospitalization - , and even predict the probability of sars-cov- infections that enables the the timely quarantine of high rate infections and prevents their spreading - . but none of these studies aim to provide a solution for rational triage of patients in places where the medical resources are limited. in light of this unmet need in efficient triage of covid- cases, the study is sought to develop and validate a learning-based model that evaluates patients' priority of being admitted to hospital care due to their appearance or susceptibility toward severe covid- . the model, provided with a simple user interface, can efficiently assess the severity of covid- and predict the disease progression, with high rates of accuracy. our study is expected to have a prolonged social impact under the current circumstances, when the simple and practical model becomes accepted to assist clinicians in quick and efficient triage of covid- patients. this study was approved by the guizhou provincial people's hospital ethics committee. the patients cohorts enrolled in this study were comprised of covid- cases retrieved from world health organization (who) covid- database ( figure s , table s ) and covid- cases recruited from a multi-center dataset in china. amongst the patients, . % of patients had developed severe disease on admission and . % of patients were presented with non-severe diseases on admission but progressed toward severe disease after admission. the minimal, medium and maximal time from hospital admission to severe disease progression were less than day, days and days, respectively. the prevalence of underlying comorbidities was diseases ( . %). the medium age was years. fever was the most common initial symptom ( . %), followed by cough ( . %), fatigue ( . %) and dyspnea ( . %). table shows the baseline laboratory results obtained on or soon after admission. all the patients were laboratory-confirmed covid- cases and the severity of covid- were stratified into severe and non-severe categories according to a criteria shown in table . the clinical variables of most patients were measured multiple times across different days during hospitalization to assess the prognosis. as this study was sought to identify the hospitalization priority according to the prehospital assessment of severe covid- risk, only clinical data obtained on admission were used to evaluate the importance of clinical variables in identification of severe or potentially severe cases. given the various missing data on different clinical variables and different patients, a strategy was adopted to set a threshold value alpha to remove these missing data, minimizing their impact on data analysis. it was found that as the threshold alpha increases, available variables decrease while available observations, i.e. the available covid- cases, increase ( figure s ) . a threshold alpha of was selected to remove the missing data, and to obtain as many clinical variables and observations as possible. hence, a total of clinical variables between the severe and the non-severe groups. as shown in table s , a total of clinical variables were significantly different between the two groups, including the age, fever, dyspnea, lymphocyte, neutrophil, c-reactive protein (crp), lactic dehydrogenase (ldh), creatine kinase (ck), d-dimer, alanine aminotransferase (alt), aspartate aminotransferase (ast) and albumin. these clinical variables could be used to discriminate between the severe and non-severe covid- cases. extreme gradient boosting (xgboost), which is a high-performance machine learning algorithm and works with a sequence of decision trees where the latter tree tries to minimize the net error from prior trees, was used to generate the risk assessment model. accuracy, f score, sensitivity, specificity, and the area under curve (auc) score of receiver operating characteristic (roc) curve. the definition of these evaluation metrics . % in discriminating severe covid- cases from their non-severe counterparts (table ) . moreover, it outperformed other classifiers in the aforelisted evaluations, excepting the specificity (table , and figure ). our study was in agreement with a reported conclusion that the xgboost algorithm had high discriminative performance , and thus could be used to assess the hospitalization priority with precision. next, the assembly of variables was minimized to ease the clinical use. for this purpose, a sequential variable selection approach was used to find the optimal variable set based on its assessment performance. briefly, important variables ranked by xgboost ( figure ) were sequentially assembled in an individualized manner to investigate their incremental effects in terms of auc scores by cross validation. the auc scores ceased to grow when the count of assembled variables increased to (figure ). thus the previous -variable models shrinked to the selected -variable models, where xgboost classifier achieved an accuracy of . % in the identification of severe covid- cases. table compares the performance of various classifiers in the - variable model. the auc score of xgboost was slightly decreased compared with others models, but xgboost achieved the highest f score and accuracy among the classifiers (table , figure ). an over % accuracy indicated that the -variable xgboost model could play a crucial role in distinguishing the majority of cases that require immediate medical attention. overall, the -variable xgboost model was evaluated to be the most competitive and easy-to-use establishment throughout comparison with other prevalent choices. was effective to predict the risk of deterioration for patients who were presented with non-severe symptoms on admission. for this purpose, a total of patients who had non-severe covid- on admission but experienced deterioration during hospitalization were enrolled as an external testing set for analysis ( figure s ). the -variable xgboost model achieved % accuracy in predicting the risk of rapid deterioration (table ) . for patients who had complete time course of exacerbation, the minimal, medium and maximal prediction horizon were less than day , days and days, respectively, suggesting that our model could predict the risk of disease deterioration, for as long as days earlier than its occurrence. to test whether a clinical operable single-tree xgboost classifier based on the lymphocyte count, crp level and ldh level as reported by li et al. was able to accurately identify the risk of severe disease on admission, we performed the single- tree xgboost in identification of severe covid- cases as well as in prediction of risk of in-hospital deterioration from non-severe to severe disease. table s shows the single-tree xgboost had % accuracy in identification of severe covid- cases on admission, but only . % accuracy in prediction of risk of in-hospital deterioration. it suggested that a model established for other purposes or reported in other works does not fulfill our goal in this study, that is, identifying hospitalization priority for covid- collectively, the -variable xgboost model is the first computation model established to assess hospitalization priority that enables rational triage of infected patients and prioritize hospitalization to the most needed. shapley additive explanations (shap), as a game-theoretic approach that interpreted an impact of each input variable toward the model output, had been relied upon for the model interpretation. in figure s , each dot corresponds to an individual case in the study. different colors encoded different values of input variables, while the shap value represented the impact of each variable on the prediction outcome. as shown, the risk of severe covid- was found associated with a decrease in the lymphocyte count, and an increase in the ldh level, the crp level and the neutrophil count. next, the t-distributed stochastic neighbor embedding (t-sne) algorithm as a technique for dimensionality reduction was used to project the four-dimensional data (lymphocyte, ldh, crp, neutrophil) into a -dimension ( d) feature space for visualization . it enabled to visualize the difference in features among the three groups of patients, including the severe cases, the non-severe cases and the progressed severe cases. the progressed severe cases referred to patients with non-severe disease on admission but developed severe disease afterward. there was a clear separation between the non-severe (distributed in the core) and severe cases (distributed in the periphery), whereas the progressed severe cases distributed in between the core and the periphery ( figure s , video s ). suggests that the previously reported prediction models are not suitable for identification of the hospitalization priority for the severe or potential severe covid- cases. (table s ). more recently, a nomogram developed by gong et al. can assist the early identification of severe covid- cases with a sensitivity of . % and a specificity of . % , this study was prematurely accomplished before all the participants had fully experienced the outcome of event. therefore, our study becomes the first to focus on the triage patients, achieving a relatively high sensitivity and specificity, and meanwhile, without negatively impacting its performance by involving participants still in treatment. in this study, the clinical features of covid- were screened to identify a total of critical variables that were found to be associated with the risk of severe covid- severe cases that require immediate medical attention (table ) . importantly, it precisely predicts the risk of progression toward rapid deterioration for as long as days ahead of its occurrence (table ) . our study is in line with the previous studies that showed the increased inflammatory figure s ). in this way, the impact of missing values on data analysis can be reduced. third, there was no significant difference in the prevalence of comorbidies between the severe and the non-severe covid- cases in our datasets (table s ) shaohua ma; ma.shaohua@sz.tsinghua.edu.cn. no new unique reagents were generated in the present study. the clinical data used in this study were obtained from the who covid- database (table ) . specifically, a severe case of covid- was defined by the presence of any of the following conditions in the quiescent state, such as an increased respiratory rate of ≥ breaths/minute, decreased oxygenation index ≤ mmhg or declined spo of ≤ %. moreover, patients who developed shock, multiple organ failure (mof) that were required to be admitted to intensive care unit (icu), or respiratory failure that warranted mechanical ventilation were stratified into the severe category in the present study. finally, patients with pulmonary lesions that showed rapid progression of over % within - hours were considered to have severe disease. as there were various missing data in the datasets, a threshold alpha was set to remove to these missing data ( figure s ). first, the clinical variables with missing data which exceeded the threshold alpha were removed (bottom figure) . second, the observations (that were covid- cases) with missing data for any of the resulting clinical variables were removed. an optimal threshold alpha should be selected to obtain as many clinical variables and observations as possible. next, covid- cases with non-missing variable values were grouped into the severe and non-severe categories, according to the severity of disease. the difference in the clinical variables between the two groups was identified via the univariate descriptive statistics (table s ) . a p-value of less than . was considered statistically significant and was used as a threshold to identify key clinical variables for model development. these key clinical variables were assembled to generate the risk assessment models based on the xgboost classifier as well as other classifiers, such as the lda, logistic regression, svm, random forest, and decision tree. set at a ratio of : . the different models were trained in the training set and evaluated in the holdout testing set by comparing the values of accuracy, f score, sensitivity, specificity, and auc score of roc curve (table s ) . subsequently, the assembly of key clinical variables was minimized to generate the simplified models. for this purpose, all the key clinical variables were ranked according to the importance calculated by xgboost (figure ) , followed by the sequential variable selection approach (figure ) . this was to minimize the variable set while optimize the model performance. the simplified models based on the minimized variable set were trained and evaluated in accordance with a method mentioned above. to assess the effectiveness of models in early prediction of severe progressions, patients who were presented with non-severe symptom on admission but developed severe disease during hospitalization were enrolled as an external testing set for analysis. the performance of models was reflected by accuracy (table ) . as for comparison, the performance of a previously validated single-tree xgboost model in identification of severe covid- risk was assessed in our datasets (table s ) . variable models in discriminating the severe covid- cases. the variables included age, fever, dyspnea, lymphocyte, neutrophil, c-reactive protein, lactic dehydrogenase, creatine kinase, d-dimer, alanine aminotransferase, aspartate aminotransferase and albumin. highlights: a model was developed to evaluate hospitalization priority in covid- pandemics. this model used easily accessible biomarkers to evaluate the risk of severe covid- . the evaluation can be rapidly proceeded using an online program. performance of different algorithms in evaluation of covid- severity was explored. etoc blurb: the authors proposed a learning-based model to assist clinicians in quick and efficient triage of patients in places where the medical resources are limited in covid- pandemics. this model used four easily accessible biomarkers to assess the severity of covid- , and was found effective to identify the risk of severe covid- . it will enable the healthcare administrators to distribute hospitalization resources to the most needed. bigger picture: covid- pandemic is threatening millions of lives and stressing the medical systems worldwide. though the infection growth in some areas has creased, the risk of second wave of outbreak is under threatening. so, a sustainable strategy to defend the pandemic using current limited but effective healthcare resources is in high demand. our study is deemed to find a solution that triages patients to hospitalization by identifying their severity progression. in this study, a model that used four easily accessible biomarkers to assess the risk of severe covid- was successfully developed. this model is easy to use and it eliminates the dependencies on exquisite equipment to make a decision. it was found effective to identify the risk of severe covid- . so, it is practically applicable for general practitioners to effectively distribute the infections and allocate in-patient cares to the most needed. our study is expected to have a prolonged social impact under the current circumstances. association of radiologic findings with mortality of patients infected with novel coronavirus in wuhan prediction models for diagnosis and prognosis of covid- infection: systematic review and critical appraisal real-time tracking of self-reported symptoms to predict potential covid- chest ct for typical -ncov pneumonia: relationship to negative rt-pcr testing a model to predict sars-cov- infection based on the first three-month surveillance data in brazil. medrxiv deep learning-based model for detecting novel study. medrxiv development and utilization of an intelligent application for aiding covid- diagnosis. medrxiv, covid- early warning score: a multi-parameter screening tool to identify highly suspected patients. medrxiv an artificial intelligence-based first-line defence against covid- : digitally screening citizens for risks via a chatbot. biorxiv development and validation of a diagnostic nomogram to predict covid- pneumonia. medrxiv rapid and accurate identification of covid- infection through machine learning based on clinical available blood test results. medrxiv development and validation of chest ct-based imaging biomarkers for visualizing data using t-sne diagnosis and treatment of novel coronavirus pneumonia clinical characteristics of coronavirus disease in china covid- : treating and managing severe cases clinical management of severe acute respiratory infection when covid- is suspected: interim guidance v laboratory testing for coronavirus disease ( covid- ) in suspected human cases: interim guidance clinical characteristics of hospitalized patients clinical features of patients infected with novel coronavirus in wuhan covid- : towards understanding of pathogenesis cough expectoration hemoptysis dyspnea catarrh fatigue anorexia nausea/emesis myalgia dizziness/headache pharyngalgia abdominal pain/diarrhea laboratory findings, mean± std white blood cell count, /l lymphocyte count, /l neutrophil count, /l erythrocyte sedimentation rate, mm/h c-reactive protein, mg/l procalcitonin, ng/ml d-dimer, ug/ml alanine aminotransferase, u/l aspartate aminotransferase, u/l total bilirubin, umol/l albumin, g/l lactate dehydrogenase, u/l blood urea nitrogen we thank the above-mentioned cooperating hospitals for kindly sharing the data with us, in accordance with the declaration of helsinki. the work was supported by the national the authors declare no competing interests. definitions non-severe covid- patients have non-specific symptoms such as fever, cough, fatigue, myalgia, pharyngalgia, but have no signs of dehydration, sepsis or shortness of breath. the radiological examination showes no signs of severe pneumonia. severe covid- adult cases meeting any of the following criteria:( ) respiratory rate ≧ breaths/ min;( ) oxygen saturation ≤ % at rest;( ) fio ≦ mmhg.( ) pulmonary lesion progression exceeds % in - hours ( ) respiratory failure that requires mechanical ventilation; ( ) shock; ( ) organ failure that requires to be managed in intensive care unit key: cord- -glmshsh authors: yin, r.; yang, z.; wei, y.; li, y.; chen, h.; ma, d.; dan, m.; zhang, y.; liu, x.; leng, h.; xiang, d. title: clinical characteristics of patients with neurological diseases and co-morbid coronavirus disease : a retrospective study date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: glmshsh objectives:to describe the clinical characteristics of patients with coronavirus disease (covid- ) with co-morbid neurological symptoms. design:retrospective case series. setting:huoshenshan hospital in wuhan, china. participants:from february to april , patients with neurological diseases were enrolled from all patients in the hospital with confirmed covid- and divided into a severe group and a nonsevere group according to their covid- diagnosis. main outcome measures:clinical characteristics, laboratory results, imaging findings, and treatment methods were all retrieved through an electronic medical records system and recorded in spreadsheets. results:the mean (standard deviation, sd) age of patients was . ( . ) years, and patients were male ( . %). among patients with co-morbid neurological diseases, had a previous cerebral infarction ( . %), had dementia ( . %), had acute cerebral infarction ( . %), had sequelae of cerebral haemorrhage ( . %), had intracranial mass lesions ( . %), had epilepsy ( . %), had parkinsons disease ( . %), and had myelopathy ( . %). fever (n = , . %) was the most common symptom. the most common neurological symptoms were myalgia (n = , . %), followed by extremity paralysis (n = , . %), impaired consciousness (n = , %), and positive focal neurological signs (n = , . %). eight patients ( . %) died. there were more patients with altered mental status in the severe group than in the non-severe group ( [ . %] vs. , p = . ). the inflammatory response in the severe group was more significant than that in the non-severe group. there were more patients taking anticoagulant drugs ( [ . %] vs. [ . %], p < . ) and sedative drugs ( [ . %] vs. [ . %], p = . ) in the severe group than in the non-severe group. amid all patients with cerebrovascular diseases, only ( . %) were taking aspirin, ( %) taking clopidogrel, and ( . %) taking statins. conclusions:patients with covid- with co-morbid neurological diseases had an advanced age, a high rate of severe illness, and a high mortality rate. among the neurological symptoms, altered mental status was more common in patients with severe covid- with co-morbid neurological diseases. coronavirus disease (covid- ) due to -novel coronavirus ( -ncov) infection has caused a pandemic. [ ] [ ] [ ] from december to april , the number of covid- patients worldwide reached million, and the mortality has exceeded , . the number of patients with covid- in most regions of the world is continuing to rise rapidly. given the high infectivity of -ncov, the basic reproduction number of the infection (r ) is between . and . . the large number of patients with covid- has overwhelmed the health care system, and the mortality rate in some regions exceeds %. therefore, it is very important to study and understand this unfamiliar disease. on february , the world health organization announced that the disease caused by -ncov is named covid- . a growing body of evidence has demonstrated that covid- affects not only the respiratory system but also the digestive, urinary, haematological, circulatory, and nervous system. , [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] studies have reported neurological symptoms since the onset of covid- , including myalgia, dizziness, headache, numbness and weakness in extremities, , anosmia and ageusia, impaired consciousness, and meningeal irritation, and these symptoms may even be first-onset, solitary symptoms. mao et al. found that among patients with covid- , had neurological symptoms. clinical case studies of covid- showed that elderly patients and patients with co-morbid neurological diseases had a high rate of severe and critical illness and a high rate of mortality. , , , most patients with neurological diseases have co-morbid underlying diseases, including hypertension, diabetes mellitus, coronary heart disease, and metabolic syndrome, which in turn increase their risks of contracting -ncov, converting into severe illness, and dying. , to the best of our knowledge, except for a few case reports, there has been no clinical analysis of patients with neurological diseases and co-morbid covid- . diagnostic information of patients with covid- hospitalised in huoshenshan hospital who had completed treatment was retrieved in this study. the clinical characteristics of patients with covid- with co-morbid neurological diseases were analysed. antibiotics, corticosteroids, oxygen support, and symptomatic treatment), special treatment methods (tuzumab, convalescent plasma, stem-cell therapy, and traditional chinese medicine), prophylactic medication for cerebrovascular diseases, and clinical outcomes. this study also collected the results of laboratory tests and imaging studies first conducted within hours after admission. upon admission, subjective symptoms were provided by patients who had clear consciousness, normal cognition and psychology, and normal speech; if patients showed impaired consciousness, aphasia, or dementia, the subjective symptoms were provided by their immediate family members who had no problems in cognition, psychology, or speech, and the subjective symptoms were clarified by reviewing previous medical records and communicating directly with their close relatives and physicians. all included data were obtained with verbal consent from the patients or their family members. each patient's data were collected and cross-checked by two well-trained and experienced neurologists. in the event of any inconsistency in cross-checking the two sets of data, a third independent expert was designated to review the electronic medical records for data approval. the patients' data were analysed by a clinical research team comprising epidemiologists. this study was conducted in accordance with the principles of the helsinki declaration. this study was approved and written informed consent was waived by the huoshenshan hospital ethics committee on april . owing to enable a rapid response to the novel infectious disease and the urgent need to collect date. for continuous variables, those conforming to a normal distribution and a non-normal distribution are expressed as mean (standard deviation) and median (interquartile range [iqr]), respectively. unpaired t-tests and non-parametric tests were performed as appropriate. categorical variables are expressed as percentages and analysed using the χ test. two-sided p-values < . and < . were considered statistically significant and extremely statistically significant, respectively. data analysis was performed using the spss version . software. . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) respectively. amid the neurological co-morbidities, a previous cerebral infarction was most common, occurring in patients ( . %); dementia occurred in patients ( . %), acute cerebral infarction in ( . %), sequelae of cerebral haemorrhage in ( . %), intracranial mass lesions in ( . %), epilepsy in ( . %), parkinson's disease in ( . %), and myelopathy in ( . %). fever (n = , . %) was still the most common symptom. other common non-neurological symptoms were as follows: coughs in patients ( . %), fatigue in patients ( %), chest tightness in patients ( . %), expectoration in patients ( . %), diarrhoea in patients ( %), and dyspnoea in patients ( . %). the most common neurological symptoms were myalgia in patients ( . %), extremity paralysis in patients ( . %), and impaired consciousness in patients ( %). positive focal neurological signs occurred in patients ( . %). headache (n = , . %), nausea (n = , . %), vomiting (n = , . %), dizziness (n = , . %), altered mental state (n = , . %), and exacerbated neurological symptoms (n = , . %) all occurred in this enrolled group. the mean pulse oxygen saturation was % ( %). the mean duration from onset to cure was ( ) days. eight patients ( . %) died. disease was classified clinically as mild (n = ), moderate (n = ), severe (n = ), and critical illness (n = ). the patients with mild and moderate disease were combined into a non-severe group, totalling patients ( . %). patients with severe . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) the copyright holder for this preprint this version posted may , . . disease and critically ill patients were combined into the severe group, totalling patients ( . %). comparisons were conducted between the two groups (table ) . there was no significant difference in age between the severe group and the non-severe group ( . ( . ) vs. . ( . ) years, p = . ). there was a higher percentage of male vs. female patients in both groups: . % (n = ) vs. . % (n = ), respectively (p = . ). in terms of past medical history, there was no significant difference in either neurological diseases or other underlying diseases between the two groups. the severe group had more cases of dyspnoea ( ( . %) vs. , p = . ) and expectoration ( ( . %) vs. ( %), p = . ) than the non-severe group. per neurological symptoms, the severe group had significantly more patients with altered mental state than the non-severe group ( ( . %) vs. , p = . ), and there were no significant differences in other neurological symptoms and general symptoms between the two groups. with respect to vital signs after hospitalisation, the pulse oxygen saturation in the severe group was % ( %), significantly lower than % ( %) in the non-severe group (p < . ). the systolic and diastolic blood pressure values in the severe group ( ( ) and ( )) were lower than those in the non-severe group ( ( ) and ( )), and the difference in diastolic pressure between the two groups was statistically significant (p = . ). all patients ( . %) who subsequently died were in the severe group, and the percentage was significantly higher than that in the non-severe group (p = . ). amid the deceased patients, were critically ill before death ( / ; . %), and was severely ill ( / ; . %). . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) the copyright holder for this preprint this version posted may , . . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint table shows that out of patients ( . %) were administered nasal catheter oxygenation, ( . %) were administered face mask oxygenation, ( . %) were administered high-flow nasal catheter oxygenation, ( . %) were treated using a non-invasive ventilator, and ( . %) required mechanical ventilation with endotracheal intubation. the proportion of patients administered face mask oxygenation, high-flow nasal catheter oxygenation, and non-invasive and invasive mechanical ventilation was significantly higher in the severe group than in the non-severe group. only patient ( . %) in the non-severe group was administered face mask oxygenation and high-flow humidified oxygenation, and no patients were treated using a non-invasive ventilator or required mechanical ventilation. in the severe group, the number of patients administered face mask oxygenation, high-flow nasal catheter oxygenation, and non-invasive and invasive mechanical ventilation were ( %), ( . %), ( %), and ( . %), respectively. regarding antiviral treatment, there was no significant difference in the use of arbidol and interferon inhalation between the two groups. the proportion of patients taking antibiotics was significantly higher in the severe group than in the non-severe group . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . this study presented an analysis of the clinical characteristics of neurological diseases co-morbid with covid- in the largest sample size so far. of the patients enrolled in this study, ( . %) had severe and ( . %) had non-severe disease, and the average age was . ( . ) years. fever was still the most common non-neurological symptom. however, the proportion of patients with fever was only . %, which was significantly lower than the . %- . % reported by other studies, but similar to the proportion of . % in patients in a study by mao et al. mo et al. also found that the proportion of patients with fever in the refractory group was . %, lower than the % in the moderate group, and the average age of the refractory group was years, significantly higher than the years of the moderate group. the low occurrence of fever in this group was linked to an advanced average age, a decrease in pulmonary defence function, and a decline in systemic immunity. in this study, the proportions of patients with expectoration and dyspnoea in the severe group were significantly higher than those in the non-severe group, and moreover, the severe group had significantly higher neutrophil ratios and higher levels of infection indicators (crp, hypersensitive crp, and procalcitonin). this suggested that the severe group was likely to have a higher rate of co-morbid bacterial infections than the non-severe group, which in turn explained why there was a higher proportion of patients using mechanical ventilation, non-invasive ventilators, high-flow humidified oxygenation, face mask oxygenation, expectorants, antibiotics, and steroids in the severe group than in the non-severe group. owing to co-morbid neurological diseases, most patients had different degrees of motor dysfunction, resulting in long-term bed rest and fluid accumulation in the dorsal regions of the lungs, which was more likely to cause hypostatic pneumonia and reduce pulmonary reserve. in the meantime, bulbar palsy further exacerbated the inflammation of the lungs. in addition to the fact that patients had many co-morbidities at an advanced age, the above observations could be a main reason why there was a higher mortality rate ( . %) in patients with severe covid- with co-morbid neurological diseases than in other populations. in this study, altered mental status occurred in patients ( . %), all of whom were in the severe group. our research team has reported a case of covid- presenting with intracranial infection that resulted in mental and behavioural abnormalities. helms et al. performed head magnetic resonance imaging (mri) on patients with severe covid- and found that patients exhibited cerebral . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . leptomeningeal enhancement, and the electroencephalogram (eeg) of patient showed diffuse slow waves in the bilateral frontal lobes, which was similar to the eeg of encephalopathy. although no patient presented with meningeal irritation and obvious inflammatory changes in the cerebrospinal fluid, the above observation was still highly indicative of meningeal involvement. this suggested that -ncov could involve the central nervous system (cns), especially in patients who had blood-brain barrier breakdown due to a neurological disease. the pathophysiology pathways of -ncov involvement in the nervous system is still unclear. the viruses -ncov, sars-cov, and mers-cov are all coronaviruses, and they have highly homologous sequences. most coronaviruses have similar viral structures and transmission routes. a previous study revealed that sars-cov could invade the mouse brain tissue through olfactory epithelial cells in the nasal cavity, resulting in neuronal death. autopsy results also revealed that sars-cov existed in the cns. with the in-depth understanding of covid- , it has been reported that patients with covid- could experience olfactory and taste impairment. therefore, -ncov invasion of the brain tissue through the lamina cribrosa close to the olfactory bulb may be a pathway of cns infection. olfactory and taste impairment was not observed in the participants in the present study, suggesting that the above-mentioned pathway may not be the main pathway for the virus invasion of the cns. another possible reason for the lack of olfactory and taste impairment may be that there were few frontline neurologists and thus such symptoms were likely to be omitted. angiotensin-converting enzyme (ace ) has been identified as a functional receptor of sars-cov. it was reported that ace receptors are expressed on glial cells and neurons in the brain. hamming et al. studied ace protein localisation in different organs of the human body and found that ace existed in the endothelial cells of arteries and veins as well as arterial smooth muscle cells of all organs. it is difficult for the virus to invade the cns owing to the presence of the blood-brain barrier. however, the meninges are rich in blood vessels, which may explain why cerebral leptomeningeal enhancement appeared in patients with severe covid- in a previous report. ace receptors also appear in skeletal muscles. it was found in this study that patients ( . %) had myalgia, with no significant difference between the two groups and with lactate dehydrogenase levels in the normal range. however, the lactate dehydrogenase level was significantly higher in the severe group . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint than in the non-severe group, which further confirmed that this kind of injury may be related to ace in skeletal muscles. this result was similar to that of mao et al., but it still remains to be verified by conducting autopsies to find corresponding evidence in skeletal muscles. meanwhile, it cannot be ruled out that infection may cause an excessive immune response, damaging the nervous system, and an increase in the level of cytokines may damage skeletal muscles. recent reports have shown that -ncov infection may cause a cytokine storm, resulting in multiple organ dysfunction, including cellular immunity deficiencies, coagulation activation, myocardial injury, liver injury, and kidney injury. , , in the present study, the severe group had a decrease in red blood cells, lymphocytes, and monocytes as well as an increase in interleukin- levels, prothrombin time, and d-dimer levels. this suggested that the patients with severe disease were more likely to develop a cytokine storm, which caused coagulation activation and an increase in red blood cell destruction. the elevated d-dimer levels also increase the risk of cerebrovascular disease. although the proportion of patients taking anticoagulant drugs in the severe group was significantly higher than that in the non-severe group, the proportion of patients on prophylactic medications (anti-platelets and statins) for secondary prevention of cerebrovascular disease was very low in the patients, which may be a reason for the high incidence of acute cerebrovascular disease ( . %) and high mortality rate of patients with covid- with co-morbid neurological diseases in this study. the proportion of patients taking sedative drugs was high in the severe group, mainly because patients needed to be sedated when undergoing mechanical ventilation. after these data were excluded, there was no significant difference between the two groups. however, it should be noted that patients with neurological diseases may develop central respiratory depression, which is also a cause of increasing mortality. , limitations this study had several limitations. first, it was a retrospective study, with a sample size of only cases, which may lead to a bias in clinical observation. second, given that some patients had dementia, aphasia, and impaired consciousness, it was inevitable that some information failed to be collected. in an infectious disease hospital, the collection of neurological symptoms may be inaccurate and incomplete, and when extracting data from electronic medical records, clinical symptoms may be underestimated. for example, smell and taste impairment and peripheral nerve . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . . damage were not observed in this study. third, because there was no mri equipment in a temporarily built hospital, it was impossible to verify the affected areas in the cns, and to avoid the risk of infection, a lumbar puncture for cerebrospinal fluid analysis was rarely performed, and the diagnosis rate of intracranial infection was low. furthermore, owing to a lack of neuroelectrophysiological testing, it was impossible to determine peripheral nerve injury. fourth, the completion rate of a blood gas analysis was low in the non-severe group, and this index was removed to avoid clinical bias. finally, considering that there were only critically ill patients, their clinical characteristics were not compared with those of other patients, as the comparison would otherwise introduce inter-group bias, but this practice may prevent a deeper insight into the clinical data. in summary, patients with covid- with co-morbid neurological diseases had an advanced age, a high rate of severe illness, and a high mortality rate. regarding neurological symptoms, altered mental state was more common in patients with severe covid- with co-morbid neurological diseases. for patients with neurological diseases who have no typical respiratory symptoms, it is necessary to pay attention to neurological symptoms and the physical examination, improve nucleic acid testing for virus in a timely manner, and perform pulmonary imaging studies, which would be beneficial in the early detection of covid- and avoidance of infection. moreover, attention should be paid to the prevention and treatment of primary neurological diseases, which would help to reduce the mortality of such patients. as of april, more than million patients had been infected with covid- worldwide, and the death toll exceeded , . covid- can involve the nervous system and affect the outcome. the clinical characteristics of covid- co-morbid with neurological diseases have not yet been reported in a large sample size. patients with covid- with co-morbid neurological diseases had an advanced age, a high rate of severe illness, and a high mortality rate. it is necessary to pay attention to patients with covid- with altered mental states. . cc-by-nc-nd . international license it is made available under a is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. the copyright holder for this preprint this version posted may , . during the diagnosis and treatment of patients with covid- , it is necessary to pay attention to the history and physical exam of the nervous system as well as the prevention and treatment of primary neurological diseases. we thank all the patients and their families involved in this study, as well as many doctors, nurses, and civilians working together to fight against covid- . contributors: ry, zqy, yxw, yml, hc, zl and bz contributed equally to this paper, as did ry, zqy, hc and dwx designed the study, had full access to all data in the study, and takes responsibility for the integrity and accuracy of the data analysis. is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. (which was not certified by peer review) the copyright holder for this preprint this version posted may , . . https://doi.org/ . / . . . doi: medrxiv preprint the preparation of this manuscript; no other relationships or activities that could appear to have influenced the submitted work. ethical approval: the case series was approved by the institutional review board of the th hospital of joint logistic support force of the pla ( kyll ). written informed consent was waived owing to the rapid emergence of this infectious disease. data sharing: no additional data available. the lead author (the manuscript's guarantor) affirms that the manuscript is an honest, accurate, and transparent account of the study being reported; that no important aspects of the study have been omitted; and that any discrepancies from the study as planned (and, if relevant, registered) have been explained. no study participants were involved in the preparation of this article. the results of the article will be presented at relevant conferences. world health organization. coronavirus disease (covid- ) situation report- clinical characteristics of coronavirus disease in china the epidemiology and pathogenesis of coronavirus disease (covid- ) outbreak estimation of the transmission risk of the -ncov and its implication for public health interventions world health organization. novel coronavirus ( -ncov) situation report- covid- : gastrointestinal manifestations and potential fecal-oral transmission clinical features of patients infected with novel coronavirus in wuhan, china the neuroinvasive potential of sars-cov may play a role in the respiratory failure of covid- patients first case of novel coronavirus disease with encephalitis a new symptom of covid- : loss of taste and smell analysis of heart injury laboratory parameters in covid- patients in one hospital in wuhan clinical characteristics of deceased patients with coronavirus disease : retrospective study neurological manifestations in covid- caused by sars-cov- neurologic manifestations of hospitalized patients with coronavirus disease clinical characteristics of hospitalized patients with novel coronavirus-infected pneumonia in wuhan, china epidemiological, clinical and virological characteristics of cases of coronavirus-infected disease (covid- ) with gastrointestinal symptoms concomitant neurological symptoms observed in a patient diagnosed with coronavirus disease new coronavirus pneumonia prevention and control program clinical characteristics of refractory covid- pneumonia in wuhan, china neurologic features in severe sars-cov- infection cryo-em structures of mers-cov and sars-cov spike glycoproteins reveal the dynamic receptor binding domains severe acute respiratory syndrome coronavirus infection causes neuronal death in the absence of encephalitis in mice transgenic for human ace evidence of the covid- virus targeting the cns: tissue distribution, host-virus interaction, and proposed neurotropic mechanisms composition and divergence of coronavirus spike proteins and host ace receptors predict potential intermediate hosts of sars-cov- tissue distribution of ace protein key: cord- -up jotp authors: gillissen, adrian; ruf, bernhard r. title: das schwere akute atemwegssyndrom (sars) date: journal: med klin (munich) doi: . /s - - -z sha: doc_id: cord_uid: up jotp severe acute respiratory syndrome (sars) is a viral disease, observed primarily in southern china in november , with variable flu-like symptoms and pneumonia, in approx. % leading to death from respiratory distress syndrome (rds). the disease was spread over more than states all over the globe by sars-virus-infected travelers. who and cdc received first information about a new syndrome by the end of february , after the first cases outside the republic of china had been observed. a case in hanoi, vietnam, led to the first precise information about the new disease entity to who, by dr. carlo urbani, a co-worker of who/doctors without borders, who had been called by local colleagues to assist in the management of a patient with an unknown severe disease by the end of february . dr. urbani died from sars, as did many other health care workers. in the meantime, more than , cases have been observed worldwide, predominantly in china and hong kong, but also in taiwan, canada, singapore, and the usa, and many other countries, and more than of these patients died from rds. since the beginning of march , when who and cdc started their activities, in close collaboration with a group of international experts, including the bernhard-nocht-institute in hamburg and the department of virology in frankfurt/main, a previously impossible success in the disclosure of the disease was achieved. within only weeks of research it was possible to describe the infectious agent, a genetically modified coronavirus, including the genetic sequence, to establish specific diagnostic pcr methods and to find possible mechanisms for promising therapeutic approaches. in addition, intensifying classical quarantine and hospital hygiene measures, it was possible to limit sars in many countries to sporadic cases, and to reduce the disease in countries such as canada and vietnam. this review article summarizes important information about many issues of sars (may th, ). Übersicht der weltgesundheitsorganisation (who) und mitglied von "Ärzte ohne grenzen", der in hanoi einen erkrankten amerikanischen geschäftsmann behandelte, wurde das sars erstmals als neue lungenerkrankung beschrieben. urbani verstarb am . märz an der folgen des im rahmen der behandlung der erkrankten in hanoi erworbenen sars. am [ , ] . in kanada ist vorwiegend toronto mit fällen, davon ca. verstorbene, betroffen [ ] . alle patienten hatten engen kontakt mit sars-kranken. in toronto erkrankten häufiger ältere patienten, die z.t. an chronischen erkrankungen litten. inwieweit der immunstatus die erregerausbreitung einerseits und die individuelle krankheitsanfälligkeit andererseits fördert oder hemmt, ist gegenstand aktueller forschung [ ] . weitere fälle wurden aus vietnam, singapur und den usa (hier mation about a new syndrome by the end of february , after the first cases outside the republic of china had been observed. a case in hanoi, vietnam, led to the first precise information about the new disease entity to who, by dr. carlo urbani, a co-worker of who/doctors without borders, who had been called by local colleagues to assist in the management of a patient with an unknown severe disease by the end of february . dr. urbani died from sars, as did many other health care workers. in the meantime, more than , cases have been observed worldwide, predominantly in china and hong kong, but also in taiwan, canada, singapore, and the usa, and many other countries, and more than of these patients died from rds. since the beginning of march , when who and cdc started their activities, in close collaboration with a group of international experts, including the bernhard-nocht-institute in hamburg and the department of virology in frankfurt/ main, a previously impossible success in the disclosure of the disease was achieved. within only weeks of research it was possible to describe the infectious agent, a genetically modified coronavirus, including the genetic sequence, to establish specific diagnostic pcr methods and to find possible mechanisms for promising therapeutic approaches. in addition, intensifying classical quarantine and hospital hygiene measures, it was possible to limit sars in many countries to sporadic cases, and to reduce the disease in countries such as canada and vietnam. this review article summarizes important information about many issues of sars (may th, ). [ , ] . die letalitätsrate liegt bei ca. - %. tabelle zeigt die am häufigsten beschriebenen befunde. die cdc veröffentlichen auf ihrer internetseite die aktuellen ergebnisse von krankheitsverläufen (www.cdc.gov/ mmwr/index.html). demnach kommt es schon während der frühen erkrankungsphase zu einem anstieg der lymphozyten, im allgemeinen jedoch nicht der leukozyten. bei ca. % der erkrankten zeigen sich eine leuko-und thrombozytopenie [ , ] . obwohl die nierenund auch die leberfunktion bei den meisten patienten normal blieben, wurden passagere erhöhungen von kreatin-phosphokinase (bis iu/l) und lebertransaminasen (zwei-bis dreifach erhöhte werte) beobachtet [ ] . seit der erreger definiert ist, wurden weltweit tests zum sars-nachweis entwickelt [ ] . ein positives testergebnis wird von der who als beweis einer aktuellen und kurz zurückliegenden infektion gewertet. ein negatives ergebnis schließt ein sars allerdings nicht aus. die who empfiehlt, bei allen verdachts-und wahrscheinlichen fällen mehr als die übliche probenmenge zu sammeln und diese auch länger als üblich für evtl. spätere untersuchungen aufzubewahren. aktuell wurden folgende tests entwickelt und stehen spezialisierten labors schon zur verfügung: • mittels elisa ("enzyme-linked immunosorbent assay") oder immunofluoreszenzassay werden spezifische antikörper im serum der patienten detektiert. die tests (igg-titer-anstieg) werden erst bis zu tage initial ist der röntgen-thoraxfilm meist unauffällig. bei den meisten patienten zeigen sich jedoch früh fokale, intrapulmonale infiltrate, die sich im verlauf der krankheitsprogression flächenhaft bis zum vollbild eines ards ausweiten können [ ] . beispiele positiver röntgen-thoraxfilme im entwicklungsverlauf sind im internet abrufbar (www. droid.cuhk.edu.hk/web/atypical_pneumonia/ atypical_pneumonia.htm). wenn eine diagnostisch anderweitig erklärbare diagnose vorliegt, scheidet das sars als mögliche ursache aus [ ] coronavirus main proteinase ( cl pro ) structure: basis of design of anti-sars drugs beijing doctor alleges sars cases cover up in china centers of disease control. preliminary clinical description of severe acute respiratory syndrome epidemiological determinants of spread of causal agent of severe acute respiratory syndrome in hong kong identification of a novel coronavirus in patients with severe acute respiratory syndrome infectious diseases: deferring competition, global nets closes on sars early therapy with neuraminidase inhibitor oseltamivir maximizes its efficacy in influenza treatment guideline on management of severe acute respiratory syndrome (sars) efficacy and safety of zanamivir in patients with influenza -impact of age, severity of infections and specific risk factors sars-associated coronavirus a novel coronavirus associated with severe acute respiratory syndrome a major outbreak of severe acute respiratory syndrome in hong kong textbook of influenza sars: imaging of severe acute respiratory syndrome the use of corticosteroids in sars coronavirus as possible cause of severe acute respiratory syndrome identification of severe acute respiratory syndrome in canada comparative fulllength genome sequence analysis of sars coronavirus isolates and common mutations associated with putative origins of infection a cluster of cases of severe acute respiratory syndrome in hong kong antivirale therapie und prophylaxe der influenza severe acute respiratory syndrome (sars): infection control eine proteinaseninhibition die ausbreitung des virus im infizierten körper verhindern oder reduzieren könnte. dieses prinzip ist bei den neuraminidaseinhibitoren zur therapie der influenza bekannt und klinisch umgesetzt [ , , ] . die kristalline struktur und die bindungsstellen von cl pro sind mittlerweile bekannt. eine modifikation des strukturähnlichen und verfügbaren rhinovirus- cl pro -inhibitors wäre daher für die zukunft ein theoretisch möglicher sars-therapieansatz [ ] . das robert-koch-institut hat folgende empfehlung zum procedere bei sars-verdacht herausgegeben, welche aber kurzfristig an die aktuelle situation angepasst werden sollte (www.rki.de "hinweise für untersuchungen"; abbildung [ , ] . key: cord- -dyxfsojh authors: ahamad, shakir; branch, scotty; harrelson, shea; hussain, mohd kamil; saquib, mohammad; khan, saeed title: primed for global coronavirus pandemic: emerging research and clinical outcome date: - - journal: eur j med chem doi: . /j.ejmech. . sha: doc_id: cord_uid: dyxfsojh the global effort to combat and contain the coronavirus disease (covid- ) pandemic is now proceeding on a war footing. the world was slow to react to the developing crisis, but once the contours of the impending calamity became evident, the different state and non-state actors have raced to put their act together. the covid- outbreak has blatantly exposed the shortcomings of our healthcare system and the limitations of medical science, despite considerable advances in recent years. to effectively tackle the current epidemic, almost unprecedented in the modern era, there is an urgent need for a concerted, sustained, and coordinated effort towards the development of new diagnostics, therapeutic and vaccines, and the ramping up of the healthcare infrastructure, especially in the poorer, underprivileged nations. towards this end, researchers around the world are working tirelessly to develop new diagnostics, vaccines, and therapeutics. efforts to develop a vaccine against covid- are presently underway in several countries around the world, but a new vaccine is expected only by the end of the year-at the earliest. new drug development against covid- and its approval may take even longer. under such circumstances, drug repurposing has emerged as a realistic and effective strategy to counter the virus menace in the short run, and several antiviral and antimalarial medicines are currently in different stages of clinical trials. researchers are also experimenting with nutrients, vitamins, monoclonal antibodies, and convalescent plasma as immunity boosters against the severe acute respiratory syndrome coronavirus- (sars-cov- ). this report presents a critical analysis of the global clinical trial landscape for covid- with an emphasis on the therapeutic agents and vaccines currently being tested at pandemic speed. where the virus originated, to check whether this vaccine could stimulate antibody production and boost immunity against sars-cov- . in preclinical studies, ad -ncov showed an acceptable safety profile and generated a robust immune response in animal models. currently, a randomized, double-blinded, and placebo-controlled phase-ii clinical study with ad -ncov (registered on april , ) is ongoing. this trial will evaluate the immunogenicity and safety of ad -ncov in healthy adults over years of age (table , entry ).[ - ] the university of oxford's jenner institute, along with the oxford vaccine group, has also developed a single-dose vaccine, chadox ncov- , from a non-replicating adenovirus calmette-guérin (bcg) vaccine to protect people against covid- are also advancing in six countries (table , entry ). the institute of biotechnology, amms, china, registered a randomized, double-blind, placebo-controlled phase-ii clinical trial of recombinant novel coronavirus ( -ncov) vaccine (adenovirus vector) in healthy adults aged and above on april , , (table , entry ). the same day, biontech and pfizer also secured approval from the german the use of a vaccine to prevent covid- and tackle pandemic could be a potential fool proof strategy. however, effective vaccine development for clinical use is projected to take about - months or even more, a significant drawback in this time of crisis. similarly, the discovery, development, and approval of a new drug against covid- would take even interestingly, these results concur with the findings from earlier studies on favipiravir, which reported a reduction in the disease duration from days to - days and faster recovery from covid- disease. [ ] table and figure - , which are currently undergoing clinical validation (table , entries - ). (table entry - ) . vitamins and other cofactors play a significant role in boosting the immune system. therefore, clinical trials with vitamin a, b, c, d, and e, and cofactors lipoic acid, calcifediol, and zinc are currently underway for the treatment of covid- (table entry - ). another recent study using teicoplanin ( ) the species severe acute respiratory syndrome-related coronavirus: classifying -ncov and naming it will novel virus go pandemic or be contained? influenza: the mother of all pandemics a pneumonia outbreak associated with a new coronavirus of probable bat origin johns hopkins hospital and medicine, coronavirus covid- global cases by the global patterns in coronavirus diversity genetic recombination, and pathogenesis of coronaviruses how did coronavirus start and where did it come from? was it really wuhan's animal market? mystery deepens over animal source of coronavirus an updated estimation of the risk of transmission of the novel coronavirus ( -ncov) covid- : one 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media briefing on epidemiologic features and clinical course of patients infected with sars-cov- in singapore role of lopinavir/ritonavir in the treatment of sars: initial virological and clinical findings a systematic review on the efficacy and safety of chloroquine for the treatment of covid- hydroxychloroquine and azithromycin as a treatment of covid- : results of an open-label non-randomized clinical trial remdesivir and chloroquine effectively inhibit the recently emerged novel -ncov) in vitro a pilot study of hydroxychloroquine in treatment of patients with moderate covid- efficacy of hydroxychloroquine in patients with covid- : results of a randomized clinical trial outcomes of hydroxychloroquine usage in united states veterans hospitalized with covid- observational study of hydroxychloroquine in hospitalized patients with covid- covid- ) update: fda revokes emergency use authorization for chloroquine and hydroxychloroquine advances in the development of nucleoside and nucleotide analogues for cancer and viral diseases early peek at data on gilead coronavirus drug suggests patients are responding to treatment remdesivir for the treatment of covid- - gilead announces results from phase trial of investigational antiviral remdesivir in patients with severe covid- gilead's suspension of covid- trials in china should serve as a bellwether for studies in other countries montefiore and einstein assess remdesivir and baricitinib for covid- global coronavirus covid- clinical trial tracker remdesivir in covid- : a critical review of pharmacology, pre-clinical and clinical studies [ ] recovery: randomized evaluation of covid- therapy covid- treatments could be fast-tracked through new national clinical trial initiative effect of dexamethasone in hospitalized patients with covid- : preliminary report world health organization. who director-general's opening remarks at the media briefing on covid- covid- : demand for dexamethasone surges as recovery trial publishes preprint world first coronavirus treatment approved for nhs use by government coronavirus | government approves use of dexamethasone bergenbio starts bemcentinib dosing in accord trial major covid- drugs trial begins in namilumab and infliximab selected for catalyst trial in uk vidofludimus calcium, a next generation dhodh inhibitor for the treatment of relapsing-remitting multiple sclerosis receives first regulatory approval from german health authority bfarm to initiate a phase clinical trial of its selective oral dhodh inhibitor german-health-authority-bfarm-to-initiate-a-phase- -clinical-trial-of-its-selective-oral- dhodh-inhibitor-imu- -in-covid- -patients germany's bfarm approves covid- trial by immunic announces first patients dosed in its phase , calvid- clinical trial of imu- in covid- austrian coronavirus adaptive clinical trial (covid- ) (acovact) a human monoclonal antibody blocking sars-cov- infection cryo-em structure of the -ncov spike in the prefusion conformation antigenicity of the sars-cov- spike glycoprotein unexpected receptor functional mimicry elucidates activation of coronavirus roche initiates phase iii clinical trial of actemra/roactemra plus remdesivir in hospitalised patients with severe covid- pneumonia genentech announces fda approval of clinical trial for actemra to treat hospitalized patients with severe covid- pneumonia effective treatment of severe covid- patients with tocilizumab meplazumab treats covid- pneumonia: an open-labelled, concurrent controlled add-on clinical trial first clinical use of lenzilumab to neutralize gm-csf in patients with severe covid- pneumonia a novel protein drug, novaferon, as the potential antiviral drug for covid- atyr pharma announces phase study of atyr in covid- patients with severe respiratory complications following fda acceptance of ind application atyr-pharma-announces-phase- -study-of- atyr -in-covid- -patients-with-severe-respiratory-complications following-fda-acceptance-of-ind-application.html how blood from coronavirus survivors might save lives the convalescent sera option for containing covid- us study finds convalescent plasma safe for covid- patients trials of mw vaccine for treatment of covid- patients to be initiated soon discovering drugs to treat coronavirus disease sars-cov- rna dependent rna polymerase (rdrp): a molecular docking study an orally bioavailable broad-spectrum antiviral inhibits sars-cov- in human airway epithelial cell cultures and multiple coronaviruses in mice broad spectrum antiviral agent niclosamide and its therapeutic potential specific plant terpenoids and lignoids possess potent antiviral activities against severe acute respiratory syndrome coronavirus skp attenuates autophagy through beclin -ubiquitination and its inhibition reduces mers-coronavirus infection identification of antiviral drug candidates against sars-cov- from fda-approved drugs inhibition of severe acute respiratory syndrome coronavirus replication by niclosamide tilorone: a broad-spectrum antiviral invented in the usa and commercialized in russia and beyond htcc as a highly effective polymeric inhibitor of sars-cov- and mers-cov sars-cov- infected host cell proteomics reveal potential therapy targets brilacidin: background and scientific rationale for brilacidin as a potential novel covid- ) treatment vanquishing the virus: + covid- drug and vaccine candidates in teicoplanin potently blocks the cell entry of -ncov teicoplanin: an alternative drug for the treatment of covid- ? discovery and development of safe-in-man broad-spectrum antiviral agents the fda-approved drug ivermectin inhibits the replication of sars-cov- in vitro auranofin: repurposing an old drug for a golden new the fda- approved gold drug auranofin inhibits novel coronavirus (sars-cov- ) replication , virology drug evaluation: apilimod, an oral il- /il- inhibitor for the treatment of autoimmune diseases and common variable immunodeficiency characterization of spike glycoprotein of sars-cov- on virus entry and its immune cross-reactivity with key: cord- -kejcwlng authors: akbari, hamed; tabrizi, reza; lankarani, kamran b.; aria, hamid; vakili, sina; asadian, fatemeh; noroozi, saam; keshavarz, pedram; faramarz, sanaz title: the role of cytokine profile and lymphocyte subsets in the severity of coronavirus disease (covid- ): a systematic review and meta-analysis date: - - journal: life sci doi: . /j.lfs. . sha: doc_id: cord_uid: kejcwlng aims: this study aimed to make a comparison between the clinical laboratory-related factors, complete blood count (cbc) indices, cytokines, and lymphocyte subsets in order to distinguish severe coronavirus disease (covid- ) cases from the non-severe ones. materials and methods: relevant studies were searched in pubmed, embase, scopus, and web of science databases until march , . cochrane's q test and the i( ) statistic were used to determine heterogeneity. we used the random-effect models to pool the weighted mean differences (wmds) and % confidence intervals (cis). key findings: out of a total of initial records, articles ( studies) with patients (ranging from to ), were included. our meta-analyses with random-effect models showed a significant decrease in lymphocytes, monocyte, cd + t cells, cd + t cells, cd cells, cd cells, and natural killer (nk) cells and an increase in the white blood cell (wbc), neutrophils, neutrophil to lymphocyte ratio (nlr), c-reactive protein (crp)/hs-crp, erythrocyte sedimentation rate (esr), ferritin, procalcitonin (pct), and serum amyloid a (saa), interleukin- (il- ), il- r, il- , il- , il- , il- , tumor necrosis factor-alpha (tnf-α), and interferon-gamma (inf-γ) in the severe group compared to the non-severe group. however, no significant differences were found in il- β, il- , and cd /cd t cell ratio between the two groups. significance: decrease in total lymphocytes and lymphocyte subsets as well as the elevation of crp, esr, saa, pct, ferritin, and cytokines, but not il- β and il- , were closely associated with covid- severity, implying reliable indicators of severe covid- . j o u r n a l p r e -p r o o f j o u r n a l p r e -p r o o f pediatric or pregnant cases due to the diverse presentation of covid- in these groups, ( ) inadequate information on inflammatory-related laboratory parameters in either severe or non-severe disease groups, ( ) coronavirus strains other than covid- , ( ) and studies with unusable data. nonetheless, the diagnostic criteria for covid- were explained on the basis of laboratory approved sars-cov- infection. if two or more studies were published by the same authors or institutions, only the study having the largest sample size was selected. the data from the incorporated studies were extracted by two reviewers (sv and sf) independently. also, a third reviewer (rt) was used to solve any arisen argument. the details of each study were collected which involve author, publication date, study location, study design, sample size, sample characteristics (age, gender, comorbidities), exposure characteristics (study definition of severity of covid- , the timing of classification of disease severity [on admission or otherwise], number of cases with non-severe covid- , number of cases with severe or critical covid- ) , the timing of blood sample collection (on admission or otherwise). moreover, inflammatory-related laboratory factors, cytokines, lymphocyte subsets, and cbc indices were grouped by covid- severity (mean [sd] ) and finally, all the extracted data were transferred into microsoft excel. furthermore, through re-checking the primary studies, as well as discussions, any inconsistencies in the extracted data were resolved. it is worth mentioning that, using web plot digitizer online software, some graph data were converted to numerical data (https://apps.automeris.io/wpd/). in case the relevant data were missing, authors of selected studies were contacted via email. also, it should be noted that due to inaccuracies in the research methodology for some of the studies, we reported the type of study in some articles, especially those submitted in the medrxiv, by inference. the included studies differed in the way they defined patients' disease status, and classified the disease into 'mild, moderate, severe and critical', 'ordinary and severe/critical', 'common and severe', and 'non-severe and severe', categories. the first outcome measure adopted was severe (including both severe and critical cases) vs. non-severe disease. the definition provided j o u r n a l p r e -p r o o f patients as shown by their symptoms and the imaging examination; ( ) severe: patients with any of the following factors: (i) respiratory rate equal to or higher than /min; (ii) resting pulse oxygen saturation (spo ) equal to or lower than %; (iii) oxygen partial pressure (pao ) / fraction of inspired oxygen (fio ) equal to or lower than mmhg ( mmhg = . kpa); (iiii) imaging process showing a % progression in multiple pulmonary lobes of a lesion in hours; ( ) critical: patients with any of the following factors: (i) the need for mechanical ventilation in case of respiratory failure (ii) shock; (iii) admission to the intensive care unit (icu) due to simultaneous failure in another organ. it is noteworthy that mild or moderate patients were included in the non-severe group, while severe or critical patients were included in the severe one. the newcastle-ottawa scale was used to evaluate quality, and moreover, assessment scores of - , - , and - represented poor, fair, and good studies, respectively. additionally, discrepancies were resolved through consensus. all statistical analyses were conducted using stata version . (stata corp., college station, tx). laboratory factors were considered as the mean (sd) difference with % confidence intervals (cis) between the severe group and the non-severe group. to pool the mean differences (sd), weighted mean difference (wmd) statistic with the random-effect model (dersimonian-laird method) were used. cochrane's q test or the i statistic was used to assess heterogeneity among included studies. i above % and cochrane's q test with p < . was considered as the existence of significant heterogeneity. sensitivity analysis was used to evaluate the robustness of meta-analyses findings with applying the leave-one-out method after removing one by one included study on the pooled wmds. egger regression and begg's rank correlation tests were applied to detect the potential evidence of publication bias between included studies. we yielded a total of records through initial online search in databases. of these, were duplicate. after screening based on title and abstract, articles were selected as the candidates for assess according to inclusion and exclusion criteria. finally, articles ( studies) were identified to be eligible for current meta-analysis. figure shows the flowchart of study identification and selection process. all selected studies contained a total of (ranging from to ) patients including in the severe group and in the non-severe group. forty-three of all included articles were conducted in china and one [ ] of them was performed in usa. most of assessments on laboratory tests among included patients were conducted on admission period/before treatment. the characteristics of included studies are summarized in table . using random-effects model, our meta-analyses indicated a significant decrease in the wmd of the pooled finding on cytokines showed a significant increase in the wmd of il we found no significant differences between the pre-and post-sensitivity pooled effect sizes by , [ ] the study on tnf-α (wmd= . pg/ml, %ci: - . , . ), the sensitivity findings showed that there was a significant differences between pre-and post-sensitivity pooled wmd for these outcomes. potential publication bias across included studies was examined using the egger's regression crp/hs-crp and esr have been found to be increased in a vast number of inflammations/infections [ , ] . in this new pandemic pneumonia, the levels of crp and esr significantly increased in severe cases compared to non-severe covid- patients [ , ] , which greatly coincides with those found in the present systematic review and meta-analysis. in the present study, pct concentrations were significantly higher in severe/critical patients than in non-severe cases. as it was previously shown, that is, the pct does not increase with virus infections, it may indicate superimposed bacterial infection for the critically ill patients [ , ] . saa, another important factor capable of improving inflammatory response through activation of chemokine and induction of chemotaxis even at a very low concentration [ ] , was found to have elevated circulating levels in severe patients and both were significantly related to covid-j o u r n a l p r e -p r o o f apoptotic factor [ ] . consequently, these inflammatory-related factors might function as a biomarker to monitor the progression of respiratory diseases. the higher level of il- in covid- patients is possibly indicative of t cell activation. an important pro-inflammatory cytokine, il- can put an end to the activation of normal t cells, which may be a reason for the presence of lymphopenia. a study carried out by gong et al. [ ] showed that although levels of il- r and il- were associated with the severity of the disease, and il- ) that suppress inflammation; a finding which coincides with that of the present study but differs from sars-cov infection [ ] . an important anti-viral cytokine generated by cd + t cells, cd + t cells, nk cells, and macrophages, ifn-γ has been reported to contribute to the cytokines storm in sars patients [ , ] . the present study showed that levels of ifn-γ in severely infected cases were higher than those of the non-severe covid- patients, suggesting that ifn-γ may efficiently indicate the status of the disease. besides, il- β and il- were found not to be significantly associated with the covid- severity, which might be due to few numbers of included studies, hence the need for further studies to explain the role that these cytokines play in the progression of the disease. albeit no significant association was found by some studies between the covid- pneumonia severity and il- , il- , and tnf-α, this systematic review and meta-analysis indicated that il- , il- , and tnf-α could be used to assess the severity of covid- and that they might be potential targets for immunotherapy of covid- . the association found between lymphopenia and severity of the covid- implies that, as does sars-cov, sars-cov- might act on lymphocytes, especially t types, hence possibly leading to the depletion of cd + t and cd + t cells [ ] . the exhaustion of cd + t cells in severe patients j o u r n a l p r e -p r o o f may reduce their cellular immune response to sars-cov- . the study conducted by li et al. [ ] showed that these multi-functional cd + t cells were much frequently seen in patients severely [ , ] . the elevated levels of nlr found in the present study suggest that the internal environment was seriously disturbed and that the severely infected cases were in a potentially critical condition. liu et al. [ ] revealed that the area under curve (auc), c-index, sensitivity, and specificity for nlr were at a high level, suggesting that nlr is a reliable index of predicting the incidence of severe illness in an early time. these results point out that the easily accessible tests are potentially easy-to-use, low-cost for early screening and prognosis of the severe and/or critical covid- infected cases. whereas covid- was initially recognized as a pulmonary disease followed by a storm of pro-inflammatory cytokines, resulting in ards, mods, and death [ ] , recent evidence indicates that the disease is a systemic disorder affecting many organ systems including kidneys, gastrointestinal tract, liver, nervous system, and skin among others [ ] [ ] [ ] [ ] . the mechanism of these systemic effects is not clear yet and many might be related or mediated by the effects of the cytokines and dysregulated immune system [ ] . there are several limitations for this review. as most of the evidence came from china, this lack of evidence from outside china might be a limitation in the way of generalizing our results, particularly with regard to the shortage of costly laboratory tests in the context of nations with low resources. the heterogeneity of the included studies was another limitation, and the need for further studies is greatly felt. one of the main causes might be the poor description of the analytical performance features of the methods applied among the included studies. furthermore, aging as a condition related to the inflammation was the basis of the previous studies [ , ] , showing that severe patients were older than non-severe ones. since the proinflammatory response is believed to initiate sars-cov- infection, it is logically possible that the aged cases have an overwhelming inflammatory reaction. additionally, as the age of some of the patients were not included in some included studies, the comparison of the age differences between the two groups was not possible, which would be another probable limitation. accordingly, the findings showed that more comprehensive clinical studies are covid- is considered as a global health threat, consequently it is essential that clinicians have access to reliable quick pathogen tests and feasible differential diagnoses based on the clinical descriptions in their first contact with suspected patients. although it has not been witnessed that pro-inflammatory cytokines and chemokines are directly involved in lung pathology during covid- , the changes in laboratory parameters, including the reduced total lymphocytes, lymphocyte subsets, and the elevated nlr, il- , il- , il- , il- , and tnf-α, as well as the routine inflammatory-related parameters in infected patients were remarkably associated with the severity of the disease. likewise, irrespective of the crucial role that hyper-inflammatory responses play in covid- pathogenesis, there could be a protective role for the innate immune system. clinical display, 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wilson title: psycho-neuroendocrine-immune interactions in covid- : potential impacts on mental health date: - - journal: front immunol doi: . /fimmu. . sha: doc_id: cord_uid: bqh jkds coronavirus disease (covid- ) is caused by the severe acute respiratory syndrome coronavirus (sars-cov- ). the impacts of the disease may be beyond the respiratory system, also affecting mental health. several factors may be involved in the association between covid- and psychiatric outcomes, such as fear inherent in the pandemic, adverse effects of treatments, as well as financial stress, and social isolation. herein we discuss the growing evidence suggesting that the relationship between sars-cov- and host may also trigger changes in brain and behavior. based on the similarity of sars-cov- with other coronaviruses, it is conceivable that changes in endocrine and immune response in the periphery or in the central nervous system may be involved in the association between sars-cov- infection and impaired mental health. this is likely to be further enhanced, since millions of people worldwide are isolated in quarantine to minimize the transmission of sars-cov- and social isolation can also lead to neuroendocrine-immune changes. accordingly, we highlight here the hypothesis that neuroendocrine-immune interactions may be involved in negative impacts of sars-cov- infection and social isolation on psychiatric issues. in december , a new outbreak of severe acute respiratory syndrome (sars) emerged in wuhan, china. caused by severe acute respiratory syndrome coronavirus (sars-cov- ), the coronavirus disease (covid- ) caused a national outbreak of severe pneumonia in china and quickly spread worldwide. according to the world health organization (who) official website, on may th, , , , people have been tested positive for sars-cov- infection and , deaths have resulted from sars-cov- worldwide ( ). the disease, initially restricted to china, is now a pandemic, comprising all continents so far except for antarctica, thus having become a major planetary health issue ( ) . the most common symptoms of covid- are fever, cough, dyspnea, sputum production, myalgia, headache, diarrhea, rhinorrhea, anosmia, and ageusia ( , ) . nevertheless, symptoms of post-traumatic stress disorder (ptsd), anxiety and depression have also been prevalent in patients infected with covid- ( , ) . besides, sars-cov- rna was detected in the cerebrospinal fluid of a patient ( ) and increasing evidence points out that coronaviruses (covs) may invade the central nervous system (cns) ( ) . thus, we describe here the likely routes by which sars-cov- can invade the brain. since covid- is associated with increased levels of pro-inflammatory cytokines ( ) , an immune signature shared with several psychiatric disorders, we propose how the relationship between sars-cov- /host can possibly impair interactions between the immune, nervous and endocrine systems, leading to psychiatric symptoms. furthermore, once millions of people worldwide are isolated in quarantine to minimize the transmission of sars-cov- ( ), we also discuss herein evidence on the negative impacts of social isolation measures upon mental health, gathering evidence that explains how social isolation can also lead to neuroendocrine-immune changes, impairing mental health. accordingly, it is likely that both sars-cov- infection and social isolation epidemiological measures to contain the pandemic can lead to changes in psychoneuroendocrine-immune circuits with impact on the appearance and/or evolution of mental health impairments in infected subjects, as well as in those individuals that, even though not being infected, are subjected to social isolation due to one or more risk factors. finally, we provide some suggestions for how future research could confirm the hypotheses outlined here, as well as intervention strategies that mitigate the impact of covid- pandemic on mental health. coronaviruses (covs) comprise a large enveloped nonsegmented positive-sense rna virus, which belong to the family coronaviridae, within the order nidovirales ( ). they are classified in four genera, namely alphacoronavirus, betacoronavirus, gammacoronavirus, and deltacoronavirus, based on their phylogenetic relationships and genomic structures ( ) . the α-cov and β-cov are able to infect mammals, whereas the γ-cov and δ-cov tend to infect birds ( ) . previously, six covs have been identified as capable of infecting humans (human coronaviruses-hcovs): α-cov hcov-nl and hcov- e, and β-cov hcov-oc , hcovhku , middle east respiratory syndrome coronavirus (mers-cov), and severe acute respiratory syndrome coronavirus (sars-cov). the last two hcovs are considered the most lethal among them. however, the novel sars-cov- has shown a mortality rate that is presently also expressive ( ) . sars-cov- infection leads to a clinical picture characterized by highly lethal pneumonia with symptoms similar to those reported for sars-cov and mers-cov ( ) . genomic analysis show that sars-cov- shares highly homological sequence with sars-cov ( ) . although the existence of more than one receptor for this virus cannot be excluded by now, evidence so far reveals that sars-cov- enters human host cells using the same receptor of sars-cov, the human angiotensin-converting enzyme (hace ) ( ) . consequently, most of the infection mechanisms detailed for sars-cov could be applied to this novel virus. hcovs may enter the cns through distinct routes: hematogenous and/or neuronal retrograde dissemination ( ) . the neuronal route can occur through at least two different pathways: (a) via olfactory nerves and/or (b) via enteric nervous system ( , ). an experimental study using k -hace transgenic mice for the expression of hace (i.e., human sars-cov receptor) showed that sars-cov, when given nasally, could invade the brain, likely via the olfactory nerves ( ) . however, the non-expression of ace in neurons in the olfactory system ( , ) leads to question whether this is really a possible route for sars-cov- entry into cns, although it is not yet possible to rule out the possibility that other ace -independent mechanisms are involved in the entry of sars-cov- into host cells. by contrast, ace expression is abundant in small intestine endothelial cells ( ) , which connect with neurons in the enteric nervous system. in addition, gastrointestinal symptoms are commonly seen in a part of patients with covid- ( , , ) and sars-cov- was isolated from oral and anal swabs of these patients ( ) . in this way, the enteric nervous system, via the vagus nerve, can also be a possible pathway for sars-cov- to enter the cns. similarly, the hematogenous route can occur by at least two mechanisms: (a) through infected leukocytes that cross the blood-brain barrier carrying the virus to the brain and/or (b) through direct infection of brain microvascular endothelial cells, which express ace ( ) . nonetheless, the hematogenous route does not seem to be involved in the cns invasion by sars-cov, since virtually no viral particles were detected in non-neuronal cells of the infected brain areas in the early stage of infection ( ) ( ) ( ) . yet, the precise route(s) by which sars-cov enters the cns remain(s) to be determined. the recent sars-cov- rna detection in the cerebrospinal fluid of a patient with covid- ( ), as well as its similarities with the sars-cov, emphasizes the need to conduct studies aiming at evaluating the neuroinvasive potential of sars-cov- in animal models and humans. sars-cov genomic sequences in human brain tissues were found mainly in neurons of the cerebral cortex and hypothalamus, but not in the cerebellum ( , ) . however, pre-clinical studies with k -hace mice infected by sars-cov revealed viral particles during acute phase in other brain regions besides the cortex and hypothalamus, such as cerebellum, midbrain (e.g., dorsal raphe and substantia nigra), thalamus, amygdala, hippocampus, basal ganglia (e.g., caudate-putamen and nucleus accumbens), cortex (e.g., frontal, infralimbic, and cingulate), and olfactory bulb ( , ) . in these animals, a rapid spread throughout the brain was accompanied by significant neuronal loss in the cingulate and infralimbic cortices and the anterior olfactory nucleus ( ) . interestingly, high levels of cytokines and chemokines, most notably interleukin- (il- ) and interferon gamma (inf-γ) were found in brain of k -hace transgenic mice infected by sars-cov ( , ) . rather surprisingly, minimal signals of local inflammation were observed, and apoptotic or necrotic cells were not detected ( ) . considering the high-expression of inflammatory mediators along with a lack of other inflammatory signals, how sars-cov can be leading to neuronal death remains unknown. cell death non-inflammatory processes, such as autophagy, may be an explanation ( ) . since autophagy is related to several neurodegenerative and psychiatric diseases ( ) , evaluating whether infection by sars-cov- can lead to neuronal death by autophagy may also be important for future relationships between sars-cov- infection and mental health outcomes. several studies have demonstrated psychiatric manifestations in patients with mers or sars during the acute phase, such as increased stress levels, impaired memory, symptoms of depression, anxiety, ptsd, psychoses, and suicidal behavior ( ) ( ) ( ) ( ) ( ) ( ) . long-term damage has also been seen in these patients. survivors of sars, months or years after the acute phase of the infection, may also exhibit impaired memory, sleep disturbances, increased levels of stress, depression, anxiety, and ptsd symptoms ( , ( ) ( ) ( ) ( ) ( ) . to date, few studies have evaluated the possible mental health outcomes of sars-cov- infection. however, corroborating the data observed in patients with sars, a study recently demonstrated a prevalence of . % of ptsd symptoms in patients with covid- during acute phase ( ) . another study reported a prevalence of . and . % of anxiety and depression symptoms, respectively, in patients with covid- ( ). taken together, these data indicate that infection with these hcov, especially sars-cov- , can yield a negative impact on mental health, both in the short-and longterm time windows. supplementary table summarizes studies that reported mental health outcomes in patients with mers, sars, or covid- . many factors can influence the results of studies that have reported symptoms or development of psychiatric disorders in patients with mers, sars, or covid- . among them (a) the work directly with health care, (b) the presence of family history of psychiatric illnesses, (c) less social support, (d) older age, (e) the isolation, and (f) the use of high doses of steroids during the acute phase (see supplementary table ) . however, some patients who survived sars displayed psychiatric manifestations that appear to be disproportionate to the extent of lung infection or expected side effects of corticosteroid therapy ( , , ) . furthermore, it has been reported that one patient developed progressive neurological symptoms starting at day after the onset of the disease. this patient eventually died due to the sars-cov infection, and an autopsy revealed the presence of the virus in the brain, together with neuronal necrosis, glial hyperplasia, and edema ( ) . although the studies cited above have been conducted with small samples of patients, they suggest that the psychiatric manifestations seen in at least some patients might be a direct effect of the infection of sars-cov. also, studies with humans are important to evaluate and highlight the possible psychiatric outcomes in patients with sars-cov- infection. a "cytokine storm" has been proposed as a key mechanism in the sars-cov- pathophysiology and related to lung damage and lethality observed in patients bearing covid- ( ) . accordingly, increased circulating levels of several cytokines have been found in patients with mers, sars, or covid- (see table ). interestingly, high levels of pro-inflammatory cytokines (e.g., il- and inf-γ) were also found in the cns of k -hace transgenic mice infected by sars-cov ( , ) . this evidence supports the existence of an immune signature characterized by increased levels of pro-inflammatory cytokines involved in the pathophysiology of different pathogenic sars-cov in humans. furthermore, higher serum levels of pro-inflammatory cytokines (e.g., il- and ifn-γ) and chemokines were found in sars patients with severe disease, as compared to individuals with uncomplicated sars ( ) ( ) ( ) . recently, dysregulation of the immune response similar to sars-cov infection has been observed in patients with sars-cov- in wuhan (china). particularly, a significant increase in the serum levels of several pro-inflammatory cytokines, or corresponding cytokine receptors, in severe patients (n = ) than the non-severe ones (n = ), including il- , tumor necrosis factor alpha (tnfα) and interleukin- receptor (il- r) ( ) . similarly, intensive care unit (icu) patients (n = ) with severe sars-cov- infection displayed higher plasma levels of cytokines, such as il- and tnf-α, when compared with non-icu patients (n = ) ( ) . a previous study identified psychiatric manifestations (e.g., psychosis, cognitive impairments, depression, and anxiety symptoms) in patients during the acute phase of sars-cov infection ( ) . the authors also found an association between the severity of symptoms and some psychiatric outcomes. if the increase in cytokine levels and the manifestation of psychiatric symptoms are related to the severity of the symptoms of sars-cov infection, the "cytokine storm" might also be related to the "mental health thunderstorms" seen in patients with covid- ? accordingly, a possible mechanism concerning the relationship between sars-cov- infection and mental health outcomes is the involvement of neuroimmune networks. table shows that increased levels of various cytokines can be seen in several psychiatric disorders, an immune signature shared with the sars-cov- infection. soluble cytokines that reach the brain, or corresponding local altered levels can influence synthesis, release and reuptake of several neurotransmitters, including monoamines, such as dopamine, norepinephrine, and serotonin ( ) . changes in the metabolism of neurotransmitters are involved in the pathophysiology of various psychiatric disorders, such as depression, anxiety, ptsd, and obsessive-compulsive disorder ( , ) . since changes in cytokine levels can lead to a disruption in the metabolism of neurotransmitters, triggering behavioral deficits, we hypothesize than the immune system can be placed as a link between sars-cov or sars-cov- infection and mental health impairments. evidence shows that cytokines also play a key role in learning and memory processes. in healthy conditions, an increase in gene il- n = icu vs. n = healthy n = severe vs. n = moderate n = critical > n = severe > n = mild n = / icu n = severe vs. n = mild n = severe vs. n = non-severe n = severe vs. n = moderate n = spo < % vs. n = spo ≥ % il- β n = infected vs. n = healthy n = severe vs. n = moderate tnf-α n = infected vs. n = healthy n = icu vs. n = non-icu n = severe vs. n = moderate n = critical vs. n = severe vs. n = mild n = severe vs. n = non-severe n = severe vs. n = moderate il- n = infected vs. n = healthy n = icu vs. n = non-icu n = severe vs. n = moderate n = critical vs. n = severe vs. n = mild n = / icu n = severe vs. n = non-severe n = severe vs. n = moderate n = spo < % vs. n = spo ≥ % il- n = icu vs. n = healthy n = icu vs. n = non-icu n = severe vs. n = non-severe il- r n = severe vs. n = moderate n = critical > n = severe > n = mild n = severe vs. n = moderate expression of il- β, il- receptor antagonist, il- , and il- occurs in hippocampus during long term potentiation (ltp), a process considered to underlie certain forms of learning and memory ( ) ( ) ( ) . while il- β is related to ltp maintenance, acquisition of learning and memory consolidation, il- has opposite effects. however, during peripheral and central diseases in which the brain levels of il- β and il- are increased, both cytokines tend to inhibit the synaptic plasticity, learning, and memory ( ) . importantly, high levels of il- were found in blood of sars-cov and sars-cov- infected patients (see table ), as well as in cns of k -hace transgenic mice infected by sars-cov ( , ) . impaired memory has also been observed in both acute and convalescent phases of sars infection in humans (see supplementary table ). therefore, it is possible that the increased levels of il- are related to the cognitive impairments observed in sars patients. such issue should be evaluated in future studies. interleukin- is a well-known pleiotropic cytokine expressed in low levels in healthy individuals, in the presence of homeostasis alterations it becomes higher and rapidly detected, and even after stress agent removal, its levels can be maintained elevated and cause diseases ( , ) . accordingly, a dysregulation of this cytokine expression counts for the development of psychiatric disorders ( ) , as seen in table . recently, gao et al. ( ) showed increased levels of cytokines in patients with sars-cov- , especially il- , which seems to be directly related to the severity of the disease. evaluating the blood parameters of adult patients positive to sars-cov- and subdivided in groups (mild and severe) they found a significant increase in the combined detection of il- and d-dimer specially in the severe cases, pointing out the il- and d-dimer combination as a potential biomarker to identify early stages or the prognosis of the covid- disease ( ). in another study, patients were subdivided in three groups (mild, severe, and critical) and had hematological parameters followed up during disease evolution. it was shown that the more severe the case was, the higher was the il- level ( ). liu et al. demonstrated that not only increased levels of il- related to the severity of covid- , but also that decreased levels of il- were positively correlated with the treatment effectiveness and remission of the disease ( ) . in this sense, the humanized anti-interleukin- -receptor (il- r) monoclonal antibody (tocilizumab), a drug used against rheumatoid arthritis ( ) that inhibits il- signaling, has been administered experimentally in treatment of covid- ( ) . the retrospective evaluation of patients demonstrated that tocilizumab was able to improve the respiratory function and restored the levels of lymphocytes in the blood, which can be promising ( ) . in a second vein, a meta-analysis study pointed out that treatment with anti-cytokine drugs, including tocilizumab, may have an antidepressant effect ( ) . accordingly, we can conceive that this type of treatment may represent a promising therapeutic alternative to be attempted in humans, not only has beneficial effects for respiratory symptoms associated with covid- , but also for possible depressive symptoms related to the disease. thus, it would be interesting for future clinical studies to evaluate the effects of tocilizumab and other pharmacological treatments not only on symptoms and tests related to respiratory and immune functions, but also on the psychiatric symptoms. it is important to notice that some individual biological characteristics associated with impaired immunity may influence not only the natural history of covid- , but also the associated psychiatric outcomes. in this context, obesity, which is linked with systemic inflammation and impaired immunity, can increase vulnerability for covid- ( , ) , contributes to neuroinflammation and constitutes an important risk factor for the development or worsening of psychiatric disorders [for review, see ( ) ]. another important factor is aging, which is related to an imbalance in the levels of proinflammatory (high levels) and anti-inflammatory (low levels) cytokines and decrease in t-cell-mediated function ( ) . these immunosenescence-dependent changes in the elderly may be associated with higher susceptibility to viral diseases, including covid- ( ) , as well as neuropsychiatric disturbances, such as cognitive impairments ( ) . it has been demonstrated the relationship between aging and symptoms of anxiety and depression in patients infected with sars-cov- during the acute phase ( ). therefore, both obesity and older age may increase the risk of psychiatric symptoms in patients with covid- ; and one hypothesis is that neuroimmune circuits may be involved in this association. in addition, since poor nutrition and sedentary lifestyle are frequent in the elderly population and in overfat individuals, actions that promote the practice of physical activity and adequate nutrition are crucial, as they can potentially be associated with a lower risk for covid- and mental health impairments. pregnancy is another important potential factor that can affect the neuropsychiatric outcomes of covid- . maternal immune activation (e.g., in response to infection) is a risk factor for neurodevelopmental disorders such as autism spectrum disorder (asd) ( ) . autism has a complex etiology, involving environmental, and genetic factors. one of the proposed etiologies for asd is viral infection in early stages of development ( ). although the mechanisms by which viral infection can lead to autism are not yet known, it is believed that they may occur through (a) direct infection of the infant cns, or (b) due to the inflammatory response of the mother and/or the fetus, which can lead to neuroinflammation, triggering changes in brain development ( ). in fact, clinical evidence supports the participation of the neuro-immune mechanisms in the pathophysiology of asd [for review, see ( ) ]. while increasing evidence supports the neuroinvasive potential of sars-cov- , there is still no consistent demonstration of vertical transmission of this virus. in this sense, a recent study reviewing the effects of sars, mers, and covid- on gestational outcomes, including vertical transmission, and demonstrated that fortunately this transmission mechanism does not appear to occur in these betacoronaviruses ( ) . however, the controversial data on this aspect and the high expression of ace detected in the human placenta ( ) revealed that the possibility of vertical transmission needs to be further explored in clinical settings. accordingly, it is important to point out that there is still insufficient evidence to support the association between sars-cov- infection during pregnancy and the development of asd. nonetheless, we cannot rule out that changes in the maternal immune response triggered by the sars-cov- infection may affect neurodevelopment, another aspect that also deserves the attention of the medical and scientific communities. in any case, since increased levels of cytokines have been observed in covid- and in psychiatric disorders, it is likely that changes in neuroimmune axes may be involved in the mental health outcomes occurring in covid- patients. although this hypothesis is based mainly on studies with other betacoronaviruses, it will be interesting if future clinical studies, for example, include the search for correlations between the levels of inflammatory markers and psychiatric symptoms in covid- patients and survivors. studies in animal models infected with sars-cov- may also assist in the investigation of possible pathological mechanisms involved in neurobehavioral disorders related to the viral infection. the activation of the hypothalamic-pituitary-adrenocortical (hpa) axis has been observed during pathologies involving an immune/inflammatory process, including viral infections ( ) . the activation of this neuroendocrine axis by pro-inflammatory cytokines causes increased glucocorticoid production, a physiological response that contributes to avoid the deleterious effects of excessive production of inflammatory mediators and a non-specific recruitment of cells with no or low affinity for triggering antigens ( ) . in this respect, it seems reasonable to imagine a state hyperactivity of the hpa axis in infected patients, due to the "cytokine storm" observed in these individuals ( figure a) . a second aspect deserving discussion is the fact that ace overexpression in corticotropin-releasing-hormone (crh)producing neurons in the hypothalamic paraventricular nucleus alters the processing of psychogenic stress in mice, decreasing the crh content in the hypothalamus and corticosterone plasma levels (i.e., less hpa axis activation), as well as anxiety-like behaviors ( ) . sars-cov infection decreases the expression of ace in the lungs and myocardium of infected mice ( , ) . also, patients who died from sars and had sars-cov detected in the hearts exhibited reduced ace levels, when compared to patients who died from a non-sars related sepsis ( ) . although sars-cov genomic sequences have been found in the hypothalamus of humans ( ) , it remains to be determined whether the virus also decreases ace contents in this brain region. in any case, a downregulation of hypothalamic ace levels may be considered as another potential mechanism by which sars-cov/sars-cov- induces hyperactivity of the hpa axis with consequent psychiatric disturbances that are observed in these patients, such as the anxiety for example ( figure b) . however, the role of ace in the sars-cov- pathogenesis is still unknown and more studies are needed to test this mechanism. by contrast, in a study that prospectively assessed the presence of hormonal changes in sars survivors (without pre-existing endocrine disorders) months following recovery, patients ( . %) displayed late hpa axis hypoactivity, with hypocortisolism ( ). this alteration appeared to be a pathological effect of sars-cov, since nearly two-third of the patients did not use steroids and the majority were young (mean age: . years) and previously healthy ( ) . retrospective data from sars survivors do not support changes in hpa axis activity during the acute phase, suggesting that sars-associated hypocortisolism is a late onset phenomenon ( ) . since the "cytokine storm" is seen in the acute phase of sars (see table ), increased cytokine levels are unlikely to be secondary to hpa axis hypofunction. although proinflammatory cytokines classically increase the activity of the hpa axis (i.e., a downregulation mechanism of the inflammatory figure | hypothetical mechanisms by which sars-cov- may lead to changes in the activity of the hypothalamus-pituitary-adrenal (hpa). (a) during a viral infection (e.g., sars-cov- ), pro-inflammatory cytokines are released by immune cells present in the periphery (e.g., macrophages, t and nk cells) and/or in the brain (microglia). these cytokines can act at three levels of the hpa axis: increasing (i) the secretion of the corticotrophin-releasing hormone (crh) in the hypothalamus, (ii) the secretion of adrenocorticotropic hormone (acth) in the pituitary, and (iii) release of glucocorticoids (e.g., cortisol) through the adrenal cortex. by any of these actions, the result is an increased release of glucocorticoids, which bind to their receptors present in immune cells, suppressing the synthesis and release of pro-inflammatory cytokines. therefore, it is possible that increased pro-inflammatory cytokine levels in covid- may lead to hyperactivity of the hpa axis. however, due to a dysfunction in the negative feedback between the hpa axis and the immune system, this neuroendocrine axis is not able to reduce the production of inflammatory mediators, a possible explanation for why sars-cov- infection leads to cytokine storm. (b) hypothalamic ace overexpression decreases the activity of the hpa axis in mice, reducing the crh content in the hypothalamus and corticosterone plasma levels. since sars-cov infection is able to reduce the expression of ace in other tissues, one hypothesis (based on molecular similarities between sars-cov- and sars-cov) is that sars-cov- can induce a decrease in hypothalamic ace levels, thus contributing to hpa hyperactivity. (c) although pro-inflammatory cytokines classically increase the activity of the hpa axis, some cytokines (e.g., tgf-β) can decrease the activity of this neuroendocrine axis under specific conditions that remain unclear. this is another mechanism by which the sars-cov- infection, inducing an exacerbated inflammatory response, may lead to changes in the hpa axis, in this case, hypoactivity. continuous arrows: stimulation; dashed arrows: inhibition. response), under some conditions, tnf-α and transforming growth factor beta (tgf-β) may induce hpa axis hypoactivity ( ). therefore, it is possible that some cytokines that are increased in sars patients play a causative role in sars-associated hypocortisolism. as both hyperactivity and hypoactivity of the hpa axis are associated with depression ( , ) , hypocortisolism can also be associated with depressive symptoms that can be in sars survivors. in addition, due to the similarities between sars-cov- and sars-cov, it is possible that this mechanism involved in hpa axis hypoactivity can also be observed in covid- ( figure c) . thus, studies that simultaneously evaluate the axis hpa activity, cytokine levels, and psychiatric disturbances in patients and survivors of covid- will certainly improve the current knowledge. in the above context, it is noticeable that long-term survivors of the acute respiratory distress syndrome often report traumatic memories from the icu. interestingly, these patients displayed lower baseline cortisol levels and higher incidence of ptsd ( ) . such an information leads to questions related to the hypocortisolism observed in sars-cov infected patients, which may reflect an exhaustion of the adrenal cortex function, as a result of the viral infection or distress associated with hospitalization. clearly, future studies are needed to assess whether sars-cov- can affect the functioning of the hpa axis and whether this is involved in the association between sars-cov- infection and mental health outcomes. in clinical settings, it will be important to observe and measure the stress associated with hospitalization, as well as the presence of traumatic memories, as these factors may also be associated with changes in the hpa axis. a dysfunctional glucocorticoid-immune circuitry has been observed in schizophrenia. after a stress paradigm, while healthy patients experienced an increase in cortisol levels, negatively correlated to the subsequent changes in il- levels, patients with schizophrenia had elevated cortisol positively correlated to subsequent changes in il- levels, suggesting an inability to down-regulate inflammatory responses to psychological stress in this psychiatric condition ( ) . it is well-known that stressful life events may precipitate subsequent exacerbations of the illness ( ) . interestingly, elevated levels of circulating il- have been found in early episode psychosis patients ( ) . increased levels of stress or il- have also been described in sars or covid- patients (see supplementary table and table ). in addition, several studies reported symptoms of psychosis during the acute or long-term phase in sars patients (see supplementary table ). therefore, it is possible that sars-cov- infection and stressors related to hospitalization may increase the risk of psychosis by increasing levels of cytokines and/or by disrupting the glucocorticoid-immune circuits. since infections are associated with increased risk of developing schizophrenia ( ) , it seems important that future studies further assess the potential association between sars or covid- and the development of schizophrenia, as well as highlighting the importance of measures that prevent or reduce the impact of covid- on mental health. therefore, it is possible that increased pro-inflammatory cytokine levels in covid- lead to hypoactivity or hyperactivity of the hpa axis and, due to a dysfunction in the negative feedback between the hpa axis and the immune system, this neuroendocrine axis is not able to reduce the production of inflammatory mediators. in this sense, we hypothesize that such a dysfunction in the negative feedback between the hpa axis and production of pro-inflammatory cytokines may also be associated with mental health outcomes of the sars-cov- infection, thus conceptually corresponding to a psychoneuroendocrine-immune dysfunction. pre-clinical studies will hopefully provide more consistent clues to define a putative causal association between sars-cov/sars-cov- infection and behavioral deficits. in addition, animal models should allow a better control of variables that could also affect this association, such as the isolation of infected patients, since social isolation per se can also lead to both immunological and behavioral dysfunctions. in the current scenario, where social isolation measures are being strongly implemented worldwide, it is also important put into focus the potential damage to the mental health of isolated individuals, infected or not, applied the psycho-neuroendocrine-immune approach discussed herein. the exponential increase in the number of people infected with sars-cov- is leading to saturation of health services worldwide. to prevent human-to-human transmission and, in this way, slow down the growth of the pandemic, who has recommended that people avoid getting outside as much as possible ( ) . although such a measure is necessary to contain the advance of the pandemic, social isolation can cause negative impacts on mental health of individuals. studies on mental health outcomes of the quarantine during other epidemics, including sars and mers, revealed negative psychological effects, such as symptoms of ptsd, depression, stress, anxiety, and fear. some of the predictors of psychological impact included having a history of psychiatric illness, healthcare work, longer quarantine duration, infection fears, boredom, inadequate supplies, inadequate information, and financial resources ( ) . results of an online survey that assessed the levels of psychological impact and stress during the initial stage of covid- outbreak were recently reported ( ) . the responses of , subjects showed that . , . , and . % had moderate to severe stress levels, anxiety and depression symptoms, respectively. moreover, the general public with no formal education had a significant greater likelihood of depression during epidemic and higher satisfaction with the health information received was associated with a lower mental health impact of outbreak. people that presented sars-cov- -related symptoms like coryza, cough, dizziness, and myalgia or reported a history of chronic illnesses showed significant high levels of anxiety, depression, and stress. these results suggest an importance of accurate health information to reduce the impact of rumors and show the need for the media to provide, not only true information, but also information in simple language so that to support those people with less educational background during the epidemic ( ) . in addition, these data lead to the urgent need of psychological and psychiatric interventions, together with measures to prevent the spread of sars-cov- , so that to provide, as much as possible, well-being to both infected and non-infected socially isolated people. several studies show that living alone (vs. living with a family member) is associated with elevated levels of depressive symptoms ( ) ( ) ( ) , higher risk of depression ( ), and higher mortality ( ). yet, it has been emphasized the need for caution in arguing for a negative association between living alone and mental health ( ) . one reason is that other factors may influence the association between living arrangements and mental health, such as social networks ( , ) , social support ( , ) and neighborhood environment ( , , ) . in a study using data from more than , individuals in the united kingdom or england, it was shown that prevalence of common mental disorders was higher in people living alone vs. people not living alone. this association occurred regardless of age and gender but was largely mediated by loneliness. therefore, we believe that people living alone may be more vulnerable to the effects of quarantine on mental health than people living with a family member. accordingly, it would be interesting for future studies to assess the influence of different living arrangements on outcomes of quarantine on mental health. in this framework, loneliness has been associated with several psychiatric disorders, such as depression, anxiety, and suicide behavior ( ) . importantly, it has been showed that lonely people present several immune dysregulations, such as upregulated expression of pro-inflammatory cytokine genes ( ) . on the other hand, several studies have revealed that changes in the immune system play a key role in mental disorders ( ) . therefore, it is possible that changes in the immune system are involved in the negative impacts of loneliness on mental health. accordingly, it is conceivable that inflammatory mediators are also involved in the impact of quarantine on mental health, during covid- . studies with animal models have provided important clues on the neurobiological and the behavioral consequences of social isolation. in rodents, the stress of social isolation is able to lead to changes in several neurotransmitter systems (e.g., dopaminergic, adrenergic, serotonergic, gabaergic, glutamatergic, nitrergic, and opioid systems). indeed, the synthesis, release and even the corresponding receptor expression can be altered in several brain regions (e.g., hippocampus, cortex) of animals submitted to social isolation stress [for review, see ( ) ]. disturbances in neuroplasticity-related signaling pathways are also observed in these models ( ) . for instance, rats submitted to chronic social isolation stress displayed brain morphological changes such as decreased number of dendritic spines in the hippocampus and prefrontal cortex, as well as decreased brain-derived neurotrophic factor (bdnf) and phosphorylatedprotein kinase b (p-akt) in the dorsal hippocampus ( ) . the bdnf/trkb/pi k/akt pathway had already been described to be an important pathway in the maintenance of synaptic plasticity through translation and transport of synaptic proteins ( , ) . in this context, a metanalysis study reported a positive correlation between lower bdnf serum levels and depressive symptoms ( ) , and patients who present depressive symptoms may have reduced hippocampal volume ( ) , which supports the association between neuroplasticity and depressive disorders. the social isolation stress can also lead to hyperactivity of the hpa axis through an increase in corticosterone production and release in rodents ( ) . the abnormal levels of glucocorticoid have been related to depressive-like behavior and can affect the hippocampal neurogenesis ( ) . additionally, social isolation stress can lead to neuroinflammation, with higher levels of tolllike receptors, il- and tnf-α in the hippocampus ( ) , as well as increased plasma levels of tnf-α, il- , il- , and acth in isolated rats ( ) . a recent systematic review reported that social isolation and loneliness may be linked to systemic inflammation (i.e., high levels of c-reactive protein and il- ) in the general population ( ) . accordingly, it is conceivable that nervous, immune and endocrine systems can be interacting with each other, mediating neurobehavior impairments induced by social isolation stress. thus, these interactions may be part of the mechanisms by which social isolation during quarantine, via changes in neuroendocrine-immune circuits, can trigger damage to mental health. yet, future studies are needed to understand the mechanisms associated with the psychological damage caused by quarantine. although the whole population can be affected by the psychological impacts of covid- , some vulnerable groups may experience the same pandemic scenario differently. a recent study based on a multidisciplinary approach called attention for measures that can support the population susceptibilities such as ( ) older adults with multicomorbidities, ( ) children and women that stay at home and suffer domestic violence, ( ) people with preexisting mental health issues, ( ) people with learning difficulties, which might be affected by disruption to support and by loneliness, ( ) front-line health care workers that can be affected by the fear of infection, and ( ) groups that have hard socio-economic difficulties ( ) . as previously mentioned, financial problems may enhance the impact of social isolation on mental health during quarantine ( ) . interestingly, studies demonstrated that a worse socioeconomic status is directly related to higher systemic levels of inflammatory markers such as il- and c-reactive protein [for review, see ( ) ]. thus, it is possible that neuroimmune interactions may also be involved in the impacts of financial stress during covid- on mental health. this represents a novel possibility, that for sure requires future investigation. in addition, higher levels of inflammatory markers associated with worse socioeconomic conditions may also explain why lower social support is also associated with symptoms of anxiety and depression in patients infected with sars-cov- ( ). even though the biological mechanisms involved in the impact of socioeconomic status on mental health are still unclear, actions aiming at reducing socioeconomic inequalities should be a priority, in order to mitigate the impacts of covid- on mental health. finally, it is important to note that the evidence highlighted here does not contradict the need for the isolation measures that are necessary to control the pandemic. however, they call attention to the usefulness of strategies aiming at reducing the harmful effects of social isolation on mental health of the general public, including the improvement of psychological intervention and the reduction of socioeconomic inequalities. in summary, previous studies have reported psychiatric manifestations in patients infected with sars-cov- , such as anxiety, depression and ptsd symptoms ( , ). since increased levels of cytokines have been observed in covid- and in psychiatric disorders, we can place immune/inflammatory pathways as one of the mechanisms involved in mental health outcomes of covid- . changes in the hpa axis have also been observed in sars patients, indicating that alterations in neuroendocrine-immune circuits may be related to the psychiatric symptoms observed in these individuals. therefore, the hypothesis of the present article is that sars-cov- infection can lead to neuroinflammatory and endocrine changes, which in turn may reflect poor mental health. however, it is important to note that related biological factors (e.g., older age, female gender, and overfat), together with other factors inherent to covid- (e.g., social isolation, financial stress, and adverse effects of treatments) can influence psychiatric outcomes. accordingly, it is likely that the psychiatric symptoms observed in covid- patients are due to processes involved in the virus-host relationship, as well as to psychosocial and therapeutic issues associated with the pandemic. a further important aspect to be pointed out is the impact that the covid- pandemic can have on people who are isolated to prevent the transmission of the virus and to prevent health system overload. similar to possible mechanisms involved in the impacts of sars-cov- infection on mental health, social isolation may also be associated with dysfunctional psycho-neuroendocrine-immune interactions, which in turn can contribute to the development or the worsening of psychiatric disturbances (figure ) . it urges to put all ours efforts in understanding the pathophysiology of covid- , including cns infection and the risk of mental health compromise, but also the effects of this pandemic in the healthy isolated individuals, including children and adolescents, so that to prevent a "new generation" of groups in which the risk of developing mental disturbances, as anxiety or depression, could be increased. if nothing is done, we will probably be doomed to face a new mental health "pandemic" in the future. in terms of social aspects, a number of short term simple attitudes or initiatives, can comprise the encouragement to: (a) strengthen bonds using social media and start thinking positively ( ); (b) sleep properly and exercise regularly ( ); (c) balance the diet, regular daily routine, relaxation exercise and other healthy lifestyle measures ( ) . on the other hand, people should be avoid: substance use, eating too much fast food, excessive online activity, excessive watching television, and believing fake news ( ) . it is also important to look for strategies that mitigate the impacts of covid- on frontline healthcare providers. for instance, as recommended by ho et al. ( ) , healthcare organizations should introduce shorter working periods, regular breaks, and rotating shifts. individuals who experience moderate to severe and/or persistence distress should seek help from mental health professionals or in hospitals in cases of emergency situations ( ) . in addition, online consultation can be a potential alternative of delivering therapy ( ) . based on the similarity of sars-cov- and sars-cov, hematogenic or neuronal retrograde dissemination routes (via olfactory nerve) may be involved in the entry of the sars-cov- into the central nervous system (cns). in the cns (left) the virus can lead to increase in cytokines levels (e.g., il- , il- , tnf-α, il- β, inf-γ, and il- ) due to its local or peripheral (right) actions. increased cytokine levels are associated to neuronal death, synaptic plasticity impairments, dysfunction in the neurotransmitter metabolism and in the hypothalamic-pituitary-adrenocortical (hpa) axis. likewise, social isolation can also lead to these neuroendocrine-immune disturbances, for instance: increase in cytokine levels, changes in neurotransmitter systems, hpa axis hyperactivity and disturbances in neuroplasticity-related signaling pathways. through these common mechanisms, both sars-cov- infection and social isolation can lead to mental health impairments [e.g., impaired memory, depression, psychoses, anxiety and posttraumatic stress disorder symptoms (ptsd)]. il, interleukin; tnf-α, tumor necrosis factor alpha; inf-γ, interferon gamma. we also believe that art (especially music) can be an ally in the quest to improving mental health, whether for inpatients, health care workers, or isolated people. a meta-analysis study reported that music can modulate cytokine levels (including reducing il- levels), as well as neuroendocrine-immune responses triggered by stress, including physical stress caused by viral infection ( ) . in addition, it has been reinforced that music interferes positively in the immune system when subjected to acute stress (co stress test), also regulating the function of il- and the hpa axis ( ) . therefore, music therapy can be a further relevant and simple strategy that might be adopted on a largescale basis, for individuals in social isolation (also including medical staff). overall, it is important that political and health authorities pay attention to the mental health of infected and uninfected individuals during the pandemic, looking for prevention and treatment strategies, since poorer mental health can be associated with shorter life expectancy ( ) ( ) ( ) and high economic burden ( , ) . beyond the immediate and fundamental task of saving lives during sars-cov- pandemic, the due care of his mental health should be timely addressed. protocols aiming at minimizing mental problems during the infection as well as during recovering after hospitalization must be designed. in addition, studies that evaluate the impact of isolation during sars-cov- pandemic on mental health are important as they can guide new strategies to preserve population mental health in other critical situations that we can live in the future. finally, it is noteworthy that the approach applied herein, related to psychoneuroimmunology in covid- , should be convergent with a social sciences approach so that to better understanding and to better tackling this disease. hopefully, future studies may test the hypothesis outlined herein to better understand and consequently mitigate the impacts of covid- on mental health. the original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author. 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health strategies to combat the psychological impact of covid- beyond paranoia and panic video consultations for covid- the psychoneuroimmunological effects of music: a systematic review and a new model the impact of acute stress on hormones and cytokines, and how their recovery is affected by music evoked positive mood understanding excess mortality in persons with mental illness: -year follow up of a nationally representative us survey mental disorders and cause-specific mortality excess mortality, causes of death and life expectancy in , patients with recent onset of mental disorders in denmark, finland and sweden a review of the economic impact of mental illness the economic burden of mental illness we are grateful to arnaldo p. andrade for his valuable suggestions in writing this article. the supplementary material for this article can be found online at: https://www.frontiersin.org/articles/ . /fimmu. . /full#supplementary-material conflict of interest: the authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.copyright © raony, de figueiredo, pandolfo, giestal-de-araujo, oliveira-silva bomfim and savino. this is an open-access article distributed under the terms of the creative commons attribution license (cc by). the use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. no use, distribution or reproduction is permitted which does not comply with these terms. key: cord- - ey gzw authors: lo, anthony wi; tang, nelson ls; to, ka‐fai title: how the sars coronavirus causes disease: host or organism? date: - - journal: j pathol doi: . /path. sha: doc_id: cord_uid: ey gzw the previous epidemic of severe acute respiratory syndrome (sars) has ended. however, many questions concerning how the aetiological agent, the novel sars coronavirus (cov), causes illness in humans remain unanswered. the pathology of fatal cases of sars is dominated by diffuse alveolar damage. specific histological changes are not detected in other organs. these contrast remarkably with the clinical picture, in which there are apparent manifestations in multiple organs. both pathogen and host factors are important in the pathogenesis of sars. the choice of specific receptors and the unique genome of the sars‐cov are important elements in understanding the biology of the pathogen. for the host cells, the outcome of sars‐cov infection, whether there are cytopathic effects or not, depends on the cell types that are infected. at the whole‐body level, immune‐mediated damage, due to activation of cytokines and/or chemokines and, perhaps, autoimmunity, may play key roles in the clinical and pathological features of sars. continued research is still required to determine the pathogenetic mechanisms involved and to combat this new emerging human infectious disease. copyright © pathological society of great britain and ireland. published by john wiley & sons, ltd. severe acute respiratory syndrome (sars) is a new viral disease caused by a novel coronavirus, sars-cov ( figure ) [ , ] . the saga of sars has officially come to an end, as no more new cases have been reported since . many questions, particularly those related to how sars-cov causes disease, however, remain unanswered. the disease caused by sars-cov differs from the diseases caused by the previously known human coronaviruses, e and oc . sars-cov infection results in severe and potentially fatal lung disease [ , ] . although the majority of patients recovered after - weeks of debilitating febrile illness, a substantial proportion (up to one-third) developed severe inflammation of the lung, requiring ventilator support and intensive care. many patients in this group deteriorated into acute respiratory distress syndrome (ards). the mortality of this group of patients is high [ ] . manifestations in other organ systems are characteristic. lymphopenia [ ] , gastrointestinal symptoms [ ] , impaired liver function [ , ] , and impaired renal function [ ] are common. the possibility of viral infection in multiple organs has been raised and viral replication in the lung, kidney, and gastrointestinal tract was reported [ , ] . in addition, prolonged shedding of virus was found in some convalescent patients [ ] . however, chronic infection by sars-cov has not, to date, been documented in humans. moreover, asymptomatic carriage of sars-cov is rare [ ] . there are significant age differences in the prognosis of sars. children have a good prognosis [ ] , while elderly patients with chronic illnesses fare badly. sars is predominantly a lower respiratory tract disease, yet the most consistent and powerful prognostic indicator reported so far is blood lactate dehydrogenase (ldh) concentration [ ] , which is most likely a surrogate indicator and may reflect the extent of ongoing tissue damage. both pathogen and host factors are important for the progression of an infection. here, we review the pathology of sars infection. specific features of the pathogen sars-cov itself are then addressed. finally, host factors, particularly an emerging understanding of immunological and inflammatory responses to sars-cov infection, are discussed. virus particles can also be seen budding through the cytoplasmic membrane (b). each virion particle is - nm in size by transmission electron microscopy and is characterized by the numerous club-shaped projections on the outside, a ring beneath the envelope, and an electron-lucent centre. scale bars = nm (a) and nm (b) diffuse alveolar damage is the most characteristic pathology in sars most data on the human pathology of sars come from autopsy studies of fatal cases [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] . these reports thus reflect the terminal stages and are likely to represent only the more severe end of the spectrum of sars. treatment and co-morbid conditions might also modify the pathological changes. diffuse alveolar damage at different stages of organization is the most consistent finding in the lungs of sars patients in the terminal stage (figures a- f ). multinucleated syncytial cells ( figures g and h ) are characteristic, although these cells are rare. apart from when secondary infection occurs, the lack of a prominent inflammatory response is also distinctive. sars-cov is explicitly detected in the alveolar lining cells ( figures i and j ) [ , [ ] [ ] [ ] [ ] [ ] [ ] . no specific pathology is identified in the gastrointestinal tract ( figure ) [ ], urinary system [ ] , or other organ systems [ ] , apart from that related to end-stage multi-organ failure or those changes secondary to treatment. it is important to note that in some organs such as the liver, while definitive and distinct morphological and functional changes are observed, sars-cov may not be unequivocally demonstrable [ ] . it is clear that our understanding of the pathology of sars is incomplete. an obvious large gap is the lack of information on the early pathological changes of sars. during the epidemic, very few biopsies were obtained from patients with clinically active sars. the study of animal models is important in a number of ways. it has allowed the establishment of sars-cov as the aetiological agent [ ] . it also provides controlled conditions for the study of early changes in the disease. initial studies of macaque models were promising. the histology of infected lung tissue is similar to that in humans [ ] [ ] [ ] . both acute and organized stages of diffuse alveolar damage were seen when the macaques were sacrificed on the sixth day after a heavy dose of the virus. sars-cov was detected in the alveolar epithelial cells and in the intra-alveolar syncytial cells. however, detailed morphological studies and viral distribution in other organs in these animal studies are lacking. in studies involving longer observation times, the disease in macaque models appears self-limiting and different from the genuine human disease. the usefulness of the macaque as a model of the disease remains to be established [ , ] . civet cats, domestic cats, and ferrets are thought to have been potential reservoirs of the virus during the epidemics and subsequent smaller outbreaks in mainland china [ ] . the animal coronavirus identified in civet cats shows high sequence identity with, but is distinct from, sars-cov [ , ] . recent evidence also suggests that wide chinese horseshoe bats harbour a closely related bat-sars-cov which might also act as the animal reservoir [ ] . again, details concerning the distribution of virus in different organs in these animals and the information on the pathology in the diseased or carrier animals are, surprisingly, sparse [ , ] . other common small laboratory animal models, such as the mouse, are not particularly useful. sars-cov has a low virulence in ordinary laboratory mice and very high levels of inoculation are required to produce self-limiting diseases. these features may be although the latter finding may be related to pre-morbid lung pathology, a correlation with interstitial fibrosis and disease duration has been demonstrated [ ] . diffuse alveolar damage at different stages of organization, from fibrin deposition (c, h&e, original magnification × ), to interstitial fibrosis (d, h&e, original magnification × ) and cellular organization (e and f, h&e, original magnification × ), can be detected. atypical pneumocytes with enlarged nuclei and prominent nucleoli are often seen and some pneumocytes coalesce into syncytial multi-nucleated cells (g, h&e, original magnification × ). multi-nucleated histiocytes may also be found (h, h&e, original magnification × ). sars-cov can be detected in pneumocytes by in situ hybridization (i, using a dna probe against the m gene, original magnification × [ ] ). a large array of antibodies against the viral proteins including nucleocapsid n, spike s, membrane m, and sars- a [ ] , has been developed for the detection of sars-cov in formalin-fixed, paraffin-embedded tissue sections (j, showing immunohistochemical staining with an anti-peptide antibody against n, original magnification × ) related to differences in the affinities of sars-cov for human receptors and their murine homologues [ ] . sars-cov uses a protector of lung damage, angiotensin-converting enzyme , as a receptor characterization of the functional cellular receptor of sars-cov provides important clues to the pathogenesis of sars. angiotensin-converting enzyme (ace ) interacts directly with the spike (s) proteins of the sars-cov [ ] [ ] [ ] [ ] [ ] . the level of expression of ace correlates with the efficiency of sars-cov infection in cell culture models [ ] [ ] [ ] . ace proteins are expressed by alveolar epithelial cells and by surface enterocytes of the small intestine [ ] , which are the primary target cells of sars-cov. studies in the intestine cell culture model, however, suggested that, in addition to ace , unknown co-factors or coreceptors are required to convey infectivity [ ] . in addition to being a cellular receptor, ace may contribute to the pathogenesis of dad in sars through its role in the tissue renin-angiotensin system. in a mouse model of alveolar damage induced by acid aspiration, the balance of the renin-angiotensin system appears to affect the development of dad. ace , which acts as a negative regulator of the local renin-angiotensin system, protects the mouse lung against experimental damage [ , ] . sars-cov co-infection in these damaged animals downregulates ace in the lungs of infected mice and the severity of lung damage can be alleviated by blocking the system [ ] . exciting as these findings appear, the case of a new coronavirus, nl , immediately provides an example that other factors are acting in the overall mechanism of lung damage. nl utilizes the same ace protein as its receptor in the lung. however, infection with nl results in only minor cold symptoms and alveolar damage is rare [ ] . the insert/deletion genotype of the ace gene was associated with dad after sars-cov infection in a small cohort of patients [ ] . this association was, however, not replicated subsequently in a larger series [ ] . we also could not detect any association between the ace genotype and disease severity in sars-cov infection [ ] . sars-cov may also use the c-type lectins as receptors for infecting immune cells c-type lectins, including cd and cd l, are also sars-cov receptors: these were identified through the study of proteins that interact with the s (spike) protein. cd , also known as dendritic cell-specific intercellular adhesion molecule-grabbing non-integrin (dc-sign), was shown to mediate viral entry in a lentiviral pseudo-type experimental model [ ] . in chinese hamster ovary (cho) cells expressing a human lung cdna library, s protein and its fragments interacted directly with a second related cell surface glycoprotein, cd l, also known as l-sign or dc-signr [ ] . cd l acts in conjunction with lsectin (liver and lymph node sinusoidal endothelial cell c-type lectin) and enhances viral infection [ ] . tissue cultures expressing cd or cd l were also susceptible to sars-cov infection [ , , ] . the possible involvement of dendritic cells is particularly interesting. although sars-cov does not replicate in dendritic cells, these cells may act as a reservoir and distribute the virus to other cell types [ , ] . this is an attractive concept and similar biological behaviours have been proposed for human immunodeficiency virus i (hiv i) [ ] . no sars-cov has been detected in dendritic cells in autopsy and biopsy studies reported so far. the genome of sars-cov consists of a single . kb positive-strand rna. the genomic sequences derived from different phases of the sars epidemic revealed no association with sequence variation and virulence [ , ] . there are two large open reading frames (orfs) and potential orfs in the sars-cov genome. the two large orfs encode non-structural proteins involved in replication. these proteins have relatively higher homologies to known coronaviruses. the remaining orfs are squeezed into the end of the genome. these orfs include four genes encoding known structural proteins (envelope, membrane, nucleocapsid, and spike proteins, respectively). the remaining potential orfs encode hypothetical sars-cov-specific proteins which lack obvious sequence similarity to known proteins [ , ] . the functions of these hypothetical proteins and their roles in sars pathogenesis remain obscure [ , ] . antibodies against some of these putative proteins, notably sars a and sars , can be detected in the serum of sars patients [ ] . there is also evidence suggesting that a number of these proteins, including sars a, b, a, and b, were expressed in pneumocytes and enterocytes in deceased patients [ ] . however, differential expression patterns of these proteins in cell types showing different responses to sars-cov infection have not been confirmed. by expressing the hypothetical proteins individually in tissue culture, we are beginning to see data on the cellular functions of these proteins. sars a appears to be important in mediating apoptosis in some cell types [ ] . the sars a protein is incorporated into the viron particle and may also act as one of the structural proteins [ ] [ ] [ ] . through an unknown mechanism, host cells overexpressing sars a have increased expression of fibrinogen mrna [ ] . sars a has been implied in mediating apoptosis through the caspase-dependent pathways [ ] . the effect of sars-cov infection varies in different cell types. apoptosis and syncytial formation are seen in infected monkey renal epithelial cells (vero e ) [ ] . persistent infection with no change in cellular morphology or doubling time was detected in the colon cancer cell line lovo [ ] . in clinical specimens, sars-cov was detected in the lungs and small intestine. severe cellular damage is characteristically detected in the lungs of sars patients, while no morphological changes are observed in the small intestine. the basis of these differences in cellular responses is not clear. the tissue/cellular tropism may be partly related to differential expression of membrane receptors for the sars-cov [ ] . these observations highlight the importance of host cell responses in sars-cov infection. it is also clear from these observations that cytopathic damage alone cannot explain the pathogenesis of sars. the marked heterogeneity of the disease course and outcome after sars infection suggests that host responses may play an important role in pathogenesis. dad or ards appears to be a common pathway of lung parenchyma damage initiated by a variety of aetiologies, including sars-cov infection itself, systemic sepsis, shock, and direct lung contusion. once an inflammatory process reaches a certain intensity, it may self-perpetuate. the cellular inflammatory infiltrate releases toxic metabolites and proteolytic enzymes, which may cause further damage to the lung parenchyma. the surrounding inflamed capillaries launch the coagulation cascade and recruit more immune cells [ , ] . our previous investigation in the h n influenza outbreak showed that patients who died of the disease had lymphoid depletion associated with marked elevation of circulating concentrations of cytokines, including interleukin- (il- ), il- receptor, and interferongamma [ ] . with the observation of characteristic lymphopenia in sars, it has been postulated that the sars-cov may similarly trigger an exaggerated hyper-cytokinemic response in patients with dad after viral infection [ ] . current understanding indicates that patients with a more intense immune response are those at risk of a poor outcome, as the immune system also mounts a profound reaction to the bystander, the lung parenchyma, and causes dad [ ] . sars patients have variable humoral responses to individual epitopes [ ] . however, early sero-conversion and high peak total sars-cov igg levels were associated with more severe disease in a cohort of patients [ ] . hence, particularly strong humoral responses to sars-cov infection might not be protective but, perhaps, might be harmful to the host. the specific epitopes upon which these 'damaging' antibodies act await further characterization. there is evidence that disarray of the immune system towards the host's own antigens may play a role in the pathology of sars. in the early phase, within week of sars-cov infection, igm and igg autoantibodies against antigens located in the cytoplasm of lung epithelial cells (figure ) were detected in the sera of chinese sars patients (lo, unpublished observations). in another cohort of sars patients, immune activity against antigens from lung epithelial cell lines and endothelial cell lines was found in some patients' sera obtained approximately month after infection [ ] . moreover, high levels of these autoimmune activities in the sera were shown to be cytotoxic to lung epithelial cells and endothelial cells in culture. autoimmune antibodies may be important in mediating tissue damage at certain stages of the disease. the cause of the autoimmunity is not fully understood. these autoantibodies may be the result of humoral responses to innate antigens exposed accidentally during direct damage of the lung and, perhaps, the endothelium by sars-cov. alternatively, autoimmunity may be due to crossreactivity of antibodies against some specific epitopes of the sars-cov proteins. the chemokines are a family of small proteins that play important roles in intercellular signalling and chemotaxis. based on their protein sequences, they are broadly divided into α-chemokines with a common c-x-c (cysteine-other-cysteine) structure of amino acid residues near the amino-terminus which interacts predominantly with neutrophils, and β-chemokines with a c-c (cysteine-cysteine) structure interacting with mononuclear cells. recently, chemokines have been recognized for their roles in integrating the innate and adaptive immune responses to viral infection through a cytokine-to-chemokine-to-cytokine signalling cascade [ ] [ ] [ ] . a global view of the spectrum of expression of the immune mediators was studied in sars by measuring the circulating concentrations of these mediators at different stages of the disease. most cytokines showed only transient and short-lived activation in patients after sars-cov infection [ ] . even in patients who developed dad, most cytokine concentrations were not significantly increased [ ] . in contrast, circulating concentrations of several chemokines, including cxcl (chemokine c-x-c motif ligand or monokine induced by γ -interferon), cxcl (chemokine c-x-c motif ligand or interferoninducible protein- ), and ccl (c-c motif ligand or monocyte chemoattractant protein- ), were markedly increased in sars patients [ , , ] . remarkably, the circulating concentration of cxcl measured early after infection is an independent prognostic indicator of disease outcome [ ] . these chemokines therefore appear to be important elements of the pathogenesis of sars. in the lung tissues obtained from seven sars patients who died [ ] , chemokines cxcl ( figure ) and il- were markedly activated ( and -fold compared with controls, respectively). the important roles of chemokines are underscored by the findings in an experimental mouse model of sars-cov infection in which cxcl and a neutrophil chemokine, cxcl (chemokine c-x-c motif ligand ), were also markedly activated [ ] . these findings in sars compare favourably with the specific situation in hiv patients with lung allograft rejection and interstitial alveolitis, in which similar activation of the chemokine cxcl and its receptor cxcr (chemokine c-x-c motif receptor ) was also found [ , ] . other than pneumocytes, chemokines are also expressed and secreted by various different cell types. global gene expression profiles, generated by cdna microarray analysis of peripheral blood mononuclear cells (pbmcs) after in vitro exposure to sars-cov, also reveal the importance of chemokine activation. within day after exposure to the virus, a number of chemokines (including cxcl , cxcl , and ccl ) were activated [ ] . pbmcs and macrophages do not support productive infection as viral replication is abortive and no infectious virus is produced. the roles of these cell types in the pathogenesis of sars remain to be clarified. nonetheless, these easily obtainable cell types provide convenient experimental models and allow some insight into the patterns of host responses to the infection to be studied. similar findings were also reported in other cell types, such as dendritic cells, where the cytokine expression profiles are predominantly of inflammatory chemokines ccl (chemokine c-c motif ligand ), ccl (chemokine c-c motif ligand ), cxcl , and ccl . unlike the usual response of dendritic cells to viral infection, anti-viral cytokines, including ifn-α (interferonalpha), ifn-β, ifn-γ , and il- b, were not activated [ ] . immunogenetics of the host may affect the severity of sars other than using serum inflammatory mediators to reflect the different degree of host inflammatory reaction during an infection, the intensity of the immune response is also genetically determined. the difference in genetic makeup between individuals is mostly accounted for by single base differences (single nucleotide polymorphisms, snps). many studies have shown an association between snps and predisposition to ards, and survival after sepsis or other insults [ , ] . in the context of predisposition to ards after trauma, among parameters such as circulating concentrations of il- , tumour necrosis factor and plasminogen activator inhibitor- (pai- ), and the genotype of pai- , insertion alleles at the promoter of pai- were associated with high concentrations of pai- in the plasma and a poor survival rate [ ] . in addition to pai- , other genetic polymorphisms, such as angiotensin-converting enzyme (ace) [ ] , cd [ ] , surfactant protein [ ] , and hla genotypes [ ] , are also associated with predisposition to, severity, and outcome of ards. although sasr-cov utilizes ace as its receptor and ace is known to be an important protector of lung damage in experimental ards, we and other groups found no solid association between alleles of the two ace genes (ace and ace ) and the severity of ards after sars infection [ , , ] . several immunogenetic studies have been reported in association with sars infection. among taiwanese sars patients, hla-b * was associated with both predisposition to infection and severity of infection [ ] . however, the association of this allele was replicated in another chinese community of hong kong involving sars patients [ ] . hla-b * was found to be a predisposition allele in the latter study. it should be noted that this latter allele is rare and is found in ∼ % of the general population. hence, this allele cannot be considered a major predisposition factor for sars infection [ ] . immunogenotype may play a role in determining the severity of host responses. there is considerable variability in the prevalence of immunogenotypes among different populations and the significance of detecting so-called 'predisposing' alleles in clinical practice is questionable. more studies are needed to uncover fully the real genetic determinants for both predisposition to infection and the host-pathogen interaction after infection with the virus. a considerable amount of knowledge of sars infection has accumulated as a result of almost years of research since the emergence of sars. some key issues about the pathogen, sars-cov, have been addressed. these include the rapid discovery of sars-cov receptors and the actions of some of the specific viral proteins in different host cells. understanding the molecular basis of differences in host cell responses to sars-cov infection will be crucial in delineating its pathogenesis. it is also clear from clinical and experimental data that host immune responses may be the key determinant in disease progression after initial sars-cov infection. future studies aimed at characterization of the variability of host immune and inflammatory responses will be important in understanding this new emerging infectious disease. a major outbreak of severe acute respiratory syndrome in hong kong sars -beginning to understand a new virus treatment of severe acute respiratory syndrome haematological manifestations in patients with severe acute respiratory syndrome: retrospective analysis enteric involvement of severe acute respiratory syndromeassociated coronavirus infection retrospective analysis of liver function 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cxcl /cxcr chemokine biology during pathogenesis of acute lung allograft rejection chemokine response in children with sars genetic polymorphisms associated with susceptibility and outcome in ards genetic susceptibility to acute lung injury plasminogen-activator-inhibitor- g/ g promoter polymorphism and prognosis of severely injured patients angiotensin converting enzyme insertion/deletion polymorphism is associated with susceptibility and outcome in acute respiratory distress syndrome association between a genomic polymorphism within the cd locus and septic shock susceptibility and mortality rate polymorphisms of human sp-a, sp-b, and sp-d genes: association of sp-b thr ile with ards common features in the onset of ards after administration of granulocyte colony-stimulating factor identification of an alternative -untranslated exon and new polymorphisms of angiotensin-converting enzyme gene: lack of association with sars in the vietnamese population association of hla class i with severe acute respiratory syndrome coronavirus infection association of human-leukocyte-antigen class i (b * ) and class ii (drb * ) genotypes with susceptibility and resistance to the development of severe acute respiratory syndrome key: cord- -pjbviagq authors: lisi, lucia; lacal, pedro miguel; barbaccia, maria luisa; graziani, grazia title: approaching coronavirus disease : mechanisms of action of repurposed drugs with potential activity against sars-cov- date: - - journal: biochem pharmacol doi: . /j.bcp. . sha: doc_id: cord_uid: pjbviagq on march , , the world health organization (who) declared the severe acute respiratory syndrome caused by coronavirus (sars-cov- ) a global pandemic. as of july , sars-cov- has infected more than million people and provoked more than , deaths, worldwide. from the beginning, a variety of pharmacological treatments has been empirically used to cope with the life-threatening complications associated with corona virus disease (covid- ). thus far, only a couple of them and not consistently across reports have been shown to further decrease mortality, respect to what can be achieved with supportive care. in most cases, and due to the urgency imposed by the number and severity of the patients’ clinical conditions, the choice of treatment has been limited to repurposed drugs, approved for other indications, or investigational agents used for other viral infections often rendered available on a compassionate-use basis. the rationale for drug selection was mainly, though not exclusively, based either i) on the activity against other coronaviruses or rna viruses in order to potentially hamper viral entry and replication in the epithelial cells of the airways, and/or ii) on the ability to modulate the excessive inflammatory reaction deriving from dysregulated host immune responses against the sars-cov- . in several months, an exceptionally large number of clinical trials have been designed to evaluate the safety and efficacy of anti-covid- therapies in different clinical settings (treatment or pre- and post-exposure prophylaxis) and levels of disease severity, but only few of them have been completed so far. this review focuses on the molecular mechanisms of action that have provided the scientific rationale for the empirical use and evaluation in clinical trials of structurally different and often functionally unrelated drugs during the sars-cov- pandemic. on january , , the world health organization (who) declared the outbreak of severe acute respiratory syndrome coronavirus (sars-cov- ; initially named novel coronavirus or -ncov) a public health emergency of international concern, highlighting the need for a coordinated international intervention to limit virus spreading. few weeks later, on march , , because of the rapid diffusion of the infection, the who announced that sars-cov- infection was a global pandemic. the first cases of respiratory disease caused by -cov- , thereafter officially named covid- (corona virus disease ), likely occurred from a zoonotic transmission in china in december and since then infection has spread across countries and territories. as of july, , sars-cov- has infected more than , , people and caused more than , deaths (https://www.who.int/emergencies/diseases/novel-coronavirus- /situation-reports/ accessed july , ). coronaviridae define a family of hundreds of enveloped, positive-sense, single-stranded rna viruses that are known to cause diseases in animals. sometimes these viruses become able to overcome the species barriers (spillover event) and, so far, coronaviruses are known to cause human diseases. among these, four human coronaviruses (i.e., hcov- e, hcov-nl , hcov-oc and hku ) typically affect the upper respiratory tract and cause relatively minor symptoms. however, the other three coronaviruses [severe acute respiratory syndrome coronavirus (sars-cov), middle east respiratory syndrome coronavirus (mers-cov) and sars-cov- ] are able to replicate in the lower respiratory tract and are responsible for severe forms of pneumonia that can be fatal ( ). phylogenetic analysis indicates that sars-cov- has high similarity ( - %) with two coronaviruses circulating in rhinolophus (horseshoe bats) ( ) , but it is less closely related to the sars-cov (~ % similarity) and mers-cov (~ % similarity). based on the sequence analysis of the . kb viral genome and on the presence of bats and live animals in the seafood wholesale market in wuhan (hubei province, china), where sars-cov- was detected for the first time, this virus might have arisen from bats or materials contaminated by bat droppings in the chinese seafood market areas and transmitted to humans either directly or through an intermediate host ( ) . similar to the other respiratory coronaviruses, sars-cov- is transmitted primarily via the respiratory route in the form of droplets, with a possible, though yet unproven, fecal-oral transmission route ( , ) . the virus is stable for several hours to days in aerosols and on various types of surfaces, suggesting that transmission may occur by person-to-person droplets as well as by contact with fomites in the proximity of infected patients ( ) . although many individuals remain asymptomatic, . % of diseased patients display clinical symptoms within . days ( ) . patients with covid- may exhibit mild to moderate symptoms, most commonly fever, fatigue, dry cough, anosmia/dysgeusia, or severe pneumonia with dyspnea, tachypnea, and hypoxemia. actually, dyspnea is predictive of severe covid- and intensive care unit (icu) admission ( ) . other symptoms less frequently reported include muscle and joint pain, headache, diarrhea, nausea or vomiting, hemoptysis ( ) . severe covid- is associated to acute lung injury (ali) and/or acute respiratory distress syndrome (ards) that generally occur - days after symptom onset. as with sars-cov infection, an aggressive inflammatory reaction is responsible for the damage to the lung, indicating that the disease severity also depends on dysregulation of the host immune responses. respiratory failure is the most common cause of death (> %) of fatal covid- cases. furthermore, the massive release of cytokines by the immune system can result in cytokine storm and septic shock and/or multiple organs dysfunction syndromes in % of fatal cases ( , ) . other causes of death are cardiac failure, coagulopathy and renal failure ( ) . sars-cov- appears also to target the central nervous system with anosmia and dysgeusia as early symptoms and convulsions that may develop later on ( ) . currently, the standard of care in patients showing ards includes oxygen therapy together with the administration of parenteral fluids. furthermore, many patients with severe respiratory distress, hypoxemia and ards require invasive mechanical ventilation, and, if the situation deteriorates, extracorporeal membrane oxygenation support ( ) . therapeutic interventions including administration of drugs may vary from country to country and it is extremely difficult to harmonize the different protocols due also to the different disease stages of the patients (asymptomatic, pre-symptomatic, mild, severe, under mechanical ventilation). so far, there is not a standardized effective pharmacological treatment for covid- , a part from anecdotal evidence of efficacy. the scarce knowledge of the sars-cov- biology and of the host-pathogen interactions leading to covid- has markedly hampered the prompt identification of suitable targets for the development of new therapies. a large number of exploratory clinical trials and pivotal studies are being carried out worldwide. among them, the international "solidarity trial" launched by the who on march with the aim to find an effective treatment for covid- patients by comparing four different treatments (i.e., lopinavir/ritonavir, lopinavir/ritonavir plus interferon-β, chloroquine/hydroxychloroquine or remdesivir) against standard of care (see also sections and ). presently, regulatory authorities all over the world underline the need of common and rigorous approaches to clinical trials in order to generate more robust evidence on the safety/efficacy of the different anti-sars-cov- treatments or vaccines that are being tested. here, we review the recently published literature on the pharmacological treatments used so far and/or undergoing evaluation in clinical trials, with focus on the biochemical mechanisms of action of repurposed or investigational drugs, classified as agents directly targeting the virus ( figure and table ) and those used to treat the respiratory distress and inflammation associated with the cytokine release syndrome ( figure and table ). in addition, we summarize the main clinical trials completed or still ongoing in sars-cov- infected patients. the first step in any viral infection entails binding of the virus to a host cell through its target receptor. both sars-cov and sars-cov- entry into cells requires the interaction of the viral spike (s) glycoprotein (the envelope-associated protein conferring coronaviruses the characteristic crown-like morphology) with the angiotensin-converting enzyme (ace ) ( ) ( ) ( ) . ace is a dimeric ectoenzyme with dipeptidyl carboxypeptidase activity. although the ace mrna has been detected in a variety of tissues ( ) , the protein has not always been analyzed or detected. the ace protein is expressed at high levels on the surface of the lung alveolar epithelial cells and enterocytes of the small intestine providing an easily accessible route for sars-cov- infection ( ) . the ace protein is also present in smooth muscle, pericytes and endothelial cells of the vasculature, heart, kidney and this might account for the multi-organ dysfunction observed in severe covid- patients ( ) ( ) ( ) ( ) ( ) ( ) . other tissue sites where the ace protein was detected include, among others, the basal epithelium of the nasal, nasopharynx oral mucosa, the basal cell layer of epidermis, and testis ( , ) . the viral s glycoprotein is a trimer and each monomer contains two subunits, s and s , of which s is responsible for the virus attachment to the host cell surface though the receptorbinding domain (rbd), whereas s is required for the fusion of the viral and cellular membranes. after the attachment step, the entry process requires the s protein priming by cellular proteases, consisting in the proteolytic cleavage at the s -s boundary and at a downstream position in s ; this process leads to the exposure of a peptide that is involved in membrane fusion ( , ) . the s proteins of coronaviruses can be cleaved by various cellular proteases; in the case of sars-cov- , the transmembrane protease serine protease (tmprss ) plays a critical role in s protein priming, whilst the endosomal cysteine protease cathepsin l may replace tmprss in this function in cells other than those of the lung ( , ). moreover, it has been recently demonstrated that the host cell protease furin can cleave the sars-cov- s protein at the s /s site cleavage, an essential step for viral entry into lung cells ( ). since ace is located within lipid rafts, cell infection by sars-cov- also requires interaction of the viral s protein with different raft components, including sialicacid-containing gangliosides; this interaction also facilitates the contact of the s protein with the ace receptor ( ). after cleavage of the s protein, sars-cov- can be induced to fuse at both the plasma membrane and the endosomal membrane ( , ). various endocytic pathways have been described as being used for cell infection by different coronaviruses, including clathrin-coated vesicles, caveolae as well as clathrin-and caveolae-independent mechanisms ( , ). different antiviral agents or other drugs used for indications unrelated to virus infections have been used to block sars-cov- entry into the host cells either by i) inhibiting virus attachment and proteolytic cleavage of the s protein, ii) targeting key cellular enzymatic activities or proteins involved in the endocytic processes or iii) using a combination of both mechanisms ( figure ). umifenovir (arbidol) is a small indole-derivative molecule with a broad spectrum of activity against dna/rna and enveloped/non-enveloped viruses that prevents viral entry into the host cell (attachment and internalization), behaving as host-targeting and direct-acting antiviral agent ( , ). in particular, due to its hydrophobicity, umifenovir displays high affinity for the lipids of the host cell membranes altering their fluidity and rendering them less prone to fusion with the virus. this agent is also able to interact with aromatic residues of the viral glycoproteins involved in the attachment and in the membrane destabilization necessary for the fusion process. furthermore, umifenovir markedly affects clathrin-mediated endocytosis by hampering the release of clathrin-coated pits from the plasma membrane with consequent slowing of vesicle intracellular trafficking and accumulation of clathrin-coated structures where the viral particles remain trapped ( ). finally, based on structural similarities between the umifenovir binding sites in the hemagglutinin of the h n influenza virus and the s glycoprotein of sars-cov- , it has been suggested that this drug might block the trimerization of the s glycoprotein, which is essential for the virus cell adherence and entry ( ). umifenovir is licensed (only in russia and china) for the prophylaxis and treatment of influenza a and b infections but it has shown in vitro activity against infections by hepatitis c and b (hcv and hbv), ebola and other viruses ( ). in a clinical pilot trial conducted in sixty-nine covid- patients, oral treatment with umifenovir (n= ) showed a tendency to reduce viral load and mortality rate as compared to the control group receiving interferon or other non-specified antiviral agents ( % vs %) ( ) . the results of a retrospective cohort study in patients with covid- , without invasive ventilation, who received umifenovir plus lopinavir/ritonavir (n= ) or lopinavir/ritonavir only (n= ), showed a potential benefit of the triple combination therapy to reduce viral load and delay disease progression. in fact, after days of treatment, in the umifenovir-treated group nasopharyngeal specimens were negative for sars-cov- in % of patients (vs % of the control group) and the chest computed tomography (ct) scans were improved in % of cases (vs % of the control group) ( ) . subsequently, umifenovir was tested as monotherapy (n= ) and its activity compared to that of lopinavir/ritonavir (n= ). on day after treatment, no viral load was detected in the umifenovir group, whereas the virus was still found in . % of patients treated with lopinavir/ritonavir ( ) . conversely, in another study with non-icu patients (n= ) umifenovir failed to improve the prognosis and virus clearance compared to the control group receiving symptomatic treatment, including the most appropriate supportive care (n= ) ( ) . a similar conclusion was drawn by an observational cohort study on the real-world efficacy and safety of umifenovir used as single agent or in combination with lopinavir/ritonavir. there was no evidence that adding umifenovir to lopinavir/ritonavir could shorten the time to negative conversion of sars-cov- nucleic acid in pharyngeal swabs or improve the symptoms ( ). however, a retrospective analysis of adverse drug reactions in chinese patients with covid- , by a hospital pharmacovigilance system, reported a lower incidence of adverse effects (that were mostly at the gastrointestinal and hepatic level) for umifenovir compared to lopinavir/ritonavir ( . % vs . %) ( ) . a small retrospective cohort study has recently suggested the use of umifenovir for post-exposure prophylaxis, based on the significant reduction of infection risk observed in family members (n= in families) and health care workers (n= ) who were exposed to patients with confirmed sars-cov- infection ( ) . further clinical studies are ongoing to evaluate the role of umifenovir in covid- management, used as monotherapy [nct ] or in combination with other antiviral agents [nct , nct ]. in the randomized, double-blind, placebo-controlled clinical nct trial, umifenovir is added to a therapeutic regimen including interferon-β a, lopinavir/ritonavir and a single dose of hydroxychloroquine plus standard of care. baricitinib is a potent and selective inhibitor of the janus kinases / (jak /jak ), currently used in the therapy of rheumatoid arthritis. based on the results of a benevolentai's knowledge graph, the small-molecule kinase inhibitor baricitinib was predicted to alter virus entry by inhibiting ap -associated kinase (aak ) and cyclin g-associated kinase (gak), which are likely involved in sars-cov- endocytosis ( ) . the benevolentai's knowledge graphical method uses machine learning to integrate the scientific information on the biological processes involved in viral infection with that on the mechanisms of action of available drugs in order to identify potential new pharmacological targets and therapeutic indications. besides exerting potential direct antiviral effects, baricitinib might prevent the dysregulated production of pro-inflammatory cytokines typically observed in covid- patients via the inactivation of interleukin- (il )-jak-signal transducer and activator of transcription (stat) pathway (this activity will be more deeply discussed in section , especially regarding the jak inhibitor ruxolitinib). some clinical trials, also including placebo-controlled studies, are evaluating the safety and efficacy of baricitinb, mostly as - week add-on therapy in patients with mild to moderate covid- . results from a small study in patients with moderate covid- pneumonia, treated with baricitinib in combination with lopinavir/ritonavir [nct ], indicated that a -week oral treatment with the jak / inhibitor was well tolerated. moreover, although proper control groups were missing, the authors reported improved clinical and laboratory parameters ( ) . a favorable clinical course was also reported in an -year-old woman, with a mild-to-moderate covid- , chronically treated with baricitinib for rheumatoid arthritis, who also received other pharmacological treatments to control viral infection (i.e., lopinavir/ritonavir, hydroxychloroquine) ( ) . this patient was part of a familiar cluster of covid- , and the three other family members (husband, son and daughter) received the same antiviral therapy with the exception of baricitinib. interestingly, the patient's husband ( - chloroquine and hydroxychloroquine are among the most frequently used drugs for the treatment of covid- patients in view of their potential inhibitory activity on virus entry. however, other mechanisms appear to contribute to their antiviral activity, including impaired receptor recognition by coronaviruses due to altered terminal glycosylation of ace ( ) and inhibition of viral attachment to the lipid raft as a consequence of a reduced interaction of the sars-cov- s protein n-terminal domain with membrane gangliosides ( ). indeed, in vitro studies have demonstrated that chloroquine is able to block sars-cov- infection at lowmicromolar concentrations ( ) ( ) ( ) . furthermore, chloroquine and hydroxychloroquine exhibit immunomodulatory activity since they reduce the toll-like receptor (tlr) signaling (that plays a crucial role in the innate immune system) and production of inflammatory cytokines, as well as the expression of co-stimulatory molecules in t cells (for a comprehensive review see ) . so far, however, there is not conclusive or robust clinical evidence on the usefulness of quinolines in covid- . starting from mid-february, , chloroquine was included in the sixth version of the covid- treatment guidelines by the national health commission of the people's republic of china. according to these guidelines the initial recommended chloroquine dose was mg twice daily for no more than days; however, due to safety concerns the maximum therapy course was reduced to days and a lower dose was recommended for patients weighing less than kg. based on clinical trials conducted in china in more than hospitals, treatment of > patients with chloroquine is superior to control treatment in preventing pneumonia exacerbation, improving lung imaging results, accelerating virus-negative conversion, and shortening the disease course ( ) . however, detailed information on the study design, patient characteristics or control treatment were not provided. the results of a blinded, randomized, controlled chinese trial for covid- pneumonia, reported a significant improvement in terms of symptoms and ct findings in patients treated with hydroxychloroquine (n= ; mg/day for days) compared to the control group (n= ) ( ) . conversely, in a previous pilot study in treatment-naïve patients with confirmed covid- , hydroxychloroquine did not show any clinical benefit ( ) . a french study on a cohort of patients with severe covid- treated with hydroxychloroquine ( mg/day for days plus the macrolide antibiotic azithromycin for days) did not reveal antiviral activity or clinical benefit ( ). a published interim analysis of a double-blind, randomized, phase iib clinical trial [nct ] performed in brazil, after enrollment of the first patients with severe ards treated with high and low chloroquine doses (i.e., mg/twice/day for days, n= ; mg twice daily on day and once daily for days, n= ) indicated that the high-dosage group showed a higher incidence of cardiotoxic effects (qtc interval prolongation) and a higher mortality rate compared to the low-dosage group ( % vs %) ( ). all these patients also received the macrolide antibiotic azithromycin that may induce cardiotoxic effects. these preliminary data indicate that high chloroquine dosage should not be recommended for treating critically ill covid- patients. although hydroxychloroquine is better tolerated than chloroquine, both agents may cause in the long-term life-threatening arrhythmias (an effect increased by the concomitant use of azithromycin), leucopenia, neuropsychiatric effects and retinopathy. in addition, quinoline overdose can lead to cardiovascular collapse, seizures and coma ( ). therefore, the use of chloroquine/hydroxychloroquine for covid- management requires a careful patient selection and monitoring. based on the initial publication in the lancet of the results of a multinational registry analysis conducted by surgisphere corporation, showing that treatment with hydroxychloroquine/chloroquine (with or without a macrolide) in hospitalized covid- patients (n= , ) failed to induce clinical benefit and was associated with higher risk of death and cardiovascular complications compared to control treatment (n= , ) ( ), on may , , the who temporarily halted the solidarity trial arm with chloroquine/hydroxychloroquine. thereafter, the article was retracted by three of the four coauthors of the original article since surgisphere (owned by one of the authors) did not make available to a third-party audit the complete dataset used for the study ( ) . thus, on june , , the who announced that there was no reason to modify the solidarity trial protocol and the arm with quinolines was resumed. nevertheless, on the basis of a low benefit/risk ratio, the fda retracted the emergency use authorization (eua) previously issued to hydroxychloroquine for use in covid- hospitalized patients outside of clinical trials. to have a clear view on the overall risk-benefit ratio of using chloroquine/hydroxychloroquine especially in severely ill covid- patients we will have to wait for the conclusion of welldesigned, multi-center, randomized, controlled studies. actually, clinicaltrials.gov lists a number of phase studies testing chloroquine and more frequently hydroxychloroquine, alone six of them also received azithromycin ( mg on the first day followed by mg daily) to prevent bacterial infection. all patients treated with both drugs showed negative nasopharyngeal sars-cov- pcr conversion compared to . % of those treated with hydroxychloroquine as single agent and . % of the untreated ones ( ) . the results of a french retrospective non-randomized study in a total of patients treated for at least days with hydroxychloroquine plus azithromycin showed that early treatment with this drug combination was well-tolerated and associated with a very low fatality rate ( . %) ( ) . the mechanism underlying the potential azithromycin activity against sars-cov- still needs to be clarified; recently, it has been hypothesized that this antibiotic might inhibit cd , a glycosylated transmembrane protein that would serve as additional receptor for sars-cov- cell invasion ( ) . furthermore, azithromycin might stimulate immune responses against the virus by inducing the synthesis of type i and iii interferons, as demonstrated in epithelial cells collected from patients with chronic obstructive pulmonary disease ( ) . as mentioned above, a number of clinical trials are currently evaluating azithromycin mostly in combination with hydroxychloroquine for the treatment of covid- or as prophylaxis. another approach to inhibit sars-cov- infection consists in inhibiting the protease that cleaves the s protein, thus facilitating viral entry and activation. tmprss is an androgen- ( , ) . in addition, nafamostat is able to inhibit the coagulation and fibrinolytic systems, the kallikreinkinin system, the complement cascade, and activation of protease-activated receptors ( ) . therefore, their anti-inflammatory, anti-coagulant and fibrinolytic properties might contribute to attenuate the symptoms and complications occurring in covid- patients. both agents are approved in japan for the treatment of pancreatitis, and nafamostat is also used for disseminated intravascular coagulation and as anticoagulant in extracorporeal circulation. three case reports of elderly covid- patients with pneumonia, all taking antivirals like lopinavir/ritonavir and hydroxychloroquine, showed that the introduction of nafamostat induced clinical and radiological improvement without significant adverse effects ( ) inhibition of the viral spike protein cleavage by cathepsin l in the late endosome might also result in decreased sars-cov- entry into the cells ( ) . once sars-cov- reaches the endosomes, the cysteinyl proteinase cathepsin l is the main protease that cleaves the s once inside the cell, sars-cov- , like other coronaviruses, uses two third of its positivesense single-stranded rna genome as template to directly translate two open reading frames (orf a and orf ab), connected by a ribosomal frameshift site, into the two overlapping polyproteins, pp a and pp ab, which are afterward cleaved by viral proteases into nonstructural proteins (nsps) ( ) . some nsps (including rna-dependent rna polymerase, helicase and other enzymatic activities required for the mrna capping and proofreading) eventually contribute to form the replication-transcription complex, which is anchored to double-membrane vesicles integrated into a reticulovesicular network of modified endoplasmic reticulum membranes, also including convoluted membranes ( , ) . the viral genomic rna is encapsulated by the nucleocapsid n protein that thereafter buds into the ergic and acquires a membrane containing the s, e and m structural proteins. finally, the virus is released by exocytosis ( figure ). the cl pro /m pro is highly conserved among various coronaviruses, and mutations in cl pro /m pro are often lethal to the virus ( , ) . therefore, cl pro /m pro is indispensable for viral replication and thus represents an attractive therapeutic target for inhibiting the coronavirus infection process ( ) . this enzyme is a homodimeric cysteine protease whose recognition sequence at most sites of viral polyproteins is leu-gln↓(ser,ala,gly). several previous reports have indicated that the hiv aspartate protease inhibitors lopinavir and ritonavir have the potential to act also as sars-cov protease inhibitors through their binding to cl pro /m pro ( ) ( ) ( ) ( ) . for hiv treatment, the two drugs are used in combination, but ritonavir is administered at a dose that does not affect hiv protease activity but rather inhibits the cytochrome p a -mediated metabolism of lopinavir, thus increasing its plasma levels. both drugs bind to amino acid residues present at the active site of sars-cov- furthermore, based on a virtual docking prediction study, some hcv ns / a protease inhibitors (e.g., simeprevir, paritaprevir, grazoprevir, boceprevir, telaprevir) might also inhibit the cl pro /m pro ( , ) . however, none of these agents is currently clinically evaluated for covid- treatment. concerning the other coronavirus protease pl pro , although considered another potential therapeutic target since it is crucial for viral replication, the development of sars-cov- pl pro inhibitors is still at an early stage ( ), despite several investigational compounds have been found to efficiently inhibit the corresponding sars-cov and mers-cov enzyme ( ). another the adenosine analogue remdesivir is one the most frequently tested anti-sars-cov- agents and has been firstly approved in japan for severe covid- . remdesivir was originally developed for rna virus infections and tested for ebola during the outbreak in democratic republic of the congo but failed to show clinical benefit. it has a broadspectrum antiviral activity, including mers-cov, sars-cov and sars-cov- , both in vitro and in vivo in animal models ( , ( ) ( ) ( ) ( ) . remdesivir is a prodrug that, after diffusion into the cells, is metabolized to the alanine metabolite gs- and further converted into a nucleoside monophosphate, which is highly polar and remains trapped within the cell ( ) . host cell kinases eventually convert the monophosphate derivative into a triphosphate nucleotide that is misincorporated into the nascent rna chain by the rna dependent rna polymerase with consequent inhibition of the rna synthesis ( ) . remdesivir has been found to interact with the sars-cov- polymerase, competing with the physiological atp nucleotide, and to behave as delayed-chain-terminator, since rna synthesis is terminated after the addition of three nucleotides ( , , ) . it should be noted that the efficacy of remdesivir or of other nucleoside/nucleotide-based agents, whose activity relies on their misincorporation into the viral genome, might be counteracted by a coronavirus proofreading exoribonuclease (nsp ) that would enable the virus to evade the pharmacological inhibition ( ) . intravenous remdesivir was used to treat the first covid- patient diagnosed in the us with rapid improvement of the clinical conditions ( ) and is regarded as one of the most promising agents for sars-cov- . presently, the drug is included in the national institutes ( ) . in an uncontrolled study where patients ( % receiving mechanical ventilation) were treated with remdesivir on a compassionate-use basis, clinical improvement at days was observed in % of patients ( ) . however, this trial raised several criticisms on the study design and result interpretation, due to lack of control, small sample size, inappropriate data censoring, high variability of disease severity ( ) ( ) ( ) ( ) . in another study, remdesivir was administered as compassionate treatment to hospitalized patients ( of whom in icu) and beneficial effects were observed on sars-cov- pneumonia, mainly in non-critically ill patients ( ) . remdesvir is usually well-tolerated for short courses. however, concerns were raised about its potential toxicity in patients with kidney dysfunction related not only to the drug-mediated injury of renal tubular epithelial cells, but also to the nephrotoxicity associated with the drug vehicle (i.e., sulfobutylether-β-cyclodextrin) required for the intravenous formulation ( ) . another nucleoside analogue used for covid- is favipiravir, which acts as a competitive inhibitor of the rna-dependent rna polymerase. this agent was previously approved in japan for the treatment of influenza a and b and, in particular, for novel or re-emerging influenza viruses ( ) . presently, the drug has been approved in russia for covid- and is investigated worldwide. favipiravir, after undergoing intracellular tri-phosphorylation, exerts antiviral effects as a guanosine analogue, through several mechanisms including chain termination, slowed rna synthesis and lethal mutagenesis (due to c-to-u and g-to-a transitions favored by the low cytosine content of sars-cov- genome) ( high levels of il and il also contribute to hypercoagulation due to activation of the complement and coagulation cascades, causing disseminated intravascular coagulation ( , ) . waiting for an effective antiviral therapy or vaccine against the virus, any treatment that can decrease the severe symptoms of covid- may help to attenuate the mortality rates and to improve the quality of life of severely ill patients. in this regard, several pharmacological therapies, with different mechanisms of action, have been used in order to improve the symptoms related to covid- . in the next section, we will consider the agents directed against the: a) cytokine storm and b) cardiovascular damage. the first category includes: i) anti-cytokine drugs, such as tocilizumab, sarilumab, siltuximab, olokizumab, ruxolitinib, baricitinib, anakinra, emapalumab, mavrilimumab; and ii) immunomodulating agents, such as interferon-β, interferon-α or interferon-λ, fingolimod, ozanimod, opaganib, cd fc, allogenic mesenchymal stem cells and the lately reappraised dexamethasone. the second group comprises: the anti-c complement monoclonal antibodies (mabs) eculizumab and ravulizumab; anti-thrombotic and fibrinolytic agents; the phosphodiesterase type inhibitor sildenafil; the vasoactive intestinal polypeptide analog aviptadil; and the anti vegf-a mab bevacizumab ( figure ) . however, the effects of the two drug sets are closely interconnected, as many drugs that have an impact on the circulatory system may also reduce circulating inflammatory cytokines. the first therapeutic agent used to counteract the inflammatory reaction in patients with ( ) . furthermore, a larger study on patients with covid- pneumonia accompanied by hyperinflammatory syndrome and acute respiratory failure described improvement or stabilization of the respiratory conditions in % of patients ( ) . case reports also supported the potential benefit from tocilizumab treatment ( ) . a chinese small retrospective study in patients showed that tocilizumab is associated with rapid improvement of the clinical symptoms and hypoxemia, and prevention of clinical worsening in severe covid- patients, without serious adverse events ( ) . however, tocilizumab did not reduce icu admission and mortality rates in critically ill patients with severe covid- pneumonia ( ) . tocilizumab treatment was found to be associated with an initial rise in il levels, as an expected consequence of the mab-mediated inhibition of il interaction with its receptors, followed by a significant decrease of the c-reactive protein inflammatory marker ( ). however, the d-dimer, measured as an indicator of intravascular fibrin formation, remained unaffected suggesting that tocilizumab might have limited effect on the activation of the coagulation cascade ( ) . furthermore, the clinical response to tocilizumab seems to be negatively affected by hyperglycemia, shown to be associated with increased il levels ( ) . in regard to tocilizumab safety, concerns have been raised about the risk of candidemia, septic shock and possible occurrence of intestinal perforation (an adverse effects reported in rheumatoid arthritis patients), which may be favored by the altered hemodynamics observed in critically ill covid- patients ( , ( ) ( ) ( ) . based on these initial data, the efficacy and safety of tocilizumab in severe covid- need to be corroborated by the results of the ongoing randomized controlled clinical trials. clinical studies are also testing the other anti-il receptor mab sarilumab (fda-and emaapproved for rheumatoid arthritis) and anti-il mabs such as siltuximab (fda-and ema-approved for multicentric castleman's disease) and the investigational mab olokizumab. like tocilizumab, sarilumab is able to bind both il r and sil r, but being a fully human mab, has a lower risk of inducing neutralizing antibodies and allergic reactions compared to chimeric/humanized mabs ( strictly dependent on the jak/stat pathway ( , ) , jak inhibitors have also been used in covid- patients with the aim of reducing the excessive inflammatory reaction. among these, the orally administered ruxolitinib is a jak /jak small-molecule inhibitor approved for the treatment of myelofibrosis, polycythemia vera, and graft-versus-host disease ( ) . consistently with its mechanism of action, in patients with myelofibrosis, ruxolitinib was able to reduce il and tnfα levels and was well-tolerated ( ( ) . furthermore, in covid- patients two case reports of diffuse skin reactions with purpuras and a rapid decrease of hematocrit values were described ( ) . baricitinib is another jak inhibitor that besides interrupting the jak / -dependent signaling involved in cytokine-mediated inflammatory response to the sars-cov- infection, might also exert direct antiviral effects by blocking virus entry (see section ). anakinra is a recombinant, non-glycosylated form of the natural occurring human interleukin- interferon-γ, which inhibits its binding to cell surface receptors and the subsequent activation of intracellular pro-inflammatory signaling pathways. emapalumab is fda-approved to treat the severe inflammatory condition of primary hlh in which serum interferon-γ levels are elevated ( ). blockade also immune responses against viruses, through enhancing antigen presentation, costimulation, and cytokine production by the effector cells of innate immune system, leading to enhanced adaptive immune responses ( ) . the response mediated by type i interferons is more potent, rapid, transient, diffuse and inflammatory, whereas the type iii interferon response is less potent, slower, sustained, anatomically restricted and less inflammatory ( ) . interestingly, the cytokine storm associated with covid- is due to an uncontrolled response of the immune system to sars-cov viral infection that leads not to only to an excessive production of cytokines but also a diminished/delayed interferon response ( ) ( ) ( ) . based on preclinical studies and observations in sars-cov or mers-cov infected patients, the outcome of the interferon-mediated response to the viral infection seems to depend on the viral load and integrity of the host immune system. in particular, if the initial viral burden is low, type i interferons are promptly released and efficiently clear the infection; conversely, if the viral load is high or in elderly patients the early interferon production is hampered and a delayed interferon-mediated response may not only fail to control the infection but also result in inflammation and lung damage ( ) . thus, exogenously administered type i interferons would have protective effects as prophylaxis or in the early stage of sars-cov- infection, whereas they may deteriorate tissue injury and pneumonia when administration is delayed. concerning the potential therapeutic role of interferon-λ, the lack of pro-inflammatory systemic effects would allow its safe administration also in an advanced phase of the infection ( ) ( ) ( ) . although better tolerated than type i interferons that may cause severe systemic side effects due to the ubiquitous expression of ifnar, a possible disadvantage of interferon-λ is its lack of antiviral effects on infected alveolar macrophages or endothelial cells that do not express ifnlr but may serve as virus reservoir ( ) . . in an additional trial, interferon-α b as nasal drops is assessed for low-or high-risk medical staff exposed to sars-cov infected patients, as single agent or combined with thymosin α , respectively [nct ]. in regard to interferon-λ, the safety and efficacy of subcutaneous pegylated interferon-λ is tested as immediate/early therapy in non-critically ill hospitalized or ambulatory patients another immunomodulating agent, fingolimod (fty ), an orally administered drug approved for multiple sclerosis, was evaluated in covid- patients ( ) . once absorbed, fingolimod undergoes phosphorylation to form an analog of the naturally occurring s p, a lipid signaling molecule whose activity is mediated by the interaction with four subtypes of g protein-coupled receptors (s p and s p - ) ( , ) . after binding to s p , fingolimod initially activates the receptor and thereafter down-regulates its expression, causing retention of naïve t cells and central memory t cells in the lymph nodes and induction of lymphocytopenia ( ) . nevertheless, fingolimod does not substantially affect memory effector t cells, which play an important role in the defense against infectious agents ( ) . moreover, it does not affect humoral immune responses and does not prevent the generation of virus-specific cytotoxic t cells in the lymph nodes ( ) . thus, it has been hypothesized that patients on s p modulators might have a reduced risk of complications from sars-cov- infection. furthermore, due to the s p role in lung endothelial cell integrity ( ) , in covid- patients, fingolimod might reduce vascular permeability and consequent lung injury ( ) . in clinical trials with fingolimod for multiple sclerosis, conflicting results were reported showing either no change or increased risk of viral infections (especially herpes virus) compared controls ( ) ( ) ( ) . severe covid- cases have been reported in patients with multiple sclerosis on treatment with fingolimod that was stopped upon sars-cov- diagnosis; in all these cases patients fully recovered from infection ( , , another molecule that has raised some interest to control the inflammatory response associated with sars-cov- infection is cd fc, a recombinant fusion protein that comprises cd attached to the fc region of human igg . cd is a glycosylated membrane protein expressed in hematopoietic cells (including immature b and t cells, granulocytes, macrophages and some epithelial cells) that plays a regulatory role on b and t cell homeostasis ( ) . in humans, cd is able to suppress inflammation upon interaction with the prr siglec and several danger-associated molecular patterns (damps), helping to reduce the host immune response against proteins released by damaged cells. preclinical studies demonstrated that the chimeric molecule cd fc mitigates the graft-versus-host disease, by decreasing the overall inflammatory response, and, in particular, the release of il β, il and tnfα release ( ) . these data provided the biological rationale for the clinical testing of cd fc also for covid- , and a randomized, double-blind, placebocontrolled, phase study is currently recruiting severely ill infected patients [nct ]. a cell-based approach to modulate the damage deriving from inflammation and altered activation of dexamethasone is an old corticosteroid, i.e., a drug with broad anti-inflammatory and immunosuppressant activity that reduces cell-mediated immunity and various cytokine production. its clinical indications span from pain in the joints to asthma, irritable bowel disease/crohn disease, emesis, multiple sclerosis and various autoimmune diseases, as well as different types of cancer, to name just the most prevalent. it has also been used in the previous although the clinical manifestations of covid- are dominated by respiratory symptoms, the disease prognosis is largely influenced by the involvement of various organs, including the heart. cardiovascular complications (i.e., myocardial infarction, acute heart failure and cardiomyopathy, shock and cardiac arrest, dysrhythmias, venous thromboembolic events, acute myocarditis) are associated with a high mortality rate and occur in about % of hospitalized patients ( ) . furthermore, patients with pre-existing cardiovascular diseases are predisposed to sars-cov- -induced myocardial injury and infection is associated with a high mortality rate; in these patients, the risk of heart failure and myocardial damage increases to ≥ % ( ) ( ) ( ) . the mechanisms involved in the cardiovascular injury of covid- include: i) direct damage upon virus entry through ace present in coronary endothelial cells, cardiomyocytes and cardiac fibroblasts; ii) increased oxygen consumption deriving from fever, enhanced adrenergic tone and tachycardia; iii) increased oxidative stress as a result of ros production; iv) massive cytokine release and a state of hyperinflammation that contribute to pneumonia/ards with consequent acute heart failure, as well as endotheliitis leading to disseminated intravascular coagulation, thrombosis and infarction; v) sars-cov- -induced ace downregulation due to receptor shedding or internalization with consequent increase of angiotensin ii ( , ) . in fact, normally, ace degrades angiotensin ii to produce angiotensin - that is endowed with vasodilating and anti-inflammatory effects. therefore, a reduction in ace function after viral infection may result in a dysfunctional renin-angiotensin system, associated with an increase of angiotensin ii, which would lead to vasoconstriction and inflammation. dysregulated immunothrombosis (i.e., clot formation triggered by the interaction of innate immune system components, like macrophages, neutrophils and the complement system, with platelets and coagulation factors, that provides a first line defense against infectious agents) with diffuse microvascular thrombi formation has been also described in covid- and involved in multi-organ damage ( , in patients with severe covid- , high rates of venous thromboembolism and disseminated intravascular coagulation, due to dysregulation of the coagulation and fibrinolytic systems are furthermore, heparin has anti-inflammatory properties by inhibiting il , il , tnfα release, c-reactive protein and adhesion of neutrophils to endothelial cells ( ) ( ) ( ) . in a retrospective cohort study, the administration of lmwh to covid- patients besides producing anticoagulant effects also reduced il levels and increased lymphocyte counts suggesting a beneficial effect towards controlling the cytokine storm ( ) . in addition, ufh and lmwh have been shown to inhibit the binding of the s protein of sars-cov- to its cellular receptor, ace , in an in vitro cell system expressing ace and tmprss . these results suggest another mechanism through which heparin may slow down or prevent disease progression in the early phases of covid- . ( , ) . fibrinolytic drugs, namely tissue-type plasminogen activator (tpa) as systemic intravenous treatment or as lung-targeted nebulizer form, have been proposed for covid- patients ( , ) . several clinical trials are currently assessing different heparin regimens, other anticoagulants, systemic and local fibrinolytic approaches, and antiaggregants (e.g., rivaroxaban; defibrotide; clopidogrel, aspirin) (clinicaltrials.gov). the pde- inhibitor sildenafil, a vasodilator that is approved for treating erectile dysfunction and pulmonary arterial hypertension ( ) , is also evaluated in a phase trial for patients with mild to severe covid- [nct ]. in fact, sildenafil has a wide range of antiinflammatory, antioxidant, and vasodilatory actions resulting in cardioprotective effects and improved pulmonary circulation ( ) ( ) ( ) . aviptadil is a synthetic form of vip that increases adenosine cyclase activity with consequent smooth muscle relaxation, approved in combination with phentolamine only in certain the vegf-a is considered the most potent inducer of vascular permeability. the potential involvement of vegf-a in covid- has been related to the excessive production of angiotensin ii, consequent to the sars-cov- -mediated down-regulation of ace . in fact, angiotensin ii is able to increase vegf-a expression which in turn may exacerbate inflammation stimulating the recruitment of inflammatory cells and the release of proinflammatory cytokines ( ) ( ) ( ) . numerous studies have confirmed a key role of vegf-a as potential therapeutic target in ali and ards due do the increased vascular permeability and pulmonary edema ( ) . furthermore, vegf-a has been involved in disruption of the blood-brain barrier and may contribute to brain inflammation in the course of sars-cov- infection ( bleeding, delaying wound healing, thromboembolic events). all over the world, the scientific community is racing to evaluate a huge number of drugs or [ , ] (see table ) eidd- inhibition of rdrp -nct nct a nct: clinicaltrials.gov identifier data from clinicaltrials.gov accessed on june, . due to the rapidly evolving situation and the increasing number of clinical trials, the reported list of clinical trials does not mean to be exhaustive. b these agents might also have additional mechanisms contributing to the antiviral activity against sars-cov- rdrp; rna-dependent rna polymerase. human coronaviruses with emphasis on the covid- outbreak, virusdisease. 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severe and lifethreatening covid- : a randomized clinical trial challenges in the production of convalescent hyperimmune plasma in the age of covid- a human monoclonal antibody blocking sars-cov- infection we would like to acknowledge the support of the "fondazione airc" to the national civil protection to help tackle the covid- emergency in italy and its commitment to continue supporting cancer research during the challenging times of sars-cov- pandemic. g.graziani is principal investigator (pi) of the airc grant ig -id. project. orphan drug for the treatment of ards, ali and sarcoidosis nct nct anti-vegf-a bevacizumab cancer treatment; age-related macular degeneration (off-label) nct nct nct a nct: clinicaltrials.gov identifier data from clinicaltrials.gov accessed on june, . due to the rapidly evolving situation and the increasing number of clinical trials, the reported list of clinical trials does not mean to be exhaustive.ards: acute respiratory distress syndrome; ali: acute lung injury. key: cord- -kxnrf g authors: riggioni, carmen; comberiati, pasquale; giovannini, mattia; agache, ioana; akdis, mübeccel; alves‐correia, magna; antó, josep m.; arcolaci, alessandra; kursat azkur, ahmet; azkur, dilek; beken, burcin; boccabella, cristina; bousquet, jean; breiteneder, heimo; carvalho, daniela; de las vecillas, leticia; diamant, zuzana; eguiluz‐gracia, ibon; eiwegger, thomas; eyerich, stefanie; fokkens, wytske; gao, ya‐dong; hannachi, farah; johnston, sebastian l.; jutel, marek; karavelia, aspasia; klimek, ludger; moya, beatriz; nadeau, kari; o'hehir, robyn; o'mahony, liam; pfaar, oliver; sanak, marek; schwarze, jürgen; sokolowska, milena; torres, maría j.; van de veen, willem; van zelm, menno c.; wang, de yun; zhang, luo; jiménez‐saiz, rodrigo; akdis, cezmi a. title: a compendium answering questions on covid‐ and sars‐cov‐ date: - - journal: allergy doi: . /all. sha: doc_id: cord_uid: kxnrf g in december , china reported the first cases of the coronavirus disease (covid‐ ). this disease, caused by the severe acute respiratory syndrome‐related coronavirus (sars‐cov‐ ), has developed into a pandemic. to date it has resulted in ~ . million confirmed cases and caused almost , related deaths worldwide. unequivocally, the covid‐ pandemic is the gravest health and socio‐economic crisis of our time. in this context, numerous questions have emerged in demand of basic scientific information and evidence‐based medical advice on sars‐cov‐ and covid‐ . although the majority of the patients show a very mild, self‐limiting viral respiratory disease, many clinical manifestations in severe patients are unique to covid‐ , such as severe lymphopenia and eosinopenia, extensive pneumonia, a “cytokine storm” leading to acute respiratory distress syndrome, endothelitis, thrombo‐embolic complications and multiorgan failure. the epidemiologic features of covid‐ are distinctive and have changed throughout the pandemic. vaccine and drug development studies and clinical trials are rapidly growing at an unprecedented speed. however, basic and clinical research on covid‐ ‐related topics should be based on more coordinated high‐quality studies. this paper answers pressing questions, formulated by young clinicians and scientists, on sars‐cov‐ , covid‐ and allergy, focusing on the following topics: virology, immunology, diagnosis, management of patients with allergic disease and asthma, treatment, clinical trials, drug discovery, vaccine development and epidemiology. over questions were answered by experts in the field providing a comprehensive and practical overview of covid‐ and allergic disease. the first cases of the coronavirus disease (covid- ) , caused by the novel severe acute respiratory syndrome-related coronavirus (sars-cov- ), were reported in china in december and rapidly led to pandemic. currently, ~ . million confirmed cases of covid- and near , covid- -related deaths have been reported globally. these numbers, which are still rising, likely underestimate the cumulative incidence of covid- due to several factors; these include limitations of current diagnostic tests, the extent of population testing and reporting, and the type and timing of community mitigation strategies adopted by each country, among others. covid- shows a complex clinical profile with many different presentations. like in many other viral infections, subclinical, mild, moderate, or severe cases ( - % of patients require hospitalization and - % intensive care unit, icu) presenting with or without pneumonia are observed. asymptomatic cases are common but, to date, there is a lack of epidemiological surveys that provide a clear percentage of asymptomatic cases. , the covid- pandemic is the world's gravest public health crisis of the st century, and there is an urgent need for reliable and updated scientific and clinical information. covid- is a zoonosis to cd (known as basigin or extracellular matrix metalloproteinase inducer), which is expressed in human airway and kidney epithelium, as well as in innate cells and lymphocytes, and to tmprss , which is highly expressed in intestinal epithelial cells. in addition, antibody-dependent enhancement of sars-cov- cell entry may also contribute to infection as reported for sars-cov. sars-cov- may use receptors that have been reported for other coronaviruses, such as cd , aminopeptidase n and glutamyl aminopeptidase for cell invasion. , , among these, cd (encoded by dpp ) has emerged as a putative receptor for sars-cov- because structural analyses predict that the spike protein of sars-cov- binds to cd . this receptor has been shown to be expressed in the human epithelium and immune cells. there is limited evidence about covid- -associated polymorphisms. ace might be one of the candidate genes that influences pneumonia progression in sars. it is conceivable that the d allele influences the renin-angiotensin system via elevation of serum or local ace levels, which may damage the endothelium or epithelium of the lungs. the variance in covid- prevalence and mortality cannot be explained by an ace insertion or deletion polymorphism alone, or one polymorphism of any single gene. however, polymorphisms in genes of toll-like receptors, inflammasome, intracellular molecular sensors, interferons (ifns) and interleukins (ils) may contribute. structural proteins of sars-cov- virions, such as the spike glycoprotein, envelope, membrane and nucleocapsid, are the main immunogenic molecules (figure ) . , sars-cov- adaptive responses develop mainly to the spike protein, and immunodominant t and b cell epitopes have been reported. intracellularly, the viral rna replicase complex, and non-structural and translated proteins, activate innate immune pathways. this leads to an ifn type i response, nf-kb activation in epithelial cells, as well as activation of nlrp and other inflammasomes, in macrophages and dendritic cells. this article is protected by copyright. all rights reserved the spike protein of sars-cov- has a receptor-binding domain that binds ace with higher affinity than sars-cov. in addition, the sars-cov- spike protein harbors a polybasic furin cleavage site (prrar) with an insertion of amino acid residues, which is distinct from that found in sars-cov and other sars-like viruses. this allows effective cleavage by furin and other proteases and determines viral infectivity and host range. the severe lymphopenia observed in covid- is similar to that reported in hiv infection and acquired immune deficiency syndrome. the latter is characterized by cd + t cell lymphopenia, whereas covid- causes general lymphopenia. however, severe lymphopenia development in covid- happens in weeks, whereas hiv-induced lymphopenia takes years. hiv and sars-cov- are both rna viruses and share some similarities in their replication pathways; hence certain rna replication drugs may work in both diseases (figure ) . there are strains of sars-cov- that are clinically relevant. genome analysis of sars-cov- from human samples shows high rates of mutation and deletion in several viral genes, including the spike-glycoprotein gene. covid- treatments, including future vaccination against sars-cov- , may drive the genetic evolution of the virus affecting virulence and pathogenicity. for example, a report on a -nt deletion in orf (figure ) of sars-cov isolated from patients in singapore implied that mutations may arise as a result of human adaptation and could be associated with attenuation. nevertheless, the emergence of a sars-cov- is possible as long as there is close contact between humans and living animals that harbor coronaviruses. data from covid- patients in china show sars-cov- detection in respiratory samples for a median of days ( - days) . in this study, sputum and saliva were not analyzed separately. viral shedding was significantly longer in patients with severe disease, with a median of days ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) ( ) days), compared to mild disease, days ( - days). furthermore, glucocorticoids treatment this article is protected by copyright. all rights reserved longer than days significantly extended the duration of sars-cov- shedding. viral load differed significantly by sample type, with respiratory samples showing the highest, followed by stool samples, and serum samples showing the lowest (figure ) . another study of patients with covid- ( asymptomatic vs symptomatic) has estimated that the duration of viral shedding from nasopharynx swabs was days ( - days) for asymptomatic vs days ( - days) for symptomatic patients. the viral load range from . × copies per ml to . × in sputum of patients who died or survived, respectively. tmprss and tmprss promote sars-cov- infection of ace-expressing human enterocytes causing diarrhea in adults and children. , sars-cov- has been detected in stool samples by reverse transcription polymerase chain reaction (rt-pcr) (figure ) . the median duration of the virus in stool samples ( days, interquartile range - days) was significantly longer than in respiratory samples ( days, - days) . however, sars-cov- released into the intestinal lumen was inactivated by simulated human colonic fluid, and infectious virus was not recovered from the stool specimens of patients with covid- . therefore, the intestine is a potential site of sars-cov- replication, which may contribute to local and systemic illness and overall disease progression but unlikely to contribute to the spreading of covid- . section : immunology of covid- from previous sars studies, it is known that the median seroconversion time for detectable igg was days after infection. detectable levels of sars-specific igg and neutralizing antibodies persisted for up to days. this suggests that there is antibody-mediated protection from sars-cov recurrent infection for up to years. there are inconsistent reports on the humoral response to sars-cov- . one study with covid- patients reported that sars-cov- virus-specific igg and igm peaked - days and - days after symptom onset, respectively. on the other hand, another study of hospitalized covid- patients showed that seroconversion could take up accepted article to days. these discrepancies may be related to the time of sars-cov- diagnosis or the clinical characteristics of each cohort and warrant additional studies. systemic iga responses may play a relevant role in the pathogenesis of covid- . mucosal iga likely exerts a protective role by preventing sars-cov- adherence to epithelial cells. circulatory iga may also contribute to sars-cov- neutralization. in addition, iga has the ability to either promote inflammation, through the formation of immune complexes, or to dampen it via fc-mediated inhibitory itam-signaling. , a seroconversion study in covid- patients has found and association between disease severity and sars-cov- -specific iga levels. these were significantly higher than sars-cov- -specific-igm and -igg levels in critically ill covid- patients. whether this association, previously unseen in sars-cov infection, is due to a protective or detrimental role of iga in covid- remains to be elucidated. preliminary findings indicate that asymptomatic and mild cases of covid- can generate detectable levels of sars-cov- -specific antibodies in serum. however, seroconversion is observed less frequently in asymptomatic compared to mild or severe cases, and many asymptomatic cases yield undetectable sars-cov- -specific antibody responses. , [ ] [ ] [ ] so far, no robust data are available on the qualitative differences in humoral responses between asymptomatic and symptomatic covid- patients. it is not clear which molecular mechanisms underlie the milder symptoms of covid- in children as compared to adults. children may mount a sars-cov- antibody response characterized by more efficient production of the so-called natural antibodies, which arise from activated igm+ memory b cells. these cells, which are more prevalent in children than in adults, presumably produce broadly neutralizing antibodies early during the infection. this article is protected by copyright. all rights reserved b cell receptor-sequencing has been conducted in the blood of covid- patients. naive b cells exhibited little clonal expansion, whereas cd +cd + memory b cells showed the highest expansion levels among diverse b cell subsets. covid- patients significantly expanded specific bcell receptor clones compared to those in the healthy controls. these findings suggest that b cells experience unique clonal variable, diversity, and joining gene segment rearrangements upon sars-cov- infection. the lifespan and functionality of these b cells remain to be elucidated. the term "herd immunity" refers to the generation of population immunity that protects a region, or country, from infection. the number of confirmed covid- cases has reached approximately . million. the world population is estimated to be . billion. to ascertain the extent of herd immunity, it is pivotal to define the prevalence of sars-cov- -exposed humans. it is thought that % is the minimum percentage of symptomatic or asymptomatic covid- population required for herd immunity. that is to say that worldwide herd immunity may occur when ⁓ billion humans have a protective immune response against sars-cov- . to date, there are no reliable data, particularly on the number of asymptomatic individuals that show seroconversion, to determine the degree of herd immunity. il- is pleiotropic and could theoretically cause negative effects on immune responses. however, based on phase ii and iii studies with dupilumab (an il- rα-specific monoclonal antibody that blocks il- and il- signaling) in the context of atopic dermatitis, chronic rhinosinusitis with nasal polyps and asthma, no increased risk of infections to viral or bacterial pathogens have been documented. furthermore, dupilumab had no impact on responses to non-live vaccines. this article is protected by copyright. all rights reserved allergic airway disease patients appear to be underrepresented among covid- patients. - this could be partly attributed to the low ace expression detected in allergic patients, with or without concomitant asthma. furthermore, allergen challenge, which induces t helper (th)- inflammation, has been shown to reduce ace expression in a murine model of asthma, and ace expression was inversely associated with type biomarkers (il- , ige, exhaled nitric oxide fraction). these results are in line with previous work showing that decreased ace expression in the airway epithelium of asthmatic subjects was associated with eosinophilic inflammation. on the other hand, the analysis of nasal airway transcriptome data from children identified that tmprss is highly upregulated by type inflammation through the action of il- . therefore, the reduced ace expression seen in asthmatic patients may be compensated by an increase in tmprss production. eosinopenia has been reported in ⁓ - % of severe covid- patients. a minority of covid- patients present with eosinophilic inflammation. , the th /th cytokine balance may play a role, particularly as it pertains to il- , which promotes eosinophilopoiesis and eosinophil survival and activation. eosinophilic inflammation suggests the dominance of type inflammation, which may play a protective role against sars-cov- . on the other hand, it may be the result of a hypersensitivity reaction to drugs used to treat covid- . - anti-il- treatment, which induces eosinophil deficiency, results in a higher viral load in influenza and rhinovirus infection. this might be due to the ability of eosinophils to bind and inactivate the influenza a virus and respiratory syncytial virus (rsv). a similar role seems possible in sars-cov- infection, where type- asthma patients potentially benefit from antiviral eosinophil responses. on the other hand, covid- post-mortems did not show lung eosinophilia , which argues against its local protective role in sars-cov- infection, although it is important to control for glucocorticoid-driven eosinophil reduction in these studies. this article is protected by copyright. all rights reserved eosinopenia is commonly reported in severe covid- . , the underlying mechanisms are largely unknown and most likely multifactorial. a number of possible explanations have been proposed: decreased eosinophilopoiesis; defective eosinophil egression from the bone marrow; and eosinophil apoptosis induced by type ifn released during the acute infection. also, increased eosinophil migration and retention within inflamed tissues has been described, but disputed for the aforementioned reasons. there is no evidence for an enhanced susceptibility of patients on anti-il- /il- r treatment to develop viral infections. observational studies in covid- patients reported elevated eosinophil counts with a favorable outcome, whereas eosinopenia was observed in more severe cases. , neither was there proof of causation nor evidence for enhanced tissue presence in lungs of covid- patients. there is neither evidence for a protective effect of these biologicals nor a negative effect regarding sars-cov- infection. importantly, maintaining proper asthma control is imperative and so is to follow up on severe asthmatics during the covid- pandemic, for example via telemedicine. more than billion people worldwide are infected with helminths, with those living in resource-poor tropical areas being disproportionately affected. helminth co-infection has been shown to influence the severity of viral infection in mice. for example, murid herpesvirus respiratory infection, prior infection with schistosoma mansoni, reduced disease severity. however, immune responses to pulmonary coronaviruses and murid herpesvirus are different and therefore the impact of helminth co-infection is yet to be determined. this is particularly important as the pandemic is now spreading through the helminth-endemic regions of the word. this article is protected by copyright. all rights reserved sars-cov- infects human t cells via cd -binding. t cells are severely affected by sars-cov- , which reduces t cell counts nearly times below the reference limit. this effect is more pronounced in critically ill covid- patients. , , in addition to the reduction in t cell numbers, a recent study found that cd + and cd + t cells as well as natural killer cells displayed reduced antiviral cytokine production in covid- patients. a reduced cytotoxic potential was identified in covid- patients, particularly in those that required icu, and was associated with high il- serum levels. circulating sars-cov- −specific cd + and cd + t cells have been reported in ∼ % and % of covid- convalescent patients, respectively. cd + t cell responses to the spike protein were robust and correlated with sars-cov- -specific-igg and -iga titers. the m, spike and n proteins each accounted for - % of the total cd + response, with additional responses commonly targeting nsp , nsp , orf a and orf , among others. for cd + t cells, spike and m proteins were recognized, with at least sars-cov- orfs targeted. interestingly, sars-cov- −reactive cd + t cells were detected in ∼ - % of unexposed individuals, which indicate cross-reactive t cell recognition between circulating 'common cold' coronaviruses and sars-cov- . three sars-recovered individuals, -and -years post-infection, were analyzed for t-cell responses against sars-cov peptides that may share homology with mers-cov. sarsspecific memory t cells persisted at and years post-sars infection in the absence of antigen exposure. based on these data, it is likely that specific sars-cov- epitopes elicit a persistent t cell response, which may also confer protection against other 'common cold' coronaviruses. however, long-term studies on the natural history of sars-cov- infection are pending. different mechanisms have been proposed for lymphopenia: ) t cell exhaustion. the expression of programmed cell death- marker (also known as pd- ), which is associated with t-cell exhaustion, was higher in t cells from covid- patients than in healthy controls; the expression of pd- and tim- (another exhaustion marker) increased as covid- progressed. ) activation of p signaling in lymphocytes, which suggests a role for apoptosis for in lymphopenia. this article is protected by copyright. all rights reserved pyroptosis, which induces lymphopenia and may be proinflammatory. cov- , which may also cause a cytopathic effect on infected t cells. ) other mechanisms of lymphopenia that remain to be studied are bone marrow suppression during cytokine storm syndrome (css; see below) and sequestration in the lungs during extensive bilateral pneumonia. lymphopenia can be used as an early predictor of severity and clinical outcome. a significant reduction in lymphocyte counts was common in severe and critically ill covid- patients. a continuing or gradual decrease of lymphocyte counts was indicative of poor prognosis and usually required icu admission ( table ) . in agreement with this, a number of studies have identified lymphopenia as an independent risk factor for mortality in covid- . , in covid- patients, decreases were observed in total lymphocytes, cd + and cd + t cells, b cells and natural killer cells. t cell and natural killer cell counts were below normal levels, while b cell counts were at the low end of the normal range. a reduction in specific subsets of lymphocytes, such as cd +cd + natural killer cells and regulatory t cells, was reported in severe covid- patients. css is associated with a wide variety of diseases, both infectious and noninfectious. it is a complex cascade of multicellular activation events that leads to an excessive or uncontrolled release of proinflammatory cytokines. css-associated inflammation begins at a local site and spreads throughout the body via the systemic circulation and can cause multi-organ failure and hyperferritinemia. , css encompasses the activation of large numbers of blood cells, including b cells, natural killer cells, macrophages, dendritic cells, neutrophils, monocytes, resident tissue cells and epithelial and endothelial cells. their activation cause a massive release of pro-inflammatory cytokines, which this article is protected by copyright. all rights reserved drives pathology. the cells involved in css during covid- have not been fully determined yet. were the most important cell types releasing a large amount of proinflammatory cytokines. , which cytokines are most elevated during css? multiple proinflammatory cytokines and inflammasome activation may contribute to css pathogenesis. elevated serum ferritin, il- , il- β, ifn-γ , cxcl (known as ip- ) and ccl immunosuppression is a double-edged sword in viral infections. this article is protected by copyright. all rights reserved primary immunodeficient patients are a high-risk group in the current pandemic, but to date it is unknown if a particular immunodeficiency poses a higher risk of severe disease. international primary immunodeficiency monitoring is being carried out and few cases have been documented. patients at higher risk are those with complications resulting from their primary immunodeficiency and strict follow-up must be done in those cases. a consensus has been established that baseline chronic treatment should be continued in those patients if they are asymptomatic or mildly symptomatic. furthermore, recommendations regarding primary immunodeficient patients adhere to individual national guidelines emphasizing social distancing and strict hygiene measures. systematic testing of primary immunodeficient patients is not advised, however recommendations may change as the pandemic evolves. there are no longitudinal studies analyzing t regulatory cells in covid- systemic dysregulation of metabolism, such as that seen in obesity and diabetes is a risk factor of sars-cov- and sars-cov- infection and of covid- severity . these diseases lead to chronic systemic inflammation, upregulation of sars-cov- receptors in the lungs and the periphery, and they disturb the glucose and lipid metabolism of tissues and immune cells. , , ards is an acute life-threatening inflammation of the lung due to infection, trauma, or inflammatory conditions. excessive inflammation leads to alveolar damage and increased permeability of endothelial and epithelial cells. this results in protein-rich fluid accumulation in the interstitium and the air space, which causes impaired gas exchange and hypoxemia. reactive oxygen species, leukocyte proteases, chemokines, and cytokines also contribute to lung injury. the barrier impairment of the lung microvascular barrier is central to the pathogenesis of ards. in covid- patients, ards is more common in the elderly, those with multiple comorbidities, and those with continuing or gradually progressing neutrophilia and lymphopenia, and a higher level of creactive protein, lactate dehydrogenase, d-dimer and procalcitonin. , there are at least clinical phenotypes of ards: ) near normal pulmonary compliance with isolated viral pneumonia; ) decreased pulmonary compliance. , what specific therapies can be suggested for ards? different treatments were suggested for ards. corticosteroid treatment is generally not recommended, although widely used in critically ill patients. convalescent plasma (cp) was administered to a small number of patients and was associated with virus clearance and clinical improvement ( table ) . low tidal mechanical ventilation, positive end-expiratory pressure, prone positioning ventilation, and fluid management guidelines were associated with improved outcomes. extracorporeal membrane oxygenation could be used according to the inclusion and exclusion criteria of the eolia trial. other potential therapies such as mesenchymal stem cell therapy and cytokine inhibitors are still in trials and without definite results. , bcg is a live attenuated vaccine that was developed against tuberculosis at the beginning of the th century. bcg vaccination induces metabolic and epigenetic modifications by enhancing trained immunity (innate immunity to subsequent infections). it was hypothesized that general bcg vaccination policies adopted by different countries might have impacted the transmission patterns and/or covid- -associated morbidity and mortality. the mechanisms underlying kawasaki disease -a generalized vasculitis, in young children, of unknown, potentially post-viral etiology-are poorly understood. the rare covid- -associated inflammatory syndrome also features vasculitic changes, affects older children too and is often only associated with positive sars-cov- serology, but not viral shedding. its mechanisms need to be elucidated and may include post-infectious, antibody and immune-complex mediated pathology. in adults, there are occasional cases of covid- -associated cutaneous vasculitis, possibly a localized manifestation of the disease that leads to severe generalized vasculitis in some children. [ ] [ ] [ ] interestingly kawasaki-like disease was not reported in chinese cases and the first months of european cases. the season of the disease and environmental factors should be considered. the chinese epidemic was mainly from january to march whereas the usa epidemic started in mid-march and is still ongoing. initial results of acute phase reactants such as c-reactive protein, alanine transaminase, lactate dehydrogenase, d-dimer, procalcitonin, serum ferritin and il- on admission were used to evaluate the severity and predict the mortality. however, dynamic changes of these variables will be more precise in predicting the recovery or progression of covid- . continuing or progressively increasing levels of c-reactive protein, procalcitonin, d-dimer and lactate dehydrogenase were shown to be associated with a high risk of death in severe covid- patients. , , patients with acute respiratory illness (i.e., fever and at least one sign/symptom of respiratory disease such as cough or shortness of breath) and a history of contact with a confirmed or probable covid- case during the days before symptom onset. patients with any acute respiratory illness in the context of a pandemic should have sars-cov- infection in their differential diagnosis. special attention should be given to patients with sudden onset of anosmia, loss of taste, gastrointestinal symptoms or skin lesions without respiratory symptoms who also have epidemiological links. , , smell loss is now a well-established diagnostic symptom of covid- and can be present in otherwise asymptomatic patients, making it a useful tool in initial diagnosis. this has resulted in anosmia to be included in the list of symptoms used in early screening tools for possible covid- in many international bodies. rapidly progressive respiratory failure and sepsis, elevated serum proinflammatory cytokine levels, elevated acute phase reactants (e.g. c-reactive protein), cell-free-hemoglobin-leukopenia and markers of disseminated intravascular coagulation. rt-pcr to generate cdna from sars-cov- rna extracted from respiratory samples, followed by quantitative pcr (figure ). common gene targets for sars-cov- include the envelope, nucleocapsid, spike, rna-dependent rna polymerase, and orf genes. it is recommended to include in the analysis, at least, target genes. nasopharyngeal and oropharyngeal (throat) swabs are the primary specimens for sars-cov- rt-pcr testing. lower respiratory tract specimens (i.e. sputum, endotracheal aspirate or bronchoalveolar lavage) may have higher viral loads and be more likely to yield positive tests ( figure ). however, these locations carry a high risk of aerosolization and therefore should be reserved for severe patients with a negative test on an upper respiratory tract specimen and high suspicion for lower respiratory tract sars-cov- infection. , serology is useful to determine prior exposure to sars-cov- within a given period of time (the length of time following infection that one remains positive is unknown) (figure ) . detection of this article is protected by copyright. all rights reserved antibodies specific to the receptor binding domain of the spike protein indicates neutralization capacity, hence informing better about the development of protective immunity. , , the antibody response occurs later than initiation of symptoms as well as of the detection of viral rna by rt-pcr in respiratory tract specimens, which usually peaks within the first week of symptom onset (figure ) . although antibodies to sars-cov- have been detected as early as the first week after symptom onset, igm, iga and igg seroconversion commonly occurs between the nd and rd week of clinical illness onset. thereafter, igm starts to decline, reaching low levels by week and almost disappears by week , while iga and igg persist beyond this period. , , , the main approaches include nucleic acid amplification on respiratory samples using mobile devices (rt-pcr or isothermal nucleic acid amplification) and viral antigens or host antibodies (viral protein fragments) detection using immunoassays. however, individual tests need validation in large populations before use and their sensitivity, specificity, positive and negative predictive values have to be accurately ascertained. otherwise, they may lead to covid- under or over diagnosis, thus undermining the public health efforts to control the disease. a high rate of false negatives with antigen point-of-care assays may be due to the fact that the majority of patients produce antibodies against sars-cov- only after the second week after of infection ( figure ). furthermore, an effective antibody response is connected with several determinants, comprising severity of the disease, age and nutritional status of the patient, medications administered and concomitant infections. nucleic acid amplification using rt-pcr directly targeting the virus is not affected by the above-mentioned limitations. this article is protected by copyright. all rights reserved positive by the fifth and final test. patients with an initial positive sars-cov- result had an increased risk of progressing to severe cases. altogether, these findings underscore how the timing of the immune response influences rt-pcr tests for sars-cov- , and the importance of combining rt-pcr and seroconversion data for covid- diagnosis. the decision to discontinue home isolation/quarantine should be adapted to specific groups of patients based on factors such as symptom severity, healthcare systems´ capacity, laboratory diagnostic resources and local epidemic status. patients with suspected or confirmed symptomatic covid- can discontinue self-isolation/quarantine if all the following conditions are met: a) resolution of fever (without the use of fever-reducing medications) for at least days; b) clinical improvement in respiratory symptoms (e.g., cough, shortness of breath) for at least days; c) at least days have passed since the onset of symptoms for mild cases or at least days for severe cases and immunocompromised patients; d) negative rt-pcr tests from respiratory specimens taken hours apart. if there is limited or no testing capacity, the combined symptom/test-based strategy should be reserved to hospitalized covid- cases and healthcare workers, whereas for mild or asymptomatic covid- cases (suspected or confirmed) the symptom-based strategy (condition a) and b) and c)) without lab testing is considered acceptable to end the self-isolation period. pandemic strategies for risk minimization should be elaborated, harmonized and followed as such in allergy clinics, centers and practices. in the eaaci/aria position paper by pfaar et al. experts in the field have developed practical recommendations for optimizing allergic patients 'care whilst ensuring the safety of all health care professionals (figure ) . general guidance from national health authorities should be strictly followed (i.e., world health organization, who; european centre for disease prevention). in-person consultations should be minimized to the lowest necessary level and triaged by telemedicine whenever possible (figure ). special attention should be paid to data- protection in adherence to national data-security and -protection laws. non-delayable diagnostic and therapeutic measures should strictly follow reasonable preventive measures. several specific considerations regarding diagnostic and therapeutic measures are important in different allergic diseases ( figure ) . moreover, socio-psychological aspects play a fundamental role in the care of allergic patients during the current pandemic and should be especially recognized and followed. stress caused by isolation and stigmatization due to allergic symptoms may amplify the development of allergic symptoms. virtual doctor consultations have been regarded as an alternative to on-site clinical encounters and are increasing during the covid- pandemic. initially, pre-visit telephonic communication is helpful to screen for patients with potential sars-cov- infection. the epidemiological history should be investigated to determine if patients have fever or respiratory symptoms. in addition, previsit specific triage improves the efficiency of the patient's visit, thus reducing the length of stay in the hospital. to reduce face-to-face meetings, physicians can train some patients to self-treat at home based on the diagnosis obtained through a telephone consultation (figure ) . a strict screening protocol is needed to identify sars-cov- infected patients (figure ) . ideally, only sars-cov- negative patients (diagnosed via rt-pcr and/or rapid test) should come to the clinic. in places where systematic testing is unavailable, at least, normal temperature and negative epidemiological history should be mandatory to proceed to the outpatient departments. patients with a body temperature higher than . ºc should have additional screening examinations, including routine blood tests, chest computed tomography scanning and even throat swabs for sars-cov- rt-pcr testing. the indication and urgency of the tests for diagnosis should be considered. contraindications for skin, provocation and lung function tests can be explained beforehand to the patient, which helps to accepted article avoid unnecessary in-person consultations. any test generating aerosol particles should be avoided because it is considered high risk (figure ) . personal protective equipment (ppe) must be used when collecting biological samples. biological samples collected on-site from suspected or confirmed covid- patients (e.g. antibody assays, rna isolation, flow cytometry) should be processed following bsl- practices. during and after the covid- pandemic, the usage of bsl- facilities is mandatory for all newly arriving patient samples to prevent spreading the disease. research procedures involving sars-cov- isolation or culture should be conducted in a bsl- facility. , patients with common allergic diseases do not develop distinct symptoms or severe outcomes. allergic children show a mild course similar to non-allergic children . in a recent study of hospitalized children, of them were reported with allergies. allergic rhinitis was the most prevalent allergic disease ( . %), followed by drug allergy, atopic dermatitis, food allergy and asthma. in this study, allergic children showed a reduced increase in acute phase reactants, procalcitonin, d-dimer and aspartate aminotransferase levels compared to all patients. there were no deaths in allergic children in that study. clinical history is very helpful to identify seasonality-and exposure-related symptoms driving the diagnosis of pollen-induced allergic rhinitis. an atopy test (in vivo or in vitro) reinforces the diagnosis. however, covid- can be superimposed on allergic rhinitis symptoms. symptoms such as fever, fatigue and sudden loss of smell, are suggestive of covid- and should be closely monitored. pandemic? this article is protected by copyright. all rights reserved n facial masks have been proven useful in reducing allergen exposure by blocking pollen access to nose and mouth. on the other hand, surgical masks do not protect against inhalation of small airborne contaminants and are not designed to seal tightly against the user´s face, hence the contaminated air can pass through the gaps. there are no conclusive data on the impact of allergic rhinitis on covid- susceptibility . a recent study with allergic rhinitis patients demonstrated a reduction of ace expression in nasal brush samples following an allergen challenge. also, this study reported lower ace -expression in the epithelium of asthmatic patients. on the other hand, tmprss is highly upregulated by type inflammation through the action of il- . therefore, further studies are necessary to determine if allergic rhinitis patients have an altered risk of sars-cov- infection as compared to non-allergic individuals. although limited, the available evidence suggests that, compared to non-allergic individuals, allergic there is no scientific evidence that treatments for allergic rhinitis either increase susceptibility to sars-cov- infection or the severity of covid- . therefore, allergen avoidance measures, nasal saline douches, and background controller therapies recommended by current guidelines for allergic rhinitis, such as nasal corticosteroids or second-generation h -blockers, should be continued as prescribed, both in non-infected and covid- diagnosed patients. the loss of smell in chronic rhinosinusitis is caused by type- inflammation of the olfactory epithelium. in covid- , the exact mechanism of potential olfactory neuropathy is still unclear. however, a study found that sustentacular cells of the olfactory epithelium express ace and tmprss , which enable sars-cov- entry and may subsequently impair the sense of smell. a considerable percentage of covid- patients experience loss of smell as an early sign of the disease. in many patients smell recovers in - weeks and there is no indication that intranasal corticosteroid treatment has a positive impact on the recovery. on the other hand, there is no evidence suggesting that this treatment has a negative impact on symptomatology and/or accepted article development of covid- . consequently, it is recommended to continue regular intranasal corticosteroid treatment for chronic rhinosinusitis (figure ) . an asthma exacerbation is difficult to differentiate from covid- ards or pneumonia by the patient, especially if it is triggered by rhinovirus, or other common respiratory viruses, because both conditions have dry cough and dyspnea. the british thoracic society advises patients with asthma experiencing fever, fatigue and loss of taste or smell to alert their physician as these are indicative of covid- . the distinction can be made by the physician based on the presence of wheeze, which is generally (but not always) absent in covid- pneumonia, as well as high-resolution chest tomography and viral diagnostic tests. patients with controlled asthma are not at higher risk of severe infection than the general population. , ace expression was shown to be decreased in patients with allergic asthma and in those receiving inhaled corticosteroids. on the other hand, ace expression in asthmatic patients was increased in african-americans, in males and associated with diabetes, and type inflammation in children is associated with increased expression of tmprss . it is clear though that uncontrolled asthma is a risk factor of severe covid- , thus all efforts should be focused on treating asthma by regular use of controller medication, including inhaled corticosteroids and biologicals. , , there is no evidence available that patients on inhaled corticosteroids are at higher risk of covid- infection or of more severe symptoms than the general population. it is strongly advised by international scientific societies that patients continue with their routine control medication including inhaled corticosteroids during the pandemic (figure ) . , this article is protected by copyright. all rights reserved recent evidence indicates that inhaled corticosteroid treatment reduces the expression of viral membrane receptors used to infect the human airways in a dose-dependent manner. on the other hand, the immune suppression exerted by corticosteroids may impair anti-viral responses. however, there are no clinical studies investigating the effect of inhaled corticosteroid on sars-cov- infection rates. spirometry is essential for the diagnosis of new asthma cases as stated by the global initiative for asthma guidelines. therefore, it should be conducted, but under special conditions (negative pressure chamber, etc.) and only in areas with low sars-cov- infection incidence. healthcare providers performing lung function testing need to wear maximum ppe (filtering face-piece particles or face mask, goggles, or disposable face shield covering the front and sides of the face, clean gloves, and clean isolation gowns), and the spirometer devices should be properly disinfected between patients (figure ) . an alternative, less precise, is monitoring morning and evening peak expiratory flow variability over a week. , the global initiative for asthma guidelines state that routine spirometry should be avoided, especially in high-risk areas of covid- transmission. if spirometry needs to be performed, maximum ppe should be used (figures and ) . the treatment of asthmatic patients can be monitored using personal devices measuring forced expiratory volume and peak expiratory flow. many of these devices are equipped with remote transmission functions and thus are amenable for the telemedicine management of patients. pandemic? this article is protected by copyright. all rights reserved there is no evidence suggesting that the current approach to treat asthmatic patients during an exacerbation should change during the covid- pandemic. moreover, there is no proof that a short course of systemic corticosteroids impacts the evolution of covid- . thus, oral corticosteroids should be given as usual for the treatment of an asthma exacerbation ( figure ) . , in the few cases in which patients are treated with long-term oral corticosteroids in addition to their high dose inhaled corticosteroids this should be continued in the lowest dose possible to prevent exacerbations. the cause of the asthma exacerbation should be studied thoroughly to rule out potential exacerbations due to viral infections. the preferred treatment is a pressurized metered-dose inhaler with a spacer. each patient should have an individual spacer, and this should not be shared at home. the use of nebulizers should be avoided when possible because they increase the risk of disseminating viral particles, which could affect other patients and healthcare personnel. anti-ige treatment with omalizumab (or other biologics indicated for asthma) should be continued in non-infected patients. self-administration devices at home, whenever this option is available, are preferred, to minimize face-to-face contact in the clinic. in infected patients, omalizumab administration should be delayed until complete clinical recovery and viral clearance is achieved ( figure ) . , endotype is associated with severe asthma in obese patients, are obese asthmatic patients more likely to develop severe covid- ? obesity, as part of the metabolic syndrome, increases the risk of severe covid- . this is due to the pre-existent systemic low-grade inflammation and increased expression of sars-cov- entry receptors (ace , tmprss and cd ). , obese patients tend to have worse asthma control, increased hospitalizations and suboptimal response to standard controller therapy. thus, both difficult-to-control asthma and underlying metabolic syndrome are risk factors for severe covid- . this article is protected by copyright. all rights reserved the il- /th endotype encountered in late-onset obese asthma might be an additional risk factor. , the dermatological manifestations of covid- range from an un-specific macular erythematous rash, urticarial lesions, chickenpox-like vesicles and acro-ischemic lesions. , they can result from local inflammation due to circulating immune complexes or from systemic manifestations leading to vasculitis and thrombosis. these patients are also at increased risk of drug hypersensitivity lesions ( figure ). there is no evidence that patients with barrier defects such as atopic eczema have a higher risk for sars-cov- infection or skin complications during covid- . however, patients with atopic dermatitis are often on systemic immunosuppressants and should be monitored closely. optimal topical treatment regime should also be encouraged in all patients. hand hygiene procedures are pivotal to prevent self-infection and virus spreading. however, extensive water contact enhances dry skin, disturbs the commensal microbiota and leads to barrier disruption in healthy individuals. moreover, it exacerbates diseases with an intrinsic barrier defect such as atopic dermatitis. , effective skin-care after hand hygiene is therefore essential to prevent barrier disruption and sensitization events. here, emollients containing hyaluronic acid, vitamin e, ceramide or urea are recommended. dupilumab is approved for the treatment of moderate-to-severe atopic dermatitis. first data from italy on dupilumab-treated non-infected in high epidemic areas, and current evidence from dupilumab trials, suggest no negative effect of dupilumab regarding viral infections with reports on a reduced number of herpes simplex superinfections and less bacterial superinfections. [ ] [ ] [ ] accepted article this article is protected by copyright. all rights reserved the current eaaci statement on the usage of biologicals in the context of covid- advices no change of therapy in non-infected individuals and to withhold/delay the application of biologicals for a minimum of two weeks or the resolution of the disease in case of sars-cov- infection (figure ) . this is based on expert opinion in the light of missing data and may be adapted if more information becomes available. acro-ischemic lesions on toes and fingers have been identified in a subgroup of covid- patients. , the data available are scarce and it is unclear if preventive or active anticoagulation should be initiated. however, acro-ischemic lesions could predate other sars-cov- symptoms in children and young adults. covid- -induced skin lesions can be related to thrombovascular events (i.e. petechiae, acroischemia, dry gangrene) or to typical viral infections (i.e. erythematous rash, urticaria, maculopapular exanthema). drug hypersensitivity has to be considered as a differential diagnosis, mainly in the second group, being a distinction difficult during the acute phase. diagnosis relies mostly on clinical observations. in that regard, an accurate chronology of the reaction and the drug exposure timeline is very informative . laboratory and histopathological findings may also help. immunomodulatory drugs (including azithromycin), hydroxychloroquine/chloroquine and ifns, are the ones most frequently involved in hypersensitivity reactions. most reactions are non-immediate and further studies are required to clarify whether this increased frequency is caused by the drug immunogenicity or simply derives from a greater consumption as compared to other treatments. this article is protected by copyright. all rights reserved drug provocation tests are not recommended because reactions can occur during the tests, including the generation and spreading of virus-containing aerosols. however, they may be considered after careful risk-benefit assessment in cases of urgent need, such as chemotherapy in cancer patients, perioperative drugs and radiocontrast media in subjects needing urgent procedures, and antibiotics if no effective alternative drug is available. most ait products authorized for use in europe indicate that ait should be discontinued in case of infection; the same principle will apply to the covid- pandemic. patients on subcutaneous or sublingual ait, who are diagnosed with covid- , those suspected of sars-cov- infection or symptomatic patients with a positive contact to sars-cov- individuals, ait should be interrupted until the patient has recovered. in patients not infected or who have recovered from the infection, ait could be continued ( table ) . these recommendations are conditional and could change as clinical data evolve. , ait should continue in non-infected patients or those recovered from covid- ( figure ). this is especially important in patients with life-threatening conditions such as venom allergy. it is possible to extend the intervals between vaccines during subcutaneous ait, as done for inhalant allergens, to minimize visits to the allergy clinic. if venom ait was stopped due to sars-cov- infection, it is unclear when it should be re-initiated because data from convalescent patients is scarce. in patients diagnosed with covid- or cases with suspected sars-cov- infection, oral immunotherapy dosing should continue as indicated in the dosing plan and in coordination with the treating physician. oral immunotherapy can be continued in non-infected patients and those who have recovered from covid- ( figure ). in areas with high level of sars-cov- community transmission, visits to the allergy clinic for oral immunotherapy up-dosing should be postponed. , accepted article this article is protected by copyright. all rights reserved these patients are generally on symptomatic treatment. they need to look out for symptoms suggesting hypoxia or pneumonia, such as shortness of breath, deep shallow breathing, chest pains or persistent tachycardia. special attention needs to be given to those with risk factors for disease progression, such as patients older than years, cardiac or pulmonary comorbidities and immunosuppression. , prophylactic low molecular weight heparin, or heparin, has been recommended by the who in severe to critically ill covid- patients. however, the international society on thrombosis and haemostasis recommended that all hospitalized covid- patients, not just those in icu, should receive prophylactic low molecular weight heparin in the absence of contraindications. during the sars outbreak in , corticosteroids did not change the course of the viral infection and delayed viral clearance. on the other hand, a retrospective study on sars patients in hong kong suggested a better survival rate in patients treated with prednisolone for milder pneumonia or methylprednisolone in more severe cases. recently, chinese experts stated that, in covid- patients, systemic corticosteroids should be considered on individual indications in a low-to-moderate dose and for no longer than a week. the national institutes of health in their covid- treatment guidelines advises against the use of systemic corticosteroids in non-critically ill patients. there are over clinical trials on covid- treatment registered now in the international databases and very few have been completed. currently promoted pharmacological treatments are, at the most, based on anecdotic data collected in small numbers of covid- patients. these studies did not satisfy evidence-based medicine criteria, but caught general attention through news media, for example hydroxychloroquine (see below). tocilizumab is a humanized monoclonal antibody specific for il- r, and it is approved for the treatment of rheumatoid arthritis. a positive response to tocilizumab points towards an imbalanced innate immune response in severe covid- . luo et al. reported that of the patients treated with tocilizumab, of them critically ill, of the patients recovered within a week. prompt resolution of symptoms and encouraging results have also been reported in uncontrolled or retrospective trials. [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] [ ] these zoonotic beta-coronaviruses share structural and genomic similarities that are useful to patients? this article is protected by copyright. all rights reserved fang et al. suggested that there is ace overexpression upon treatment with ace inhibitors, thiazolidinediones and ibuprofen. there were concerns pertaining to the use of nonsteroidal antiinflammatory drugs in covid- patients. the european medicines agency clarified that no scientific evidence established a link between ibuprofen, or other nonsteroidal anti-inflammatory drugs, and a risk to worsen covid- . section : clinical trials and drug discovery in covid- adaptations for clinical trials during the pandemic must include all concerned parties such as there are drugs that interfere with ace and tmprss , which are molecules used by the virus to enter the cell. , for example, camostat mesylate is a clinically proven serine protease inhibitor with affinity for tmprss . it has shown activity against sars-cov- in human lung calu- cells. several drugs that target virus internalization are being investigated, including chloroquine phosphate and hydroxychloroquine, which have shown limited efficacy in humans and raised concerns due to side effects (see below). drugs designed to inhibit the viral replication machinery may be effective against sars-cov- . for example, remdesivir inhibits viral rna polymerases, which prevents sars-cov- replication (see below). it is uncertain whether lopinavir-boosted ritonavir and other antiretrovirals improve clinical outcomes or prophylaxis among patients at high risk of sars-cov- infection. additional potential candidates include other broad-spectrum antiviral drugs such as arbidol and favipiravir and phytochemicals with anti-viral activity such as resveratrol ( figure. ) . in a cohort of severe covid- patients, compassionate-use of remdesivir showed clinical improvement in % of patients ( out of ). of note, a double-blind, randomized, placebocontrolled trial of intravenous remdesivir was conducted in , adults hospitalized with covid- with evidence of lower respiratory tract involvement; remdesivir was superior to placebo in shortening the time to recovery in adults hospitalized with covid- and evidence of lower respiratory tract infection. furthermore, in a study of hiv-positive hospitalized patients with severe covid- , three of them were given lopinavir-boosted ritonavir and darunavir-boosted cobicistat for days. four patients recovered and remained hospitalized. meplazumab is a cd -specific humanized monoclonal antibody that has been shown to prevent sars-cov- infection of fibroblasts (veroe cells). currently, there is insufficient evidence to draw any conclusions on the benefits of meplazumab for the therapy of covid- patients. in an observational chinese study, adults hospitalized with covid- pneumonia (n= ) who were treated with an intravenous infusion of meplazumab as an add-on therapy showed a higher recovery rate compared to controls (n= ). however, these results should be interpreted with caution because they were generated in a non-randomized, non-stratified study, with a small sample size. large-scale studies are needed to assess the effectiveness and safety profile of meplazumab as a potential therapy for covid- . cp therapy for covid- treatment has yielded promising results. for example, in a trial of severe covid- patients, cp therapy was well tolerated and improved the clinical outcomes. the viral load was undetectable after cp transfusion in patients who had viremia. no severe adverse effects were observed. other clinical trials have shown the beneficial effect of cp therapy in covid- patients and ongoing clinical trials will provide additional data on its efficacy, safety and optimal timing for treatment ( table ) . in this regard, it is unclear whether in patients with a high viral load, such as severely ill patients, cp therapy may drive tissue pathology through immune complexes or complement activation. baricitinib, fedratinib, and ruxolitinib are potent and selective jak-stat signaling inhibitors approved for indications such as rheumatoid arthritis and myelofibrosis. these drugs are powerful antiinflammatory medications that may reduce the systemic levels of cytokines associated with covid- . indeed, in a pilot study of covid- patients, baricitinib limited the css and was beneficial for the patients. the use of jak inhibitors has been associated with a higher risk of opportunistic viral infections, such as herpes zoster, which suggests that the reduced inflammation caused by jak inhibitors may limit, to some extent, anti-viral responses. this article is protected by copyright. all rights reserved ivermectin (avermectin b a and avermectin b b) is an anti-parasitic drug that has shown broadspectrum anti-viral activity in vitro. in sars-cov- -infected fibroblasts (vero-hslam cells), a single addition of ivermectin at h post-infection reduced viral rna ~ -fold at hours. however, plasma concentrations of total and unbound ivermectin did not reach the ic determined in vitro, even at a -times higher dose than approved by the food and drug administration (usa). consequently, the likelihood of a successful clinical trial using ivermectin is low. in an observational study of , covid- patients, received hydroxychloroquine treatment, which did not change the risk of intubation or death. furthermore, in a brazilian randomized control study evaluating different doses of chloroquine in covid- patients with severe respiratory symptoms, mortality was . times higher in the high-dose chloroquine arm. moreover, pre-published results from us veterans health administration hospitals did not support any advantages of hydroxychloroquine administered alone or with azithromycin. in addition, the results of a clinical study conducted in individuals showed that hydroxychloroquine did not prevent illness compatible with covid- or confirmed infection when used as postexposure prophylaxis within days after exposure. however, because of the retraction of two main papers on hydroxychloroquine treatment for covid- patients, this area requires further attention by the european medicines agency and the food and drug administration. mesenchymal stem cells may exert antiviral mechanisms in the context of sars-cov- infection. the basal ifn-stimulated gene expression of mesenchymal stem cells is high, which enhances their responsiveness to ifn signaling, potentially inducing broad viral resistance. mesenchymal stem cell therapy may potentiate the low ifn-i and -iii levels and moderate ifn-stimulated gene response reported in sars-cov- -infected ferrets and covid- patients. it is is being used in some centers but its efficacy in covid- has not been proven. data available are mainly experimental with few records in humans and no reports on its efficacy in randomized clinical trials. common anti-hypertensive drugs inhibit ace, but not ace . importantly, ace opposes ace actions and lowers blood pressure by converting angiotensin-ii (a vasoconstrictor peptide) into its metabolites-angiotensin ( - ) (vasodilators). other common related antihypertensive drugs are angiotensin- receptors blockers, which block at- , a receptor for angiotensin-ii, through which it exerts its vasoconstrictor effect. however, at- is not known to be used by sars-cov- to infect cells. it was shown in animal models that ace inhibitors might increase ace expression, thus increasing susceptibility to infection. it has not been proven in humans but it raised the concerns during the covid- pandemic. based on the data available to date, antihypertensive treatment with these medications should be continued. at the moment, the animal model that resembles more closely human covid- is the rhesus macaque, whose ace receptor is identical to that in humans. this model recently showed that sars-cov- reinfection was hampered due to infection-acquired immunity and demonstrated the therapeutic effect of remdesivir in covid- prior use in human clinical trials. , the murine ace receptor is different from humans, hence humanized murine models with recombinant human ace are necessary. previous vaccine research for sars/mers facilitates rapid translation. in the who vaccine single-domain antibodies have been investigated as potential therapeutics for influenza, rsv and hiv in addition to coronaviruses. sars-cov- mainly targets the respiratory tract, hence the development of vaccines directed to the respiratory epithelia and lung parenchyma using a nebulizer has been considered to maximize bioavailability and function. although active research against respiratory viruses has focused on aerosolized plasmid dna vaccines, other forms of vaccine administration are currently further advanced in clinical trials. veterinary medicine commonly uses aerosolized coronavirus vaccines for chicken farms. a novel vaccine platform requires careful evaluation and should ideally include toxicological studies in valid animal models. early progress towards sars vaccines has facilitated a "running start" but standards of care and safety must be maintained. acceleration rather than omission of clinical trials is key. preliminary data from oxford university is anticipated by mid- . of note, a doseescalation, single-center, open-label, non-randomized, phase was conducted in healthy individuals that received an ad vectored covid- vaccine. the vaccine was tolerable and this article is protected by copyright. all rights reserved immunogenic at days post-vaccination. sars-cov- -specific antibodies peaked at day postvaccination and specific t-cell responses were detected from day post-vaccination. an important aspect is that covid- -associated mortality is very high, almost unavoidable when the pandemic control fails. this is due to rapid community spread, high community virus, especially in the elderly and co-morbid, but also in younger non-comorbid persons, including healthcare workers, young adults and children. the covid- pandemic also seems to be characterized by a significant level of asymptomatic spread. [ ] [ ] [ ] the iceberg of covid- : are there asymptomatic cases below the surface? the the differences are almost entirely due to the timing and effectiveness of public health interventions. countries that failed to control did too little, too late, and allowed sars-cov- to rip through their population, with catastrophic outcomes. those that intervened early effectively stopped the disease transmission. this article is protected by copyright. all rights reserved it is difficult to determine as it varies greatly from country to country, depending on how well countries control their epidemics with widespread testing, case isolation and vigorous contact tracing, testing and isolation if positive. in countries that do this well, the r can be very low indeed. in countries that fail to control the spread of the virus, the r is high but unknown as sars-cov- spreads untested and therefore undetected. it has been estimated to be ~ . . sars-cov- transmits more readily than either sars-cov or mers-cov. the r of sars-cov- is controversial but if left unchecked it is likely to be greater than - . however, the r number cannot be precisely defined as no country has left it to spread completely unchecked. in any case, even when preventative measures are taken, the r of sars-cov- is higher than that of sars-cov ( . - . ) and mers-cov (< ). there is a considerable frequency of very mild covid- patients as well as asymptomatic sars-cov- -infected people. this makes transmission control more challenging than either sars-cov or mers-cov, where illness is frequently more severe. children are at low risk of severe covid- outcomes. , most patients in pediatric age with sars-cov infection presented with no or mild clinical manifestations, including fever, fatigue and dry cough. they were typically managed with supportive treatments only and they had generally a favorable prognosis with a recovery within weeks. [ ] [ ] [ ] young children also frequently carry other respiratory viruses, which potentially limit sars-cov- infection, as reported for other viral infections. differences between children and adults in the regulation of ace expression may also play a role. ace mrna expression was high in type i and ii alveolar epithelial cells, in nasal and oral mucosa and nasopharynx, in smooth muscle cells and endothelium of vessels from the stomach, small intestine, colon, and in the kidney of human adults (mean age ± ). interestingly, a recent study demonstrated age-dependent ace gene expression in the nasal epithelium, which was lowest in younger children and increased with age. in addition, cd , cd and their molecular interaction proteins seem to be differently expressed in peripheral blood mononuclear cells and t cells in children in comparison with adults. many children remain asymptomatic, even when they have radiologic pneumonia detected on screening. given that children are effective transmitters of other respiratory viruses, it is expected that they will be just as good at transmitting sars-cov- . bats are likely the natural reservoir of sars- severity (see questions below). data on ethnicity and covid- are scarce and further research on ethnicity and covid- outcomes is needed. however, the data available show a disproportionate number of covid- deaths in black, asian and minority ethnic backgrounds. in fact, one third of uk icu admissions are reportedly from them. in the usa, african americans had more covid- diagnoses and deaths, after adjusting for age, poverty, comorbidities, and epidemic duration. these disparities are also seen in the hispanic and asian communities. pregnant women may be at a higher risk of poorer covid- outcomes because they have deficient ifn-α and ifn-λ responses to viral infections. however, reported pregnancy outcomes in covid- are reassuring as they appear similar to non-pregnant adult females. this article is protected by copyright. all rights reserved testing treatments is problematic because pregnant women are excluded from most trials. it is known that azithromycin doubles innate ifn production from virus-infected lung cells. it is safe for all trimesters of pregnancy and has been shown effective in high-quality clinical trials of virusinduced lung disease. , given that the human ace protein is encoded on the x chromosome, this may be relevant for malefemale differences in outcomes. particularly in males with rare ace coding variants as they will express those variants in all ace -expressing cells compared to a mosaic pattern of expression in females. males may also have differences in certain innate antiviral responses compared to female counterparts. there is reasonably robust data of covid- deaths in hospitals because most people who die in hospital are tested. deaths outside hospitals are likely underestimated as people are dying in care homes where mortality approaches ~ %, and may die without being tested and diagnosed. it is difficult to determine prevalence as testing practices vary so much from country to country. seroprevalence studies will help to collect these data. covid- was introduced rapidly to many industrialized countries as a result of air travel. most of europe and the usa probably did not react in a timely and efficient manner, resulting in the rapid spread and subsequent high mortality rates. in light of the devastating situation in many european countries and the usa, less industrialized countries had a little more time to better prepare to control the pandemic. an important factor for prevalence studies is the percentage of the population that has undergone a diagnostic test, which seems to be at lower levels in developing countries. this article is protected by copyright. all rights reserved respiratory viruses spread less readily in summer than in winter for reasons that are not well understood. dry air and higher temperatures are slowing down the spread of respiratory viruses. absence of school attendance, more time outdoors, greater household ventilation, warmer temperatures facilitating virus inactivation and higher vitamin d levels are all likely to play a part. although social distancing measures are implemented, the summer weather should play a role in hampering the spread of covid- . however, based on the analogy of previous influenza pandemic, it is unlikely that summer, on its own, could stop transmission of sars-cov- . [ ] [ ] [ ] it largely depends on the sars-cov- seroprevalence developed in each country, which is still unknown. countries that have had widespread transmission may be hit by a second wave, but presumably with less severe consequences. countries that effectively controlled the pandemic are at a higher risk of second wave of covid- if those effective controls are relaxed due to the limited viral transmission and lack of active immunization. sars-cov- has spread worldwide in humans, causing mild or no disease in many cases. it will continue circulating similar to other human coronaviruses ( e, hku , nl , oc ), and it may well become an endemic, seasonal virus. the main route of sars-cov- transmission is via respiratory droplets and aerosols. [ ] [ ] [ ] avoidance of high virus loads, acquired through aerosol and droplet transmission, is paramount to prevent severe outcomes. consequently, social distancing, masks and hand sanitation are undoubtedly effective because they prevent the droplet and surface contact-associated initial high virus load and the increased risk of severe disease. [ ] [ ] [ ] what is the evidence supporting social distancing and face mask to prevent sars-cov- infection? this article is protected by copyright. all rights reserved a systematic review and meta-analysis has found that transmission of viruses was lower with physical distancing of m or more, compared with a distance of less than (or . ) and protection was increased as distance was lengthened. in addition, face mask use could result in a large reduction in risk of infection (or . ), with stronger associations with n or similar respirators compared with disposable surgical masks or similar. eye protection also was associated with less infection (or . ). therefore, the covid- pandemic can be controlled if social distancing is combined with widespread testing, case isolation, vigorous contact tracing and personal protection. indeed, severe and critical illness among chinese healthcare workers before january th was %, a time when personal protection equipment and infectious control measures were likely not implemented. after february st , when personal protection measures were in place, the percentage of severe and critically ill chinese healthcare workers dropped to . %. sars-cov- remained viable in aerosols for h with a ~ -fold reduction in infectious titre. sars-cov- was more stable on plastic and stainless steel than on copper and cardboard; viable virus was detected up to days after application to plastic and days to stainless steel, on each surface the virus titer was reduced nearly ~ -fold. importantly, sunlight exposure inactivated % of infectious sars-cov- every . minutes in simulated saliva and every . minutes in culture media. this study suggests that persistence, and subsequently exposure risk, may vary significantly between indoor and outdoor environments. therefore, it is convenient to minimize contact with surfaces touched by others (even before sars-cov- existed), particularly at indoor environments, for example when using public transportation. in covİd- patients, the estimated median time from symptom onset to viral clearance in the nasal swabs was days, while in asymptomatic cases it was days. in patients that recovered, the median duration of viral shedding was ⁓ days, while in non-survivors it was detected until death. the longest duration of viral shedding in survivors was days. accepted article and found that a slow viral clearance is associated with an increased risk of high disease severity with a % mortality rate. the individual variation in the transmission of an infection is described by a factor called "dispersion factor or k". the lower "k" value, the more transmission comes from a small proportion of individuals acting like superspreaders. superspreading clusters have been observed in past coronavirus outbreaks (sars/mers), where a small number of infected individuals was responsible for a large proportion of secondary transmissions, with an estimated "k" of about . for sars and . for mers. it is unclear whether superspreading clusters have contributed to the covid- outbreak. a simulation of early outbreak trajectories estimated that "k" for covid- is higher than for sars and mers. however, in a recent preprint study, the estimate of "k" for sars-cov- was around . , suggesting that around % of infected patients may have been responsible for % of secondary transmissions. individual variation in infectiousness is difficult to measure, as it is mostly empirical, but the identification of any sars-cov- superspreading will be of primary importance for pandemic control. the designation of covid- -dedicated wards and personnel within hospitals is useful to limit nosocomial sars-cov- infections. it also allows other non-covid- conditions to be treated using routine healthcare resources more safely and effectively. maintaining such separation requires intensive sars-cov- testing in view of the high asymptomatic infection rate. community-based strategies are effective at controlling the transmission of sars-cov- . australia, hong kong, japan, singapore, south korea, and new zealand have all controlled effectively. their cumulative covid- mortality is > -fold less than that in belgium, france, italy, spain and the uk, countries which have had difficulties to adequately control the pandemic. , , it is important to implement measures to contain the spread of the virus, such as developing models to predict sars-cov- -related mortality. closing live animal markets is likely to reduce the risk of future viral outbreaks although this is not a practical way to prevent viral outbreaks for multiple reasons including social and economic. lifestyle factors that may influence sars-cov- infection susceptibility and covid- severity include smoking, stress, diet and alcohol intake, among others. for example, smoking has been shown to increase the susceptibility to respiratory tract infections and its severity, and it is a risk factor for severe covid- . moreover, alcohol consumption may impair anti-viral immunity; in vitro studies with human monocytes have shown that both acute and prolonged alcohol exposures inhibit type i ifn induction upon toll-like receptor- and - stimulation . dietary habits may also play a role as obese patients have been shown to have a higher risk of developing severe covid- . furthermore, there are bioactive food compounds with antiviral activity, such as resveratrol, although the amount of them obtained through the diet is unlikely to play a relevant role in covid- it is known that respiratory virus infection causes perturbations in the gut microbiota and that germfree mice are more susceptible to viral infections, which intimates a role for the microbiota in covid- . however, the impact of the commensal microbiota on sars-cov- infection susceptibility and covid- severity is unknown. an essential step is identifying the bacterial species interacting with sars-cov- . this is rather challenging given the large number of bacterial species in the lung and respiratory tract, and especially in the gut. however, a number of lung metagenomic studies have reported an abundance of prevotella in the lung of sars-cov- infected patients . while in accepted article silico analysis have revealed that prevotella proteins may promote viral infection , prospective studies are necessary to ascertain if this is a consequence of the infection or a risk factor for it. it is well-established that epithelial barrier defects and/or damage favor the development of th immunity. , increased hygiene, in general, as well as overexposure to epithelial barrier opening molecules, such as detergents, can promote the onset of allergic disease. to date, there is no evidence linking the covid- protective measures (gloves, hand-sanitizers, etc.) with increased allergy prevalence. in this regard, multifactorial epidemiological studies are needed. these studies should consider the impact on allergic diseases of virus-specific type responses and psychosocial and environmental changes caused by the pandemic and efforts to contain it. although there has been a significant change in pollution parameters, unfortunately this reduction in pollution is transient and consequently unlikely to be significant. the exposome-related allergy and asthma risk is multifactorial. it includes climate change, biodiversity, the microbiome and nutrition among others, which have not changed during the pandemic. in addition, although pollution levels have dropped, climate change still occurs at an accelerated pace. lifestyle changes during the lockdown , weight gain and increased exposure to indoor allergens and pollutants may even increase the incidence of allergic diseases in the long-run. with the rapid spread of covid- at a pandemic scale, we are overwhelmed and drowned with a wealth of information. a global fight to contain the pandemic has started in which we need international solidarity and prompt sharing of accurate scientific information. we strongly 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with convalescent plasma treatment with convalescent plasma for critically ill patients with severe acute respiratory syndrome coronavirus infection use of convalescent plasma therapy in two covid- patients with acute respiratory distress syndrome in korea treatment with convalescent plasma for covid- patients in wuhan effect of convalescent plasma therapy on viral shedding and survival in covid- patients treatment of covid- patients with convalescent plasma the authors thank the european academy of allergy and clinical immunology this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved this article is protected by copyright. all rights reserved key: cord- - s wtj authors: ruscitti, piero; berardicurti, onorina; di benedetto, paola; cipriani, paola; iagnocco, annamaria; shoenfeld, yehuda; giacomelli, roberto title: severe covid- , another piece in the puzzle of the hyperferritinemic syndrome. an immunomodulatory perspective to alleviate the storm date: - - journal: front immunol doi: . /fimmu. . sha: doc_id: cord_uid: s wtj the coronavirus disease (covid- ), an acute respiratory disease caused by severe acute respiratory syndrome-coronavirus- (sars-cov- ), has been declared as a worldwide public health emergency. interestingly, severe covid- is characterized by fever, hyperferritinemia, and a hyper-inflammatory process with a massive release of pro-inflammatory cytokines, which may be responsible for the high rate of mortality. these findings may advocate for a similarity between severe covid- and some challenging rheumatic diseases, such as adult onset still's disease, secondary hemophagocytic lymphohistiocytosis, and catastrophic anti-phospholipid syndrome, which have been included in the “hyperferritinemic syndrome” category. furthermore, as performed in these hyper-inflammatory states, severe covid- may benefit from immunomodulatory therapies. the coronavirus disease (covid- ) is an acute respiratory disease caused by a novel coronavirus (severe acute respiratory syndrome-coronavirus- , sars-cov- ), identified in wuhan, china in december ( ) . since then, the covid- outbreak has spread worldwide, becoming a pandemic, causing a public health emergency, according to the world health organization (who), and resulting in thousands of deaths ( ) . sars-cov- is a β-coronavirus, an enveloped non-segmented positive-sense rna virus, which could be transmitted from bats via unknown intermediate hosts to infect humans, using the angiotensin-converting enzyme (ace ) receptor ( ) . the latter, is more expressed in adults than children, and thus possibly explains why the disease is more aggressive in older patients ( ) . covid- shows a heterogeneous course, from patients affected by mild flu-like symptoms to patients with unremitting fever and severe respiratory involvement. on this basis, markers of poor prognosis have recently been investigated to effectively prioritize resources to patients with more severe symptoms ( ) . interestingly, this study identified hyperferritinemia and interleukin (il)- , as predictors of poor outcome, thus suggesting a hyper-inflammatory process as the major cause of death ( , ) . in severe covid- , a specific cytokine profile resembling the pattern of a secondary hemophagocytic lymphohistiocytosis (hlh) has been shown, due to significant increases of il- , il- , granulocyte colony stimulating factor (gcsf), interferon-γ inducible protein (ip- ), monocyte chemoattractant protein (mcp- ), macrophage inflammatory protein -α, and tumor necrosis factor (tnf) ( ) . contextualizing unremitting fever, hyperferritinemia, and the hyper-inflammatory process, severe covid- shows similarity to disorders comprised in the so-called hyperferritinemic syndrome ( ) . this syndrome includes adult onset still's disease (aosd), systemic juvenile idiopathic arthritis (sjia), secondary hlh, catastrophic anti-phospholipid syndrome (caps), and septic shock ( ) . hyperferritinemia is a common trait of all these forms, which could be an active pathogenic mediator and not only a consequence of the inflammation ( ). on these bases, we aimed to review the similarities between severe covid- and diseases included in hyperferritinemic syndrome, from a pathogenic, clinical, and therapeutic point of view, thus proposing new insights to improve the management of those patients. coronavirus rnas may act as pathogen-associated molecular patterns, which are detected by the pattern recognition receptors and activate downstream cascades pro-inflammatory pathways ( , ) . in the endosome, toll-like receptor (tlr) , tlr , tlr , and tlr may sense viral rna and dna ( ) , whereas, in the cytoplasm, the viral rna receptor retinoic-acid inducible gene i ( ), cytosolic receptor melanoma differentiation-associated gene , and nucleotidyltransferase cyclic guanosine monophosphateadenosine monophosphate (gmp-amp) synthase may recognize viral rna and dna ( ) . consequently, downstream cascades molecules are triggered, involving adaptor molecule myeloid differentiation primary response (myd ), transcription factor nuclear factor-κb (nf-κb), and interferon regulatory factor , leading to the production of pro-inflammatory molecules ( , ) . in fact, plasma cytokines and chemokines were increased in covid- patients, including il- β, il- , il- , il- , il- , il- , il- , il- , gcsf, ip- , interferonγ (ifn-γ), and tnf ( ) . during severe covid- , these mechanisms could be exaggerated, probably because of a specific genetic susceptibility ( ) , and these patients are characterized by very high blood levels of pro-inflammatory mediators and ferritin ( , ) . the latter is an iron-binding molecule, which is produced after pro-inflammatory stimuli, in addition to iron availability ( ) . furthermore, ferritin comprises subunits, codified according to their molecular weight into heavy (feh) and light (fel) subunits. remarkably, increased expression of feh and of cd +/feh+ macrophages may be observed in inflammatory infiltrate of aosd and secondary hlh ( , ) . additionally, a stimulatory effect of feh on nf-kb has been described, acting as a pro-inflammatory cytokine on hepatic stellate cells ( ) . in this work, ferritin was shown to regulate an iron-independent signaling pathway that resulted ultimately in nf-kb activation ( ) , thus converging on the same pathway elicited by sars-cov- rnas ( , ) . on the contrary, the deletion of feh reduced the inflammatory burden in the model of sepsis by lipopolysaccharide-induced endotoxemia, cecal ligation, and puncture ( ) . such protection was predominantly mediated by the compensatory increase in fel, associated with an inhibitory action on nf-kb ( ) . the proinflammatory cytokines, which are elevated in hyperferritinemic syndrome, have also been described in severe covid- ( ) , and may preferentially induce the expression of feh, via fer , a regulatory element acting as a binding site to nf-kb. the latter, in turn, stimulates the synthesis of further feh and pro-inflammatory cytokines, thus perpetuating a vicious inflammatory loop ( ) . in addition, feh+/il- + macrophages have been shown in the infiltrate of aosd and secondary hlh ( , ) , which may further release feh, following inflammatory stimuli ( ) , and thus contributing to the inflammatory loop ( ) . on these bases, we hypothesize that severe covid- shares common pathogenic mechanisms with other diseases of hyperferritinemic syndrome ( ), with ferritin enhancing the inflammatory burden and triggering a vicious pathogenic loop. lung involvement and hyper-inflammation are at the crossroad between severe covid- and the hyperferritinemic syndrome. as observed in other β-coronaviruses diseases, covid- is characterized by fever, dry cough, increasing dyspnoea with hypoxemia, and bilateral ground-glass opacities and patchy shadowing with a peripheral or posterior distribution, mainly in the lower lobes, on chest ct scans ( ) . in fact, an anatomy report of a covid- pneumonia cadaver showed that sars-cov- invades the respiratory mucosa and infects other cells, thus provoking an inflammatory response in the lower airway and causes lung injury ( ) . considering that coronavirus binds to the host cells using the ace receptor, which is highly represented in the lower respiratory tract, a persistent and repeated stimulation of tlrs in the lung may occur, hence triggering an aberrant immune response and the production of a cytokine storm ( ) . the latter is the result of overwhelming systemic inflammation with a massive release of pro-inflammatory cytokines, quickly progressing to multiple organ dysfunction syndrome and eventually to death ( ) . in spite of various inflammatory etiologies, cytokine release syndrome is supported by an essential underlying hypothesis: the massive release of cytokines as a consequence of: (i) excessive and repeated inflammatory stimuli, and (ii) an inadequate regulation of inflammation, (iii) an uncontrolled release of cytoplasmic cytokines from destroyed lymphocytes after anti-cancer therapies ( , ) . in addition, it has been shown that increased amounts of pro-inflammatory cytokines, including il- β, il- , il- , ifn-γ, ip- , and mcp , were associated with pulmonary inflammation and extensive lung damage in sars patients ( ) , thus suggesting a further pathogenic loop in inducing the cytokine storm. although the mechanisms of how covid- and, in other more general viral infections, would prompt the cytokine storm syndrome are not fully elucidated, it has been suggested that the ifn-γ, which is largely released by a variety of hematopoietic cells in response to viral infection, may facilitate the occurrence of hyperinflammation ( ) . in patients with sjia, lung involvement may trigger systemic inflammation and the development of secondary hlh and ifn-γ plays a central pathogenic role ( , ) . in fact, in lung biopsies in patients with sija, the analysis of expressed genes revealed that many of the up-regulated targets were in gene pathways related to an ifn-γ signature, including human leukocyte antigen (hla)-d family members and other ifnrelated genes ( , ) . two of the most highly up-regulated non-hla genes were chemokine (c-x-c motif) cxcl , and cxcl ( ) , which are ifn-induced chemokines strongly correlated with the occurrence of secondary hlh ( ) . in addition, the lung is one of the major physiological producers of il- β and il- ( ), which are also involved in pathogenic steps, leading to the occurrence of secondary hlh ( , ) . considering all of these findings, it is possible to postulate that during the acute respiratory distress syndrome of covid- , the sars-cov- may trigger a hyper-inflammatory reaction strongly resembling that observed in the lung involvement of sjia, in which the lung acts as a trigger to amplify the immune response. the final result is the uncontrolled proliferation of activated immune cells, the massive production of pro-inflammatory mediators, and the development of cytokine storm syndrome, either in severe covid- or sjia. from a clinical point of view, severe covid- and the diseases included in hyperferritinemic syndrome share a fever as the main clinical symptom. in these conditions, the analysis of fever pattern would also suggest a useful clue to assess the severity of the disease and the occurrence of complications. in sjia and aosd, a typical change from the high-spiking intermittent typical quotidian pattern, to a continuous unremitting pattern suggests the occurrence of secondary hlh, and the worsening of the clinical situation toward a life-threatening hyperinflammatory complication ( ) . during covid- , on the basis of observations from clinicians on the frontlines, the occurrence of unremitting fever would similarly identify a more aggressive subset of patients, at higher risk of a poor prognosis. in addition, in severe covid- , hyperferritinemia is observed, suggesting a marker of severity ( , ) . although it has poor specificity, a -fould increase of ferritin is strongly suggestive of the diseases included in hyperferritinemic syndrome, and is a useful marker to assess disease activity and to predict a poor prognosis ( ) . in fact, hyperferritinemia is associated with increased mortality in sepsis, multiple organ dysfunction syndrome, and critical illness ( ) ( ) ( ) . thus, the clinical phenotype, characterized by unremitting fever and hyperferritinemia, identifies the most severe subset of covid- as observed in the diseases included in hyperferritinemic syndrome. considering the lack of efficacy of antiviral therapy for severe coronavirus infection, it is reasonable to postulate the clinical usefulness of specific immunomodulatory therapies (figure ) , as observed for other diseases included in hyperferritinemic syndrome such as intravenous immunoglobulins (ivigs) and tocilizumab, the humanized monoclonal antibody against il- receptor ( ). ex juvantibus, one of the best criteria for identifying a common pathogenic mechanism, among different diseases, is that the clinical manifestations were reversed upon initiation of the same therapy. it has been shown that, after ivigs therapy, a significant reduction of hyperferritinemia, both in sepsis and secondary hlh was observed, correlating with an improvement in patients ( ). considering their proposed anti-viral activity, possibly comprising many cross-reacting antiviral antibodies and per se immunomodulatory activities ( ), ivigs has also been proposed to treat severe covid- ( ) . another therapeutic immunomodulatory possibility in severe covid- is the administration of hydroxychloroquine (hcq). this drug, has been a licensed treatment for rheumatoid arthritis for many years, and was shown to reduce the viral load, favoring the disappearance of sars-cov- ( ) . however, although it seems promising, a recent meta-analysis, including , patients, suggested that more data are required for a definitive conclusion on the use of hcq in this setting, since no difference was observed in virologic cure, death, or clinical worsening of disease between hcq-treated patients and control groups ( ) . as far as tocilizumab is concerned, the rationale for its use in severe covid- derived from evidence of its beneficial effect on cytokine release syndrome. this is a clinically significant, on-target, off-tumor side effect of the chimeric antigen receptor t-cell therapies administered for treatment of malignancies ( ) . characteristics of cytokinerelease syndrome include fever, encephalopathy, hypotension, and coagulopathy, leading to multiorgan failure, associated with very pronounced levels of hyperferritinemia and il- ( ). the latter provided an effective therapeutic target in cytokine release syndrome ( ) . mirroring this finding, tocilizumab has been used to treat severe covid- with promising results, as observed in other diseases of hyperferritinemic syndrome ( ) . furthermore, a reduction of ferritin, obtained by combing immunomodulatory drugs, was associated with a lower mortality rate in caps and hlh ( ), thus possibly suggesting the use of, in a more aggressive subset of covid- , a combination therapy with both antiviral and antiinflammatory drugs, at the same time ( ) . in addition, the repurposing of these drugs in severe covid- could benefit from the findings of previous reports, and thus, on this basis, many clinical trials are ongoing in different countries (chictr , nct , nct , and nct ). as far as other immunomodulatory strategies in covid- are concerned, il- inhibition showed benefits in sepsis, in which both hyperferritinemia and hyper-inflammation, may be observed, contributing to the dysregulation of the host immune system ( ) . a post-hoc analysis of data from a phase randomized controlled trial showed some improvement of patients with sepsis, following anakinra, a recombinant non-glycosylated form of human il- receptor antagonist, thus suggesting its possible use in those patients ( ) . as a consequence, it is possible to hypothesize that anakinra may also relieve severe covid- . reported data suggest the possible efficacy of emapalumab, a monoclonal antibody neutralizing ifn-y, approved in the treatment of hlh and its massive production of pro-inflammatory cytokines ( ) . due to the important role of ifn-y in driving hyper-inflammation during viral infections, emapalumab may be an additional immunomodulatory therapy that could be employed in the treatment of severe covid- . in addition, available literature suggests that janus kinase (jak) inhibition might affect covid- twice as much, by targeting both inflammation and cellular viral entry ( ) . it has been proposed that baricitinib, a jak /jak inhibitor, may control the hyper-inflammatory steps in those diseases, characterized by a cytokine storm, since a plethora of cytokine receptors indiscriminately use these jaks as mediators of ligands binding and consequent activation of the inflammatory cascade ( ) . furthermore, the disruption of p -associated protein kinase , a known regulator of viral endocytosis into the cell, by baricitinib, could possibly be an additional positive effect in covid- , decreasing the viral entry ( ) . finally, considering ferritin as a pathogenic mediator, this could also be proposed as a therapeutic target in these conditions. high-volume hemofiltration and plasma exchange, extracorporeal blood purification techniques, have been employed to treat secondary hlh to sepsis ( ) ( ) ( ) . interestingly, in parallel with the clinical efficacy, these procedures induce a ferritin reduction ( ) ( ) ( ) , suggesting that the mechanical removal of ferritin could have a possible therapeutic role. in this work, we discuss the similarities, from a pathogenic, clinical, and therapeutic point of view, between severe covid- and four conditions; secondary hlh, aosd, caps and septic shock, which are included in hyperferritinemic syndrome. all these diseases are characterized by very high levels of ferritin, which could not only be the product of the inflammation but rather may play a pathogenic role. possibly, in an inflammatory environment, as observed in these diseases, hyperferritinemia may be involved in a vicious pathogenic loop prompting its pro-inflammatory properties. in severe covid- , ferritin could be a further possible enhancer of the cytokine storm. clinically, unremitting fever is a common feature of severe covid- , suggesting that a change from the intermittent quotidian pattern to a continuous unremitting form would indicate a worsening toward the cytokine storm, as in aosd and sjia. the hyperferritinemia seems to be a marker of poor prognosis and response to treatment, in both severe covid- and hyperferritinemic syndrome. finally, the good response to immunomodulatory therapies, observed during severe covid- , strongly supports the link between this form and other diseases included in hyperferritinemic syndrome. in addition, targeting the hyperinflammatory process, through immunomodulatory therapies, decreases the high mortality rate of all these diseases ( , ( ) ( ) ( ) , thus proposing additional therapeutic options to improve the survival of severe covid- patients, the latter characterized by an over-exuberant pro-inflammatory response, in which the viral load is not correlated with the worsening of symptoms ( ) . in conclusion, we hypothesize that severe covid- shares pathogenic mechanisms, a clinical picture, outcomes, and therapeutic strategies with disorders included in hyperferritinemic syndrome. the hyperferritinemia, characterizing all these diseases may be a pathogenic mediator, enhancing the inflammatory burden, and, as observed in aosd, caps, and secondary hlh, its reduction is associated with a lower mortality. thus, at present, severe covid- , seems to be a new entity in hyperferritinemic syndrome. in addition, since accumulating evidence suggests that severe covid- is associated with a cytokine storm syndrome, therapeutic strategies combining immunomodulatory therapies, may improve the management of those patients. furthermore, in this setting, high levels of ferritin, identifying a more aggressive subset of covid- , may drive clinicians to apply more aggressive therapies and resources in those patients, thus balancing appropriate escalation of therapy and minimizing the exposure to iatrogenic harm. sars-cov- and consequent covid- are a new and great challenge for health systems worldwide, requiring a multidisciplinary approach and a large body of knowledge. all the authors meet all criteria for authorship in the icmje recommendations, since all authors made substantial contributions to the conception or design of the work, the acquisition and interpretation of data. all authors contributed to the critical review and revision of the manuscript and approved the final version. all the authors agreed to be accountable for all aspects of the work. the epidemiology and pathogenesis of coronavirus disease (covid- ) outbreak a pneumonia outbreak associated with a new coronavirus of probable bat origin are children less susceptible to covid- ? clinical predictors of mortality due to covid- based on an analysis of data of patients from wuhan, china covid- : consider cytokine storm syndromes and immunosuppression clinical features of patients infected with novel coronavirus in wuhan the hyperferritinemic syndrome: macrophage activation syndrome, still's disease, septic shock and catastrophic 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syndrome in patients affected by adult-onset still disease: analysis of survival rates and predictive factors in the gruppo italiano di ricerca in reumatologia clinica e sperimentale cohort key: cord- -vof qat authors: jain, vageesh; yuan, jin-min title: systematic review and meta-analysis of predictive symptoms and comorbidities for severe covid- infection date: - - journal: nan doi: . / . . . sha: doc_id: cord_uid: vof qat background/introduction covid− , a novel coronavirus outbreak starting in china, is now a rapidly developing public health emergency of international concern. the clinical spectrum of covid− disease is varied, and identifying factors associated with severe disease has been described as an urgent research priority. it has been noted that elderly patients with pre-existing comorbidities are more vulnerable to more severe disease. however, the specific symptoms and comorbidities that most strongly predict disease severity are unclear. we performed a systematic review and meta-analysis to identify the symptoms and comorbidities predictive of covid− severity. method this study was prospectively registered on prospero. a literature search was performed in three databases (medline, embase and global health) for studies indexed up to th march . two reviewers independently screened the literature and both also completed data extraction. quality appraisal of studies was performed using the strobe checklist. random effects meta-analysis was performed for selected symptoms and comorbidities to identify those most associated with severe covid− infection or icu admission. results of the studies identified, were selected after title and abstract analysis, and studies (including covid− patients) were chosen for inclusion. the icu group were older ( . years) compared to the non-icu group ( years), with a significantly higher proportion of males ( . % vs. . %, p= . ). dyspnoea was the only significant symptom predictive for both severe disease (por . , % ci . − . ) and icu admission (por . , % ci . − . ). notwithstanding the low prevalence of copd in severe disease and icu-admitted groups ( . % and . %, respectively), copd was the most strongly predictive comorbidity for both severe disease (por . , % ci . − . ) and icu admission (por . , % ci . − . ). cardiovascular disease and hypertension were also strongly predictive for both severe disease and icu admission. those with cvd and hypertension were . ( % ci . − . ) and . ( % ci . − . ) times more likely to have an icu admission respectively, compared to patients without the comorbidity. conclusions dyspnoea was the only symptom strongly predictive for both severe disease and icu admission, and could be useful in guiding clinical management decisions early in the course of illness. when looking at icu-admitted patients, who represent the more severe end of the spectrum of clinical severity, copd patients are particularly vulnerable, and those with cardiovascular disease and hypertension are also at a high-risk of severe illness. to aid clinical assessment, risk stratification, efficient resource allocation, and targeted public health interventions, future research must aim to further define those at high-risk of severe illness with covid− . the ongoing novel coronavirus outbreak is a public health emergency of international concern (pheic), involving a novel type of coronavirus originally identified in wuhan, china. at the time of writing, there have been , confirmed cases around the world with , deaths ( ) . defining the spectrum of clinical manifestations and the risk factors for severe covid- infections has been identified as an urgent research priority ( , ) . as the virus spreads globally it is likely that government strategies will shift from containment and delay towards mitigation ( ) . this will involve rapidly scaling up healthcare resources including staff, equipment, facilities, and training, to effectively identify and treat patients. to maximise the use of these limited resources it will be imperative that clinicians are able to triage covid- patients likely to recover after a mild illness from those who are not. in order to do this, a better understanding of the symptoms and comorbidities (which are the first and most routinely collected components of patient data) related to covid- severity is required. this can improve patient outcomes through three chief mechanisms: early clinical intervention in high-risk patients, designing appropriate clinical pathways and risk prediction tools, and the efficient allocation of scarce resources and expensive treatments. further still, the early identification of individuals more likely to deteriorate can help direct appropriate public health actions to protect the vulnerable and prevent further spread of infection. a recent meta-analysis of symptoms in , covid- patients from studies found that fever and cough were the most common symptoms, with . % and . % experiencing these, respectively ( ) . it also found that the case fatality rate (cfr) was . %, but the association between individual patient factors and severe infection was not investigated. most reported cases have occurred in adults (median age years) ( ) . according to most recent us centers for disease control and prevention (cdc) guidance, risk factors for severe illness are not yet clear, although older patients and those with chronic all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in the copyright holder for this this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint medical conditions may be at higher risk. ( ) the primary aim of this study is therefore to conduct a systematic review and meta-analysis, aggregating all currently available data from published studies, of symptoms and comorbidities predictive for severe illness with covid- . this study was prospectively registered on prospero. identification of relevant existing literature was performed by an online search in three databases: medline, embase and global health, for studies published from st january to th march . the mesh headings (keywords) searched were 'ncov*' or 'coronavirus' or 'sars- -cov' or 'covid*' and 'symptom*' or 'clinical' or 'predict*' or 'characteristic*' or 'co-morbidit*' or 'comorbidit*' or 'condition*'. two reviewers (vg, jmy) independently screened the list of titles and abstracts, and the full text of chosen manuscripts. disagreements on which manuscripts to include during both title and abstract screen, and the subsequent full-text analysis, were discussed until a conclusion was reached. in addition to the medline/embase/global health search, citation tracking was used to identify any remaining relevant published studies, though none were identified. unpublished studies were not retrieved due to uncertain data quality. all studies evaluating individual symptoms and comorbidities in predicting severe infection (as measured by disease severity criteria, or icu admission) were included. all studies of any design, from any time since the outbreak started (in december ) were eligible, except case reports of individual patients or literature reviews. to avoid selection bias, no subjective quality criteria were applied to the studies for inclusion. exclusion criteria included: [ ] studies of exclusively paediatric or pregnant patients, due to the varying presentation of covid- in these groups, [ ] insufficient data on symptoms/comorbidities on admission in either severe or non-severe disease groups (or icu and non-icu all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in the copyright holder for this this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint groups), [ ] coronavirus strains other than covid- and [ ] studies not written in english, because of practical limitations with translation. two reviewers independently extracted data from the included studies for both narrative synthesis and statistical analysis. from each study, various details including the study population, investigated predictive symptoms or comorbidities, and the definitions used to measure outcomes, were extracted into microsoft excel. these details are presented by study in table . the number of patients in each study, both with and without each symptom or comorbidity, was extracted for statistical analysis (described below). the symptoms or comorbidities presented were investigated in at least three included studies. where studies measured symptoms ambiguously (including abdominal pain/diarrhea ( ) , myalgia/fatigue ( ) , and nausea/vomiting ( )), this data was excluded. some studies reported heart disease and stroke separately ( ) ( ) ( ) . to allow comparability between studies for meta-analysis, these were grouped into a single predictor (cardiovascular disease). one study was excluded from the analysis of dyspnoea as a predictor of severity, as dyspnoea was part of the definition for severity used by the authors ( ) . for disease severity, the included studies varied in their differentiation of patients' disease status, with classifications of 'mild, moderate, severe and critical' ( ) , 'ordinary and severe/critical' ( ), 'common and severe' ( ) , and 'non-severe and severe', disease ( , ) . the first outcome measure used was severe (including both severe and critical cases) vs. non-severe disease. for icu admission, the included studies varied in their definition of icu admission, with classifications of 'icu, mechanical ventilation or death and non-icu' ( ) , and 'icu and non-icu' ( , ) . the second outcome measure used was icu admission all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in patient numbers were aggregated across all included studies for each group included in the meta-analysis. gender was compared between groups using the chi test in stata ( ). this was not possible for age due to a lack of individuallevel data. the predictive value of symptoms and comorbidities for each of severe disease and icu admission was estimated with random effects meta-analysis in stata. random effects models were used to account for between study heterogeneity ( ) , which was estimated with tau-squared. this provided a pooled odds ratio (por), % confidence intervals, and a p-value, for each symptom or comorbidity. a p-value of < . was used as a marker for evidence of significant association. detailed forest plots of the predictive symptoms and comorbidities common to both disease severity and icu admission are illustrated in the supplementary information file the prisma flow diagram ( figure ) illustrates the process for selection of papers in this study. the initial search on medline, embase and global health produced results. after removing duplicates and applying exclusion criteria, there were papers meeting our criteria from title and abstract analysis. on further review, the majority of these studies did not compare proportions of patients with symptoms or comorbidities between severe (or icu admitted) and non-severe disease (or non-icu admitted) groups. the reasons for all study exclusions are outlined in figure . a total of seven studies were selected for inclusion. all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in table shows details of all included studies including reported findings pertaining to symptoms and comorbidities related to disease severity or icu admission. these studies reported on a total of patients. all included studies were retrospective cohort studies in design, conducted between december and february in china, during the novel coronavirus (sars- -cov) outbreak. guan et al ( ) conducted the largest study with covid- patients, whilst huang et al ( ) included patients in their study. the number of symptoms investigated varied from in one study ( ) to in others ( , ) . the range of comorbidities investigated varied greatly with two studies not including any ( , ) and one including comorbidities ( ) . all included studies were retrospective cohort studies, and were critically appraised using the strobe checklist ( ) . the items on the strobe checklist were formulated into individual indicators, against which each study was marked. figure illustrates the proportion of included studies which met each individual appraisal indicator. each paper was assigned an overall quality score based on the percentage of strobe checklist criteria met (< % = -, - % = +, > % = ++), as outlined in table . appraising with the strobe checklist highlighted several major weaknesses in the included studies. firstly, there was no consistent definition on what constituted severe disease. the who-china joint mission on covid- ( ) defined a severe case as tachypnoea (≥ breaths/min) or oxygen saturation ≤ % at rest, or pao /fio < mmhg. critical cases were defined as all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in the copyright holder for this this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint respiratory failure requiring mechanical ventilation, shock, or other organ failure that requires intensive care. although the above criteria were used in some included studies, many defined only one out of severe and critical, with one ( ) using a definition for a single severe/critical cohort. one study reported both severity and critical cases ( ), using criteria set by the american thoracic society to judge severity. secondly, the time at which severity of disease was determined was not always clear. severity was assessed on admission in two studies ( , ) , whilst three studies did not specify when severity was assessed ( , , ) . it is possible, therefore, that the non-severe group included patients who went on to later develop severe disease. thirdly, the time point at which symptoms were measured varied from illness onset (via recall) ( , ( ) ( ) ( ) to clinical presentation ( , ) . in the study by li et al, it was not clear when symptoms were measured ( ) . finally, no study specified how each individual symptom or comorbidity was measured. for instance, it was unclear whether fever was objectively measured, and if so, how or by whom. studies may therefore have been susceptible to measurement and reporting bias. all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in tables and show the odds ratios, % confidence intervals and p-values for the individual symptoms and comorbidities that were investigated in at least three of the included studies, for both severe disease and icu admission, respectively. a total of seven symptoms were included in the model for severe disease and six for icu admission, as well as four comorbidities in both. the most prevalent symptoms in the severe group were cough ( . %), fever ( . %) and fatigue ( . %); in the icu group these were cough ( . %), fever ( . %) and dyspnoea ( . %). the most prevalent comorbidities in the severe group were hypertension ( . %) and diabetes ( . %) and in the icu group were hypertension ( . %) and cvd ( . %). although no more likely to be in the severe group, men were . times more likely than women to be admitted to icu ( % ci . - . ). dyspnoea was the only symptom significantly associated with both severe disease (por . , % ci . - . ) and icu admission (por . , % ci . - . ), being more strongly associated with the latter. cough was associated with severe disease (por . , % respectively. in contrast, diabetes was not significantly associated with icu admission, although the tau-squared value here was unusually high implying a high level of heterogeneity between studies in this particular case. all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in were the most common symptoms found in our analysis. the prevalence of dyspnoea was not investigated in sun's meta-analysis, but we found it to be all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in the copyright holder for this this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint relatively low, particularly in non-severe and non-icu groups. the prevalence of dyspnoea in the icu group, however, was . %, compared with . % in the non-icu group. whilst dyspnoea is not a particularly common symptom in covid- patients, its significant association with both severe disease and icu admission may help discriminate between severe and non-severe covid- cases, when present. the findings reported here are in keeping with current knowledge that the elderly and those with comorbidities are more susceptible to severe infection. those with: cardiovascular disease ( . %), diabetes ( . %), chronic respiratory disease ( . %) and hypertension ( . %) ( ) . unlike the china cdc study ( ) that presented case fatality rates for different groups, our findings compare those with particular comorbidities to those without, allowing us to estimate the effect of a particular comorbidity on covid- severity. although we did not investigate death (and included copd rather than chronic respiratory disease), our analysis similarly suggests that comorbidities are not uniform in terms of the risk of severe covid- disease. despite being uncommon in our study population, copd was by far the strongest risk factor for covid- severity, followed by cvd and hypertension. the foremost limitation of this study was an inability to carry out a multivariable analysis to account for the presence of several symptoms, comorbidities and all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in the copyright holder for this this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint potential confounders. although this outbreak has seen the evolution of linked data and large datasets ( ) which would be suitable for multivariable analysis, these currently lack the quality of published data: there are large amounts of missing data, a narrow range of collected variables, and uncertainty about data collection methods and consistency. our univariable analysis is therefore valuable in evaluating specific individual symptoms and comorbidities predictive for covid- severity using high-quality evidence in the form of peer-reviewed studies. secondly, the studies included here were all from china, so the generalisability of findings to other countries and populations is not clear. the chinese may differ to other populations in terms of their health-seeking behaviour, symptom reporting, prevalence of different comorbidities, as well as their access to high quality health services. nonetheless, given the current dearth of contextually specific evidence available, our findings will help to inform future research and actions in other countries as the outbreak develops. finally, it was not possible to account for the timing of presentation in the statistical analysis. if a patient presented after many days of being symptomatic, this may have affected disease severity, compared with an earlier clinical presentation. however, this limitation does not apply to comorbidities, and table shows information from individual studies on median duration of symptoms before admission, which appears similar between severe (or icu) and nonsevere (or non-icu) cases. it is therefore unlikely that this will have biased the overall results. by identifying the symptoms and comorbidities predictive for more severe disease, clinicians can better stratify the risk of individual patients, as early as their initial contact with health services. this can lead to practical changes in management, which can improve allocative efficiency as well as clinical outcomes, through the consideration of more intensive environments of care all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in the copyright holder for this this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint (e.g. high dependency unit), earlier on, for patients at highest risk of severe infection. these can also be formalised within risk stratification tools to aid clinical decision-making, such as the curb tool for community-acquired pneumonia ( ) . as the number of hospitalised covid- cases continues to increase, hospitals will increasingly need to ration limited resources and improve clinical pathways to effectively prioritise patients with greatest clinical need. this is important, as covid- is already placing increased pressure on icus, and anticipation of future demand, based on local population characteristics, may enable more timely planning and resource mobilisation ( ) . identifying those at the highest risk will also facilitate better-informed discussions between clinicians, patients and patients' families about the anticipated clinical trajectory, allowing more accurate and timely advance care discussions to occur. identifying those at high-risk will aid the public health response in controlling the spread of disease. given the ubiquity of comorbidities in the elderly population, and their increased susceptibility to severe covid- infection ( ), knowledge on the differing prevalence and risk of various conditions may help to focus and tailor public health efforts such as the screening of asymptomatic individuals, risk communication, contact tracing, self-isolation and social distancing. for instance, for copd, which is less common in the general population and very strongly associated with icu admission, a more targeted and intensive health protection strategy may be warranted, compared to other conditions (such as hypertension) that are more difficult to target due to their higher prevalence in the general population. furthermore, if it is found that severity of illness is related to infectivity, as is the case in the closely related sars-cov, then identifying patients who may develop severe illness can help guide precautions to prevent the spread of sars-cov . these include infection control decisions regarding the limited availability of isolation rooms and personal protective equipment (ppe), particularly in more resource-constrained settings. this will be of particular importance as the outbreak develops, if the prevalence of hospitalised covid- patients increases. all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in measurement tools used in studies should aim to objectively measure symptoms where possible, and details on how and when symptoms were ascertained should be made clear in all studies. there was also heterogeneity in the range of symptoms and comorbidities recorded by different studies, and some studies grouped various symptoms ( ) ( ) ( ) , limiting the utility of such data. studies investigating covid- severity should avoid using subjective or selfreported criteria (such as dyspnoea) in the definition of severity, as in one study included here ( ) . to ensure consistency between studies and comparability, the who-china joint mission on covid- definition of severe and critical cases should be used. furthermore, the distinction between cases that are severe and critical should be clearly made in studies, to enable more accurate risk stratification of the most unwell patients. for future research on predictors of severity, research should aim to include greater detail on specific conditions, including how well controlled chronic conditions were before and during admission. if the severity of covid- varies according to the severity of underlying comorbidities, there may be a case for optimising routine treatment for healthy, uninfected individuals, as a potential public health action to mitigate risk. multivariable analysis to identify which groups of symptoms or comorbidities are most associated with severe or critical disease will also be valuable. the existing literature on covid- fails to elucidate the specific symptoms and comorbidities most predictive for severe covid- cases. our analysis finds that dyspnoea is the only symptom strongly predictive for both severe disease and icu admission, and could be a useful symptom to help guide clinical management decisions early in the course of illness. the association between comorbidities all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in the copyright holder for this this version posted march , . . https://doi.org/ . / . . . doi: medrxiv preprint and severe disease is not homogenous. whilst copd, cardiovascular disease, and hypertension were all associated with severity, copd was the most strongly predictive. when looking at icu-admitted patients, who represent the more severe end of the spectrum of clinical severity, the difference in effect sizes for copd and the other included comorbidities was large, suggesting copd patients are particularly vulnerable to critically severe disease. as the outbreak develops, future research must aim to substantiate these findings by investigating factors related to disease severity. this will aid clinical assessment, risk stratification, and resource allocation, and allow public health interventions to be targeted at the most vulnerable. all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in not in english (n= ) not on covid- ( ) literature review/letter/case report (n= ) data not available for severe and non-severe (n= ) data not available for relevant predictive factors ( ) meta-analysis ( ) not yet reported (n = ) all rights reserved. no reuse allowed without permission. perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in perpetuity. preprint (which was not certified by peer review) is the author/funder, who has granted medrxiv a license to display the preprint in the copyright holder for this this version posted march , . . 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