key: cord-022084-hap7flng
authors: ARRUDA, EURICO; CINTRA, OTAVIO A.L.; HAYDEN, FREDERICK G.
title: Respiratory Tract Viral Infections
date: 2009-05-15
journal: Tropical Infectious Diseases
DOI: 10.1016/b978-0-443-06668-9.50064-8
sha: 
doc_id: 22084
cord_uid: hap7flng

nan

Acute respiratory infections (ARIs) are prevalent worldwide 1 and rival diarrhea as the leading cause of death in developing countries. 1, 2 In some impoverished urban populations in South America, ARI symptoms may be present on an almost continuous basis, making it difficult to determine symptom-free days and estimate attack rates. 3, 4 Children from these areas may spend 40% to 75% of their time with respiratory symptoms, 1,5 mostly caused by upper respiratory infections (URIs). The most striking disparity between developing and developed countries with regard to ARI epidemiology is the case-fatality rate of lower respiratory infection (LRI), mainly pneumonia, bronchiolitis, and influenza, 6, 7 in children under 5 years of age, which may reach 16% in some areas. 1, 8 Several community-based studies have established the importance of common respiratory viral infections in tropical countries 5 (Table 59-1) . In impoverished populations, these common viral infections may occur simultaneously with measles, diarrhea, and malnutrition, resulting in complex interactions of pathologic conditions that carry the potential to become life-threatening diseases. 1, 9 Unlike certain pathogens restricted to tropical areas, the respiratory viruses have worldwide distribution, efficient person-to-person transmission, and an impact on all age groups. Except for a few agents (e.g., adenoviruses, severe acute respiratory syndrome [SARS] coronavirus) and rare cases of extrapulmonary dissemination with other respiratory viruses, replication is generally restricted to the respiratory mucosa of humans.

In most health-care facility-based studies of acute LRI (ALRI) conducted in tropical countries (Table 59 -2), respiratory syncytial virus (RSV) is the virus most frequently detected (11% to 33%), followed by parainfluenza viruses (1% to 13%), adenoviruses (2% to 34%), and influenza viruses (1% to 4%). With few exceptions, human rhinovirus (HRV) and human coronaviruses (HCoV) have not been reported frequently in studies in tropical countries, 5 probably because of difficulties in their detection.

Although previous studies have shown that attending daycare centers can be a risk for ARI, 10 providing day care for children has become an important economic issue in developing countries, where mothers must join the workforce to contribute to the family income. Consistent with studies in the United States and elsewhere, a study found a high burden of ARI in young, low-income children attending day care in Salvador, Brazil. 11 Few specific interventions are available to reduce the impact of respiratory viruses, 2 and the application of these interventions may be further hampered by epidemiologic patterns in ARI and socioeconomic differences in temperate, developed countries compared with equatorial regions. For example, housing conditions and crowding pose challenges for optimizing health-care strategies in the tropics. While the incidences of some respiratory viruses, particularly RSV and influenza, show seasonal trends in some tropical areas, the association of seasonal peaks of respiratory viruses in general may be less apparent where fluctuations in temperature are smaller. 12 Nutritional and educational interventions, such as reinforcing breast-feeding, 13 vitamin A supplementation for measles, 14 and facilitation of access to oral rehydration therapy, 15 may have significant effect on the morbidity and mortality due to LRI alone or in association with diarrhea.

In this chapter we focus attention on the most common viral respiratory infections, whose main features are summarized in Table 59 -3, and try to highlight features unique to the developing world.

In tropical countries influenza activity may occur yearround as well as in outbreaks more typical of temperate regions. These infections cause serious disease in populations weakened by malnutrition, with limited access to medical care. 16 Of note, the predisposition induced by influenza to superimposed bacterial infections, mainly Streptococcus pneumoniae, may greatly affect morbidity and mortality, mainly among impoverished populations. 17 In addition, influenza viruses can reassort or sometimes cross species barriers to generate emergent strains that may cause localized outbreaks or potentially pandemics with enormous impact for health on a global scale. 18 

Influenza viruses are pleomorphic, enveloped, with segmented negative-strand RNA genomes and belong to the family Orthomyxoviridae. Influenza viruses are distributed in three genera-A, B, and C-based on the antigenicity of the nucleoprotein (NP) and matrix protein. Influenza A virus is further classified in subtypes based on its two surface glycoproteins: hemagglutinin (HA) and neuraminidase (NA). 19 Among the 15 HA and 9 NA subtypes recognized in nature, 6 HA (H1, H2, H3, H5, H7, and H9) and three NA (N1, N2, and N7) subtypes have now been identified in human isolates of influenza A viruses. 18 However, only three subtypes of HA (H1, H2, and H3) and two of NA (N1 and N2) have caused pandemics and sustained circulation in human populations in recent years. 20 The genomes of influenza viruses contain eight RNA segments in influenza A and B viruses, and seven RNA segments in influenza C. 19 The glycoprotein HA is responsible for the attachment of the virus to sialic acid-containing cell receptors, and it mediates fusion and penetration. Proteolytic cleavage of HA by cellular serine proteases exposes hydrophobic fusion domains that mediate membrane fusion. The NA cleaves terminal sialic acid from glycoconjugates present on respiratory mucins, cells, and progeny virions. This action destroys receptors recognized by HA and allows budding virus to be released from infected cells and to spread within the respiratory tract. Influenza C virus contains a single surface glycoprotein that binds to receptor, promotes fusion of membranes, and also cleaves sialic acid. 19 Virus binding to receptor is followed by internalization into endosomes, fusion of viral and endosomal membranes, and release of the genome to the cytoplasm, from where it is transported to the nucleus. In influenza A viruses, the M2 protein serves an ion channel function that facilitates dissociation of the RNA segments from the virion interior. Transcription of the negative-strand genomic RNA into positive-strand messenger RNA (mRNA) and complementary RNA (cRNA) is mediated by a viral RNA polymerase complex in the nucleus. cRNA serves as a template for the synthesis of negative-strand virion RNA genome segments, and mRNA directs viral protein synthesis. Newly assembled nucleocapsids acquire an envelope as they bud through the cell surface. Only viruses with a full complement of genome segments are infectious. 19 Influenza A viruses are primarily viruses of aquatic birds, particularly ducks and shore birds, which harbor all subtypes recognized to date. Selected subtypes naturally infect a range of terrestrial (swine, horses, humans) and aquatic (seals) mammals; influenza B virus infects humans and uncommonly seals, dogs, cats, and swine, and influenza C virus is primarily a virus of humans. Depending on the virus type and subtype, experimental infection can be induced in mice, ferrets, chickens, swine, and primates, and the viruses can be propagated in primary cultures of kidney cells, continuous cell lines (MDCK, Vero, PER.C6, and LLC-MK2), and also in embryonated hen' s eggs. 20 The biologic property of influenza viruses to bind erythrocytes is exploited for early detection of the virus in cell culture and for the development of serologic assays by hemagglutination inhibition. 20 Influenza viruses are inactivated by temperatures above 50°C and by lipid solvents, acid, formaldehyde, ionizing radiation, and ultraviolet (UV) light. 20

Influenza viruses occur throughout the world, causing highly contagious respiratory infections with high morbidity and excess mortality (in seasonal peaks), particularly in infants and the elderly. In developing tropical countries, influenza has been associated with an average of 5% of ARIs leading to physician contact. 6, 21 This apparently low proportion probably represents only the most severe cases, since 30% to 50% of children under 5 years of age in tropical Africa have been found to seroconvert in one outbreak. 21 Previously healthy children younger than 1 year of age are hospitalized for influenza at rates similar to those for adults at high risk for influenza, and influenza accounts for a great number of outpatient visits and courses of antibiotics in children of all ages. 22 When human influenza virus is introduced into a malnourished population with limited access to care, high morbidity and mortality rates can occur, as was observed in Madagascar in 2001 where a conventional influenza A/H3N2 subtype virus was associated with case-fatality rates of approximately 3%.

Contrary to the remarkably sharp seasonality of influenza A outbreaks in temperate countries, seasonal patterns in tropical countries have varied between studies. In southern India 23 and Thailand 24 influenza has occurred throughout the year with sporadic outbreaks, whereas there have been consistent outbreaks in June-July and November-January, coinciding with the winter seasons in the southern and northern hemispheres, but with no apparent association with meteorologic factors. 12 In the Philippines influenza has been more frequent between November and January, 25 while in Senegal, Nigeria, and Taiwan there has been clear association with increased rainfall. 12 In southeastern Brazil, 26 Argentina, 27 as well as in South Africa, 28 seasonal outbreaks of influenza A have occurred annually from May through August (mid autumn through winter) in association with cooler temperatures but not with rainfall. Influenza B outbreaks occur periodically, yet less frequently than influenza A, in both temperate and tropical regions, 12, 20 whereas influenza C is generally nonseasonal. 20 Minor changes in antigenicity, called antigenic drift, are caused by accumulation of point mutations in the genes coding for influenza HA and NA, generating new strains that spread in annual epidemics. Influenza viruses B and C are less prone to antigenic drift. Major antigenic changes in influenza A are called antigenic shift and result in the emergence of a novel HA subtype with or without new NA to which humans lack significant immunity. 18 This may be caused by the acquisition of new gene segments through genetic reassortment in a host infected simultaneously by a human and an animal (typically avian) virus, or by reappearance of a subtype from reservoir. Swine are susceptible to both avian and human influenza viruses and may be hosts for reassortment or serve as the mammalian species in which avian viruses can adapt. Novel influenza virus subtypes generated by shift have caused catastrophic pandemics, including three in the last century. The H1N1 "Spanish flu" pandemic of 1918 is estimated to have caused up to 100 million deaths worldwide, 18 while the H2N2 "Asian flu" in 1957 and the "Hong Kong flu" in 1958 caused an estimated 1 to 3 million deaths.

Recent clusters of human infections due to avian influenza, particularly H5N1 subtype viruses in Asia, have raised concerns about new pandemic threats. In 1997 a highly pathogenic avian influenza H5N1 virus resulting from reassortment among several avian viruses caused lethal outbreaks in domestic poultry and severe illness with 6 deaths among 18 human cases in Hong Kong. The outbreak was due to exposure to infected poultry in live poultry markets and was later contained by their slaughter. This virus was transmitted inefficiently from person to person. 18, 20 In February 2003 an H5N1 virus caused deaths in a family visiting Fujian province in China. Since late 2003 wide-scale poultry outbreaks due to H5N1 virus have occurred in at least 10 Asian countries. Interspecies transmission to humans occurred in at least two countries, causing over 45 cases and 32 deaths in Vietnam and Thailand. 29 These H5N1 viruses have continued to reassort, evolve antigenically, and extend their host range with documented infections in swine and felids. Prolonged nonsymptomatic excretion in ducks and detection in migratory birds indicate that this virus has become endemic in Southeast Asia. 18, 30 An avian H9N2 virus with human receptor specificity has also spread throughout Asia in domestic poultry and pigs and caused mild disease in humans in Hong Kong and China. 18 An avian H7N7 virus caused conjunctivitis in at least 82 people and one fatal pneumonia in The Netherlands in 2003; the outbreak was contained by culling and quarantine of poultry. In February 2004 a highly pathogenic H7N3 virus emerged in domestic poultry in British Columbia with two documented human cases of conjunctivitis and mild "flulike" illness. 18, 29 Influenza virus is transmitted from person to person by large droplets and small-particle aerosols, as well as possibly by fomites with hand contamination and subsequent selfinoculation. The relative importance of these routes is uncertain for natural influenza. Ingestion of infected birds has led to infection by avian viruses in cats. Secondary attack rates may reach over 70% in semiclosed populations, especially among schoolchildren and patients debilitated by underlying conditions who live in relative confinement, such as nursing home residents. 20 Children play a major role in influenza outbreaks with respect to propagation of the epidemic virus in families and communities. 20

Classic influenza starts abruptly after an incubation period of 1 to 2 days, with fever, chills, malaise, headache, myalgia, and prostration, often accompanied by nonproductive cough, sore throat, and mild rhinorrhea. Systemic complaints last 3 to 5 days, whereas sore throat, hoarseness, and cough, with substernal discomfort, may increase in severity as the systemic symptoms subside. Cough and asthenia often persist for 2 weeks or longer. Respiratory symptoms may be minimal or absent initially, especially in the elderly or infants. In frail elderly persons, lassitude, lethargy, confusion, lowgrade fever, and sometimes gastrointestinal complaints may be the primary findings. Influenza B tends to be milder than influenza A, and influenza C typically causes colds or bronchitis. 20 Influenza may also present as unexplained fever, croup (laryngotracheobronchitis), vomiting, diarrhea, and neurologic manifestations in young children. 31 Up to 50% of influenza virus infections in adults are subclinical. 20 Influenza causes a variety of viral respiratory complications, including otitis media, sinusitis, tracheobronchitis, pneumonia, and, in young children, bronchiolitis and croup. Secondary bacterial infections, especially pneumonia caused by Staphylococcus aureus, Streptococcus pneumoniae, and Haemophilus influenzae, are common complications and should be suspected in relapses of fever, chest pain, and cough. 20 Influenza is also associated with invasive meningococcal infections. Other complications include exacerbations of asthma, chronic bronchitis, and congestive heart failure. Myositis, myoglobinuric renal failure, meningoencephalitis, transverse myelitis, polyneuritis, parotitis, myocarditis, arthritis, and disseminated intravascular coagulation rarely occur after influenza. Reye' s syndrome occurs in fewer than 1 per 100,000 cases of influenza in patients under 18 years of age following the use of salicylates. Pregnant women, human immunodeficiency virus (HIV)-infected patients, and other immunocompromised hosts are at higher risk for severe disease and complications. 20

The virus infects the respiratory mucosa, where it causes lytic infection of cells and desquamation of the respiratory epithelium, mononuclear cell infiltrates in the lamina propria, and altered mucociliary clearance. Tracheobronchitis is a typical feature and often associated with prolonged abnormalities in small airway pulmonary function and airway hyperreactivity. Primary influenza viral pneumonia results in diffuse alveolar damage, alveolar hemorrhage and exudate, hyaline membranes, and later reactive fibrosis. Fatal cases show pathologic changes in nonrespiratory organs, such as brain congestion and swelling, myocardial inflammation, and fibrinoid changes in arterioles. 20 Viral replication in the upper respiratory tract generally peaks within 1 or 2 days of symptom onset and, depending on age and prior immunologic experience, continues for about 3 to 8 days. The severity of illness broadly correlates with upper respiratory tract viral levels. Constitutional symptoms with influenza are due in part to the release of proinflammatory cytokines and chemokines. Levels of interferon (IFN-α and IFN-γ), tumor necrosis factor (TNF-α), interleukins and chemokines (IL-1β, IL-6, IL-8, IL-10, MCP-10, MIP-1α and MIP-1β) are increased in nasal secretions, and IFN, IL-6, and TNF-α are increased in blood in human influenza. 20 The tissue tropism of a strain of influenza virus depends, among other factors, on a combination of susceptibility of its HA to be cleaved by, and tissue availability of proteases with specificity to cleave it, thus rendering the virus infectious. 32 Extrapulmonary dissemination of virus has been uncommonly documented in humans, but systemic spread is a regular feature of highly pathogenic avian viruses in chickens and sometimes in rodents or other mammalian hosts. Serum and secretory antibodies directed to HA and NA appear about 10 days after infection. Protection against reinfection by the homologous strain is durable following natural infection and is correlated with serum and nasal neutralizing antibody levels, principally directed against HA. Vaccine-induced protection may last for up to 2 to 3 years against homotypic virus. Infection also induces cell-mediated immunity, which is detectable 3 to 6 days after infection and seems to be important for recovery. 30 Cytotoxic T-lymphocyte responses against internal proteins may provide some degree of heterosubtypic immunity.

The diagnosis of influenza is frequently made on the basis of clinical and epidemiologic information. A higher index of suspicion and laboratory diagnostics is needed outside the season, particularly in sporadic individual cases or unexplained outbreaks of febrile respiratory illness. Viral isolation from respiratory specimens can be done in several types of cells (e.g., PRMK, MDCK, LLC-MK2) and remains the current standard. 20 The presence of virus may be detected in cell cultures by hemadsorption with guinea pig erythrocytes before or after cytopathic effect (CPE) is visible. Blind hemadsorption is positive 3 days after inoculation in almost all positive samples. 20 Confirmation of isolates can be done by hemagglutination inhibition or immunofluorescence with typespecific antisera. Diagnosis can also be made in 1 to 2 days by immunofluorescence of monolayers of MDCK cells inoculated by centrifugation (shell-vial). 33 Conserved influenza antigens (M or NP) directly in clinical samples can be detected by one of several techniques (e.g., immunofluorescence [IF], enzyme immunoassay [EIA]), and multiple point-of-care kits are commercially available with turnaround time of 15 to 30 minutes. One commercial assay is based on detection of influenzaspecific NA activity. The sensitivities of these assays are higher in children (up to 90%) than in adults (generally 50% to 70%) and depend on duration of illness and sample type. 20 Several formats of reverse transcription-polymerase chain reaction (RT-PCR) assays have been used for the detection of influenza A and B RNAs in clinical samples, with the advantage of detecting genomes of noninfectious virus. 20 The time to perform RT-PCR is longer but the cost may be lower than for commercial rapid antigen detection kits, especially in developing countries. Real-time RT-PCR has enabled the development of assays that provide rapid quantitative detection of influenza A and B with high sensitivity. 34, 35 These assays have great potential to replace other methods, because they are simultaneously rapid, highly sensitive, quantitative, and amenable to being used in multiplex format, which might include probes for several different respiratory pathogens. 35 However, the costs are still prohibitive for most laboratories in developing nations. Serologic diagnosis of influenza using paired acute and convalescent serum can be done retrospectively by a variety of techniques but mainly for serologic survey purposes. 20

Amantadine and rimantadine are M2 ion channel blockers that inhibit influenza A virus replication at the uncoating step. 20 In uncomplicated influenza A in adults without underlying diseases, treatment with either drug can reduce the duration of influenza illness by approximately 1 to 2 days if started early, within 48 hours from the onset of symptoms. Amantadine is excreted in an unchanged state in the urine, while rimantadine is extensively metabolized after absorption and less than 10% of the dose is excreted unchanged in the urine. Elderly persons need only half the dose to achieve similar plasma levels. Amantadine or rimantadine may cause gastrointestinal upset and central nervous system side effects. Central nervous system (CNS) intolerance is more common with amantadine and, when severe, can be manifested as agitation, psychosis, seizures, and coma. Mild complaints including insomnia, dizziness, anxiety, dry mouth, anorexia, and nausea are reversible upon discontinuation. Amantadine and rimantadine are marketed as 100-mg tablets and 10-mg/mL syrup. The recommended dose is 100 mg twice daily for adults older than 65 years of age (100 mg/day for patients ≥65 years of age). For children under age 10 years, a rimantadine dose of 5 mg/kg/day (maximum, 150 mg/day) has been suggested. 20 Dose reductions proportional to the creatinine clearance (ClCr) are suggested for patients with renal insufficiency (amantadine for ClCr <60 to 80 mL/min/1.73 m 2 ; rimantadine for ClCr <10 to 20 mL/min/1.73 m 2 ). Influenza virus resistant to amantadine-rimantadine emerges in approximately one third of treated patients; such viruses are transmissible to close contacts and cause typical influenza illness. Resistance to these drugs renders them ineffective and is sometimes present naturally, including in recent human isolates of H5N1 virus. 20 The neuraminidase inhibitors zanamivir and oseltamivir inhibit both influenza A and B viruses by blocking the active site of the enzyme for cleavage of sialic acid, thus inhibiting virus release from infected cells and spread within the respiratory tract. 36 In adults and children older than 5 years, inhaled zanamivir (10 mg twice daily for 5 days) provides 1to 2.5-day reduction in illness 37 and reduces antibiotic use for lower respiratory complications by 40%. Zanamivir is generally well tolerated but may uncommonly induce bronchospasm, particularly in those with influenza and pre-existing airway disease. 20 Oseltamivir (75 mg orally twice daily for 5 days) reduces illness severity, time to resumption of daily activities by 1 to 3 days, and rates of complications leading to antibiotic prescription and hospitalization by about 50% in adults. In children 1 to 12 years of age, oseltamivir reduces the frequency of otitis media and, consequently, antibiotic prescriptions. Side effects include mild-to-moderate nausea or emesis. Dosage of neuraminidase inhibitors does not need to be adjusted for the elderly. 20 Resistance emergence is uncommon with both drugs, 20 although a recent study of children treated with oseltamivir detected drug-resistant viruses in 18%, often in association with prolonged viral excretion, and showed that children can be a source of viral transmission, even after 5 days of treatment. 38 Antipyretic-analgesic drugs may be used for influenzainduced fever and aches. Aspirin should be avoided because of its association with Reye' s syndrome.

Immunization with formalin-inactivated or live-attenuated multivalent influenza virus vaccines and chemoprophylaxis for influenza virus A are the methods available for preventing influenza. Influenza vaccine is used prior to the influenza season and currently includes one strain of influenza B and two strains of subtypes H3N2 and H1N1 of influenza A virus, chosen by the World Health Organization (WHO) surveillance network among the viruses most likely to circulate in the next influenza season. 20, 39 The inactivated vaccine has an approximate 70% to 90% efficacy in preventing illness in healthy children and adults. 39 It also reduces influenza-related hospitalizations and mortality in elderly and high-risk patients. The Centers for Disease Control and Prevention (CDC) recommends the immunization of persons aged 50 years and older; residents of nursing homes; children and adults with chronic cardiovascular or pulmonary disease, including asthma; persons chronically ill with diabetes mellitus, renal dysfunction, or hemoglobinopathies; immunosuppressed patients including those with HIV infection; children and adolescents on chronic aspirin therapy who may develop postinfluenza Reye' s syndrome; women who will be pregnant during the influenza season; children aged 6 to 23 months; those who can transmit influenza to persons at high risk, such as health-care workers and household contacts of those at high risk including children 0 to 23 months of age; crew members of cruise ships; providers of essential services; and unimmunized travelers to areas where influenza may be circulating, including the tropics, the southern hemisphere between April and September, and those traveling in large organized tourist groups. In addition, vaccine is made available to anyone interested in reducing the likelihood of becoming ill with influenza. 20, 39 The inactivated vaccine, administered as a single intramuscular (IM) dose shortly before influenza season (two doses in previously unimmunized children <9 years of age), is safe during pregnancy but should be avoided in persons with history of anaphylactic reactions to eggs. 39 Vaccine safety and efficacy in children has been extensively evaluated and has shown a favorable safety profile with efficacy in 1-to 15-year-old children of 77% to 91%. Inactivated vaccine is not currently recommended for children younger than 6 months, but vaccination of household contacts and caregivers should reduce the risk of these high-risk children contracting influenza. Healthy people aged 5 to 49 years who are not contacts of immunosuppressed patients can receive either inactivated or intranasal live-attenuated vaccines. 39 Influenza inactivated vaccine has been recently introduced in many tropical areas of the world, with a composition based on influenza viruses circulating in the southern hemisphere. The vaccine is given prior to the influenza season, which for most countries in the southern hemisphere is between May and July. 40 In South America, annual vaccination of the elderly has reduced hospitalizations and mortality for respiratory diseases. 41 Continuous surveillance has already shown that regional variations of circulating influenza virus strains should be taken in consideration in the formulation of influenza vaccines with compositions more appropriate for South America. 42 Live-attenuated, cold-adapted vaccines administered intranasally are well tolerated, genetically stable, and rarely transmissible and have the advantage of inducing local secretory immunoglobulin A (IgA) responses. Because of potential interference between components, two doses may be required in young children. 20 This vaccine was licensed in the United States in 2003, where it has become an option for healthy persons aged 5 to 49 years, including those in close contact with groups at high risk and those wanting to avoid influenza. 39 This vaccine is not recommended for persons with asthma and other chronic disorders of the pulmonary or cardiovascular systems; persons with underlying medical conditions, including diabetes, renal dysfunction, and hemoglobinopathies; persons with known or suspected immunodeficiency diseases or who are receiving immunosuppressive therapies; children or adolescents receiving aspirin or other salicylates; persons with a history of Guillain-Barré syndrome; pregnant women; and persons with a history of hypersensitivity to eggs. 39 Cold-adapted trivalent influenza vaccine is highly effective (92% in phase 3 studies) in preventing cultureconfirmed influenza in healthy children and has provided protection against drift variant strains in some studies. In young and middle-aged adults, efficacy is generally comparable to that of inactivated vaccine. 39 Other investigational approaches have been explored in influenza vaccine development, including recombinant HA produced in insect cells, virosomes incorporating surface glycoproteins, M2 protein conjugated with hepatitis B virus core, and naked DNA encoding influenza virus nucleoprotein or HA. 20 Cell culture-based vaccines (MDCK, Vero) have been approved in Europe and may offer an alternative to the limitations of the current egg-grown vaccines. The technique of reverse genetics has been used to rapidly produce candidate vaccines against potential pandemic threat viruses.

Amantadine and rimantadine are approved for use, and are 70% to 90% effective in the prophylaxis of influenza A during outbreaks. Unvaccinated elderly persons, immunodeficient patients, patients in chronic care institutions experiencing outbreaks, persons who could not be vaccinated, and those who received a vaccine strain different from the outbreak strain may receive prophylaxis with amantadine or rimantadine. Prophylaxis should be started as early as possible at doses equivalent to those used for therapy, and continued until 1 week after the end of the outbreak for a total of at least 2 weeks. 39 Amantadine-and rimantadine-resistant mutants of influenza A virus occur in up to 30% of treated patients and may be associated with failure of drug prophylaxis. 43 Both oseltamivir (75 mg twice daily) and inhaled zanamivir (10 mg/dose twice daily) are more than 80% effective in the prophylaxis of influenza during outbreaks, but only oseltamivir has been approved for this indication in the United States. 20, 39 Antiviral agents, especially the neuraminidase inhibitors, could significantly help in the control of a future influenza pandemic by reducing lower respiratory complications and hospitalizations as well as potentially person-to-person transmission. However, supply limitations 44 pose a real difficulty. Therefore, policies to ensure a reasonable supply of these drugs, as well as directions to optimize the use of limited supplies, are important issues to be considered. 18 

Respiratory syncytial virus (RSV) is the single most important viral cause of lower respiratory disease and a major cause of morbidity and mortality in children worldwide. RSV is the leading cause of hospitalization in young children in developed and developing countries. 45 In tropical areas, RSV has been the most frequently isolated virus in hospital-based ARI studies of children. 5 AGENT RSV, the only known human pathogen of the genus Pneumovirus in the family Paramyxoviridae, is a pleomorphic RNA virus with helical nucleocapsid and lipid-containing envelope. Antigenic differences in the surface glycoprotein G permit the classification of RSV into groups A and B, each with antigenic subgroups. 46 The interaction of RSV envelope glycoprotein G with glycosaminoglycans enables adherence to the cell surface. However, G protein-independent mechanisms of attachment must exist, since mutants devoid of G protein can also enter host cells. RSV enters the cell by fusion of viral envelope with cell membranes, a process mediated by binding of the viral F protein to the cell GTPase RhoA. 46 The syncytia resulting from fusion of the infected cells to adjacent ones are the major feature of the cytopathic effect of paramyxoviruses. Once in the cytoplasm, the negative-strand RNA is transcribed by viral transcriptase into mRNAs, which then direct viral protein synthesis. An intermediate positivestrand full-length cRNA serves as a template for the synthesis of progeny negative-strand RNA. As they bud through the cell membrane, the virions acquire a glycoprotein-containing envelope. 46 RSV causes asymptomatic infection in a variety of experimental animals, but natural infection occurs only in humans and chimpanzees. 47 RSV grows well in several human heteroploid cell lines, such as HEp-2, HeLa, and A549, and is sensitive to ether, chloroform, detergents, and a pH less than 5. RSV is inactivated at 55°C, survives poorly on porous surfaces, and loses infectivity significantly by slow freezing and storage at temperatures above 4°C. 46 

RSV occurs worldwide and causes annual outbreaks in temperate climates in the winter and early spring, with sporadic cases throughout the year. 47 In tropical regions, where temperature fluctuations are smaller and the only significant seasonal variable is often rainfall, RSV outbreaks tend to occur in the rainy seasons. Such has been the case in Malaysia, Hong Kong, India, Papua New Guinea, Colombia, Kenya, and The Gambia. 12 Interestingly, in Singapore RSV peak activity occurs from March to August, a period of higher temperature, higher day-to-day temperature variation, and lower relative humidity. 48 In southeast Brazil, RSV occurs seasonally, within a broader range of months from February through July, after the rainy season and when temperatures tend to be cooler, with slight variations from year to year. 49 In regions where average winter temperatures are colder, such as in São Paulo city and the southernmost parts of Brazil, as well as in Argentina, RSV peak activity tends to occur in July and August. [50] [51] [52] Most children have specific serum RSV antibody by age 2 years, but reinfections occur throughout life. More than one subtype of either RSV group may cocirculate in one season, with group predominance changing from year to year, without apparent correlation with clinical or epidemiologic characteristics of the illness they cause. 49,53 RSV transmission requires close contact and occurs either by large-particle aerosols or by contamination of hands and inoculation into the eye or nose. Secondary infections in family contacts of an index case are common, after an average incubation period ranging from 2 to 8 days. 45 It is estimated that 30% of all infants will have RSV infection that requires medical attention and that 2% of them will be hospitalized. 45 An estimated 10% of children will have bronchiolitis in their first year of life, with 60% to 90% of those infections caused by RSV. 54 In southeast Brazil, RSV is the leading cause of lower respiratory tract infections in children younger than 1 year of age and is responsible for up to 85% of hospitalizations in this age group during peak months. 52

The spectrum of illnesses caused by RSV ranges from mild URI to severe LRI, including pneumonia, bronchiolitis, tracheobronchitis, and croup. 45 In infants and young children, URI with fever and otitis media is common. During outbreaks, RSV RNA has been detected in up to 75% of middle ear effusions in children with RSV infection and acute otitis media. 55 The most frequent LRI caused by RSV in infants is bronchiolitis, usually preceded by 2 to 3 days of URI symptoms, and progressing to lower respiratory tract involvement characterized by tachypnea, dyspnea, cough, expiratory wheezing, air trapping, and in more severe cases, intercostal muscle retractions and cyanosis. Fever is present in only 50% of infants. Chest radiographs may show hyperaeration of the lungs and sometimes segmented atelectasis. 45 Blood counts usually show lymphocytosis, and an increase in neutrophils with a left shift could be associated with bacterial superinfection. The most frequent bacterial superinfection in children with RSV infections is acute otitis media, which may be found in up to 60% of children with brochiolitis. 56 However, more serious bacterial infections that may require sepsis work-up is uncommon in previously healthy infants with RSV infections. 57 This may be different, however, in developing tropical areas, where RSV frequently causes infections in children previously debilitated by other diseases and malnutrition.

Infants with congenital heart disease, premature infants, or infants with underlying pulmonary conditions, such as cystic fibrosis and bronchopulmonary dysplasia, as well as immunocompromised hosts of any age, are at risk for severe and fatal RSV infections. HIV-infected children with RSV infections have a higher rate of pneumonia and prolonged illness and virus shedding, but the general severity of the RSV disease is not increased. 45 Differential diagnosis of acute bronchiolitis includes asthma, pneumonia, congenital heart and lung diseases, and cystic fibrosis. Particular clinical signs are generally not accurate predictors of specific viral causes, but in a study conducted in the Philippines, wheezing was a significant predictor of viral LRI, while manifestations of higher severity, such as chest indrawing and cyanosis, were more often associated with bacterial LRI. 58 The most frequent RSV illness in children over 3 years of age and adults is URI with coryza and cough, sore throat, and hoarseness, often accompanied by low-grade fever. Exacerbations of chronic pulmonary diseases and wheezing can also be seen in adults with RSV infection. 45 The role of RSV infections in causing wheezing and asthma exacerbations in infants is well established in studies conducted in temperate areas. 59 Similar observations have been made in an emergency room study conducted in Southeast Brazil, which found that infection with respiratory viruses, especially RSV, and a family history of allergy were independently associated with wheezing. 60 Similar findings have been observed in urban Nigerian preschool children. 61 RSV has been increasingly recognized as a cause of LRI in the elderly, mainly characterized by interstitial pneumonia, prolonged cough, and dyspnea in persons with chronic pulmonary conditions, and it should be considered in the differential diagnosis of flulike illnesses. 62

RSV replicates in respiratory epithelium to reach titers as high as 10 6 TCID 50 /mL in nasal secretions of infected babies, and virus shedding may be as prolonged as 3 weeks after the symptoms disappear. 46 RSV spreads from cell to cell and may involve the entire respiratory tree, reaching bronchioles in 1 to 3 days after the onset of rhinorrhea. Replication in the bronchiolar epithelium causes necrosis of ciliated cells, syncytia formation, peribronchiolar inflammation with abundant lymphocytes and macrophages, and impairment of secretion clearance, resulting in small airway obstruction and the hyperinflation characteristic of bronchiolitis. Pneumonia frequently coexists, evidenced by interstitial mononuclear infiltrate, eosinophilic cytoplasmic inclusions in epithelial cells, and multinucleated giant cells. The most severe RSV disease occurs in young babies, whose immature airways may be unable to compensate for the pathologic changes. 46 Naturally acquired immunity to RSV is incomplete and short-lived, but the severity of illness tends to decrease with reinfections. Local secretory IgA correlates better with protection than does serum antibody level and age, and pre-existing virus-specific maternal antibodies influence the development of neutralizing antibodies. Cell-mediated immune response is central to recovery from RSV infection, and patients with suppressed cell-mediated immune response are at risk of severe RSV pulmonary disease and fatal outcome. 45, 46 The type of immune response to the virus is probably a major factor in the development of wheezing and asthma exacerbations. A bias toward a Th2 cytokine response seems to be associated with more severe disease, whereas a Th1 response leads to effective viral clearance and milder illness. The virus itself generally triggers a Th1 response, but a preexisting Th1 deficiency may be associated with disease severity in some children. It has been suggested that RSV bronchiolitis may be a marker of predisposition to wheezing or asthma later in life. 46, 63 Children vaccinated with a formalin-inactivated RSV vaccine developed in the 1960s had severe disease when exposed to natural infection, apparently as a consequence of an imbalance between protective and immunopathologic T-cell responses elicited by previous parenteral immunization with inactivated RSV. This would favor a CD4+ Th2 cytokine pattern in response to subsequent RSV infections, whereas a previous natural infection would favor a CD4+ Th1 pattern in response to reinfection. 64

Nasopharyngeal aspirates or swabs, nasal washings, and lower respiratory samples are all appropriate specimens for RSV isolation. This is usually accomplished in cultures of HEp-2 cell line, in which RSV induces syncytia in 3 to 5 days. RSV antigen detection by EIA, including membrane-based EIA, is sensitive and specific and requires virtually no equipment, making it ideal for field studies. Rapid RSV detection by IF of exfoliated respiratory cells may be even more sensitive than EIA-based methods. 45, 46 The increasing use of rapid tests has facilitated the assessment of RSV in tropical areas. 1 Ideally, a combination of a rapid method with viral isolation should be used for maximal RSV detection, but the cost may still be prohibitive for the meager resources available in some tropical areas. Detection of RSV RNA by conventional RT-PCR has shown suboptimal sensitivity, especially when compared with easy-to-perform, more sensitive rapid methods. 45 However, more recently developed assays based on real-time RT-PCR are proving to be more sensitive than conventional RT-PCR assays, with the added conveniences of being rapid, quantitative, and amenable to simultaneous detection and subtyping of RSV directly from clinical specimens. 65 RSV serology has limited value for case management but may be useful for epidemiologic surveys. 45 TREATMENT URI caused by RSV requires no specific treatment, and antibiotics are needed only when bacterial otitis media or sinusitis are present. 45 The supportive treatment of infants with RSV bronchiolitis consists basically in preventing hypoxemia and electrolyte imbalance, in addition to aerosolized bronchodilators. The lack of obvious correlation between radiologic findings and disease severity suggests that a chest film should be recommended only for severely ill or deteriorating infants. 54 To prevent hypoxemia, requirements may vary from simple removal of respiratory secretions and proper positioning of the infant to mechanical respiratory assistance and even extracorporeal membrane oxygenation (ECMO). Pulse oximetry has been advocated to assess oxygen needs, but in tropical developing areas, where oximeters may not be available, serial clinical assessment is essential to monitor disease progression. For this purpose, crackles and cyanosis seem to correlate better with hypoxemia than tachypnea and intercostal retraction. 54 Correction of hypoxemia can be accomplished with 40% or lower oxygen concentrations. 45 Oxygen should be humidified with saline and delivered by mask if head boxes or tents are unavailable. The role of corticosteroids remains unclear with some evidence that they are not beneficial. 54 The only antiviral drug currently approved for the treatment of infants with RSV is the synthetic nucleoside ribavirin, delivered by small-particle aerosol via a mist tent, mask, oxygen hood, or ventilator. It is recommended only for infants and young children with an underlying condition, such as congenital heart disease, cystic fibrosis, or immunosuppression. Premature infants, infants younger than 6 weeks of age, and severely ill infants may also be considered for therapy. 66 Aerosolized ribavirin is well tolerated, but it is expensive and its prolonged administration requires facilities that may not be available in impoverished tropical areas. Passive immunotherapy with RSV immunoglobulin, in combination with aerosolized ribavirin, improved the outcome of RSV pneumonia in bone marrow transplant patients. 67 The use of RSVintravenous immunoglobulin (IVIG) or humanized monoclonal antibody against RSV has shown no benefit for the treatment of RSV infections in infants. 45

No vaccine is currently available for RSV prophylaxis. The disease enhancement caused by formalin-inactivated vaccine in the 1960s plus results of more recent unsuccessful trials of live-attenuated vaccines, have significantly slowed progress toward an RSV vaccine. 2 Purified fusion protein vaccine has been tested for safety and immunogenicity in seropositive children older than 18 months, and was associated with reduction of lower respiratory tract illness, but not of RSV infection rates, in children with cystic fibrosis. 45 These and other candidate subunit vaccines, as well as intranasal liveattenuated vaccines, should be tested in high-risk children with underlying bronchopulmonary diseases.

Passive immunization of high-risk infants with monthly infusions of RSV immunoglobulin during the RSV season reduced the incidence and severity of RSV infections in highrisk children. 68 This costly intervention is the only available means of protecting high-risk children against serious RSV LRI. Monthly intramuscular injections of humanized monoclonal antibody should be considered for passive immunoprophylaxis during RSV season for high-risk infants such as preterm infants less than 6 months old, children with congenital heart disease, and children less than 2 years of age with bronchopulmonary dysplasia. 45 Hospitalized infants with RSV infection should be isolated or grouped to prevent cross-infection. Hand washing; use of eye-nose goggles, gowns, and gloves; and decontamination of surfaces and fomites are additional nosocomial infection control measures. 45 

Human parainfluenza viruses (HPIVs) are the single most frequent cause of croup in infants and children worldwide and are second only to RSV as cause of LRI in infants. 45, 69 Little is known about the epidemiology of HPIVs in tropical countries, but these viruses have been detected in up to 13% of children in hospital-based ARI studies in developing countries. 5,21

HPIVs are distributed in two genera of the family Paramyxoviridae, sharing the structural and biological characteristics already mentioned in the RSV section. HPIVs are classified antigenically into types 1 to 4, and HPIV-4 has subtypes A and B. HPIV types 1 and 3 are classified in the genus Respirovirus, while HPIV types 2 and 4 are in the genus Rubulavirus. HPIV-1 and -3 are the types most frequently associated with LRI in children, the immunocompromised, the chronically ill, and the elderly, whereas PIV-4 causes mostly URI in both children and adults. 69 Binding of HPIV to sialic acid in the cell membrane is mediated by the glycoprotein HN, which contains hemagglutinin and neuraminidase activities. Fusion of viral and cell membranes is mediated by the viral F protein, which is cleaved by cellular proteolytic enzymes. 45 Once inside the cell, the cycle is similar to other Paramyxoviridae, as summarized in the RSV section. HPIVs can be propagated in primary simian or human kidney cells and in several cell lines, such as HEp-2, Vero, MDCK, LLC-MK2, BHK, and HeLa. 69 A variety of experimental animals undergo asymptomatic infection with PIV, but only higher primates develop symptoms. 45, 69 EPIDEMIOLOGY Primary HPIV infection occurs early in childhood, and by age 5 virtually all children are seropositive. 19 An estimated one third of all viral LRIs in children in the United States are caused by HPIV-1 and -3. 69, 70 In most temperate regions, HPIV-1 and -2 cause epidemics in the fall of alternate years, either in co-circulation or alternating with one another. The biennial pattern of HPIV-1 is found in both hemispheres. 69 HPIV-1 causes most croup epidemics, whereas HPIV-2 more frequently causes illness with milder manifestations, although it can also cause croup. 69 HPIV-3 occurs endemically throughout the year, with sporadic spring outbreaks mainly among infants, and HPIV-4 occurs sporadically throughout the year in children and adults. 45, 69, 70 In tropical areas HPIVs may account for up to 15% of child hospital admissions due to LRI. 21 Community-based ARI studies in children under age 5 years show higher HPIV activity during rainy seasons in tropical countries. 3, 24 HPIVs were the most frequent viruses detected in school-aged children with bronchial asthma exacerbations in urban Nigeria. 61 HPIVs spread mainly within families and closed communities, such as nurseries, day-care centers, and pediatric wards, with high secondary attack rates. In a longitudinal study conducted with children less than 2 years of age with ARI in a day-care center for low-income families in Northeast Brazil, HPIVs represented 11% of the viruses detected. 11 The virus does not persist long in the environment and is transmitted mainly by large droplets and fomites. 45 Viral shedding usually lasts 3 to 10 days, but shedding of HPIV for months has been reported in very young children and immunosuppressed hosts. 71

Primary HPIV infection may cause rhinitis, pharyngitis, laryngotracheobronchitis (croup), bronchiolitis, or pneumonia. Approximately two thirds of all PIV infections in children result in febrile URI with associated otitis media in 10% to 34%. The remaining one third of PIV infections are cases of croup, bronchiolitis, or pneumonia. 61, 63 HPIVs, mainly of types 1 and 2, cause up to 74% of all cases of croup. 69 Croup is the most striking clinical presentation of HPIV infection and is most common between the ages of 6 and 36 months. 72 Croup is manifested by inspiratory stridor, barking cough, and hoarseness caused by subglottic edema, preceded by rhinorrhea, mild cough, and low-grade fever. 45, 72 Most children recover in 2 to 5 days, but some may develop bronchiolitis and pneumonia and present with a bronchopneumonia-croup syndrome. 45, 69, 72 Since immunity to HPIVs is incomplete, infections tend to occur throughout life, but little is known about HPIV infections in adults. In general, adults have only nonspecific URI, commonly with hoarseness. 45 HPIVs can cause particularly severe diseases in immunocompromised hosts, especially children with severe combined immunodeficiency and bone marrow transplant patients. Mortality in bone marrow transplant patients with HPIV infection varies from 10% to 20% in most series. 45, 69 

HPIVs replicate in ciliated epithelial cells, causing cytolysis of the respiratory mucosa. The infection begins in the upper respiratory tract and tends to disseminate down the respiratory tree. The larynx and trachea are mostly involved in the croup syndrome, and extensive involvement of the lower respiratory tree may be present in tracheobronchitis, bronchopneumonia, and bronchiolitis. 69, 71, 72 Similar to influenza, factors determining the extent of HPIV infection include the susceptibility of the viral F protein to be cleaved and tissue-specific differences in the production of proteases to cleave it. 45 Host immunity is largely mediated by humoral immunity to the two surface proteins HN and F. Virtually all children by the age of 3 years will have seroconverted to HPIVs, generally first to HPIV-3 but later also to HPIV-1 and -2. At school age, a significant proportion of children will have seroconverted also to HPIV-4. Secretory antibody targeted to the HN glycoprotein is the best marker of protection against PIV, 71 but the protection conferred by antibodies is limited, and repeated infections will develop. T-cell immune response seems to be involved in the clearance of virus and additionally in the development of inflammatory infiltrate, edema, and excess mucus secretion, 69 and immunocompromised hosts may develop progressive and even lethal disease. 73 Like RSV, PIVs cause mononuclear interstitial infiltrate, epithelial necrosis, inflammatory exudate into the alveoli, and hyaline membrane formation in the lungs. 45 

PIV is present in respiratory secretions until about 8 days from the onset of symptoms and can be isolated in monkey kidney primary cells and several continuous cell lines. Virus can be detected in the monolayers by hemadsorption with guinea pig erythrocytes in around 3 days after inoculation and confirmed by IF. 69, 71 Shell-vial assays have been developed for HPIV detection but with mixed results. 69 IF of exfoliated respiratory epithelial cells has produced conflicting and sometimes disappointing results, with most studies reporting sensitivities between 50% and 75% at best. 69 Detection of viral RNA by RT-PCR, including commercially available multiplex assays for several respiratory viruses, has enhanced the sensitivity of detection of HPIV from clinical samples. 69, 74 Real-time PCR for respiratory viruses in multiplex format is sensitive and specific for HPIV. 35 

At present, only supportive and symptomatic treatment is available for PIV infections. Management of croup includes supplemental oxygen and racemic epinephrine nebulization in hospitalized patients. Mist therapy, although traditional, has no proven value. 72 Short-term, high-dose systemic corticosteroids may reduce the need for intubation, and nebulized budesonide has a rapid effect and is as safe and efficacious as nebulized epinephrine in moderately severe croup. 72 Several antiviral agents have in vitro activity against HPIVs, but none has reached clinical testing. 69 There have been anecdotal reports of reduced HPIV shedding in immunocompromised patients treated with ribavirin, but this finding has not resisted scrutiny. 69 Future possibilities include the BCX 2798 and BCX 2855 compounds, whose design is based on the threedimensional structure of the HN protein, which inhibit the hemagglutinin and neuraminidase activities of the protein and were effective in vitro and in an animal model against HPIV-1, -2 and -3. 75

No interventions are available for the prevention of HPIV infections. Early trials with inactivated HPIV vaccine in the 1960s were unsuccessful. 69 Recently, a live-attenuated, coldadapted HPIV-3 vaccine was found to be immunogenic for children as young as 1 month of age and holds promise for further development. 69 The same vaccine was tested in combination with a live-attenuated RSV vaccine candidate, showing that this approach is feasible and deserves further study. 76 Characterization of HPIV proteins HN and F has led to development of subunit immunogens that showed efficacy in animal models. 69 

Human rhinoviruses (HRVs) are the most frequent respiratory pathogens of humans. 77 They were the most frequently isolated viruses in children under 5 years of age with ARI in an urban slum in northeast Brazil. 3

Human rhinoviruses are small, nonenveloped, positivestrand RNA viruses in the family Picornaviridae, with over 100 identified serotypes. 77 HRV serotypes have been classified according to receptorspecificity into three groups. The major group includes 91 serotypes whose receptor is intercellular adhesion molecule-1 (ICAM-1); the minor group contains 10 serotypes whose receptor is the low-density lipoprotein receptor (LDLR); and the remaining serotype, HRV-87, utilizes a sialoprotein as cell receptor. Unlike other picornaviruses, HRVs are acid-labile, a property that distinguishes them from enteroviruses. 77 The HRV genome is a monocistronic single-stranded RNA, packed in an icosahedral capsid composed of 12 pentamers. Surrounding the fivefold vertex, each pentamer contains a 1.2-to 3.0-nm-wide canyon that contains the receptor binding site. Following receptor binding, the viral positivestrand RNA is released into the cytoplasm and directs the synthesis of a polyprotein, whose cleavage products include an RNA polymerase. This enzyme will produce an expanding pool of positive-strand RNA using as a template an intermediate negative-strand RNA. The positive-strand RNA can be either translated into virion proteins or packaged as a genome into newly assembled virions. The HRV replication cycle takes place in the cytoplasm, and mature virions are released when the host cell is lysed. 77 HRVs are resistant to ethanol, ether, chloroform, and nonionic detergent but are sensitive to UV light; to pH lower than 5 and higher than 9; and to halogens such as chlorine, bromine, iodine, and phenolic disinfectants. They are stable for days on environmental surfaces and for years at minus 70°C. 77 HRV infects only higher primates and causes illness only in humans. Several cell lines of primate origin support HRV propagation, but certain strains of HeLa cells and human embryonic fibroblasts provide higher sensitivity for HRV isolation from clinical specimens. 78 The optimal growth temperature for HRV is 33°C to 35°C. 77

HRV infections occur in people from all continents, including remotely located population groups, such as Bushmen from the Kalahari Desert, native Alaskans, and an isolated Amazon Indian tribe. 79 HRV has been estimated to cause up to 80% of all autumn colds in temperate climates. 80 In tropical countries, very few community-based studies of viral ARI have used adequate HRV detection methods, 5 and this has limited the assessment of the actual impact of HRV in those areas. However, available evidence indicates that HRV is frequently associated with ARI in children in Brazil. In Fortaleza, a city in northeast Brazil, HRV detected by isolation in cell culture represented 46% of the viruses in children under 5 years of age with ARI. 3 In Salvador, another city in the same region, HRV represented 52% of the viruses detected by RT-PCR in association with ARI in children younger than 2 years of age attending a day-care center for the underprivileged. 11 Data on the frequency of HRV among adults in tropical countries are even more scarce. In Singapore, HRV was detected in 20% of the samples obtained from adults with ARI symptoms attending primary care centers. 81 HRV transmission requires close exposure and occurs mainly by hand-to-hand contact, followed by self-inoculation into the eye or nose, but also by airborne spread. Once HRV reaches the nasal cavity, infection occurs in virtually 100% of susceptible subjects, and approximately 75% of those infected develop illness after a 1-to 2-day incubation. 77 Children play a central role in spreading the virus in the household.

Evidence suggests that indoor HRV transmission is favored by high relative humidity and crowding of young children, as occurs in the United States at the beginning of the school term, which may explain the autumn seasonal peak of HRV. 77 In tropical northeastern Brazil, however, where relative humidity remains above 70% reaching 90% during the rainy season, longitudinal studies have found no obvious HRV seasonality. 3, 11 

HRV colds are indistinguishable from colds of other viral causes and consist of nasal discharge, nasal obstruction, sneezing, sore or scratchy throat, hoarseness, cough, and headache. Facial and ear pressure may be present. Fever and malaise are uncommon. These symptoms last approximately 7 days but may persist for up to 2 weeks in 25% of cases. Infants and toddlers may display only nasal discharge and be otherwise asymptomatic. 77 The majority of patients have obstruction and mucosal abnormalities of the sinus cavities, eustachian tubes, and middle ear, which predispose to secondary bacterial sinusitis and otitis media, each complication found in approximately 2% of all colds. 82 HRV RNA may be detected by RT-PCR in maxillary sinus brushings in 40% of adults presenting with acute sinusitis, 83 and in 24% of the samples of middle ear fluid from children less than 7 years of age with diagnosis of acute otitis media. 84 HRV is frequently associated with exacerbations of chronic obstructive pulmonary disease and asthma attacks in children over 2 years of age and in adults. 59, 77, 85 

HRV replication is restricted to the respiratory epithelium, taking place in scattered ciliated cells of the nose and in nonciliated cells of the nasopharynx. 86 This tropism seems to be a consequence of receptor availability. Infection of a limited number of cells triggers the release of cytokines, chemokines, and inflammatory mediators, which together with stimulation of the local parasympathetic nerve endings, results in the cold symptoms. Kinins, prostaglandins, and proinflammatory cytokines and chemokines may contribute to vasodilation, increased vascular permeability, influx of polymorphonuclear leukocytes, exocrine gland secretion, and nerve ending stimulation, resulting in nasal obstruction, rhinorrhea, sneezing, cough, and sore throat. 77 Serotype-specific neutralizing IgM, IgG, and IgA antibodies develop in most infected persons in 7 to 21 days and persist for years. Protection from infection is partially attributed to the presence of IgA antibody in nasal secretions, and recovery from illness is more dependent on cell-mediated immunity. HRV-induced blastogenesis, natural killer cell activity, mitogen-stimulated cell production of IL-2 and IFN-γ have been documented during HRV infection. 77, 87 HRV induces the expression of human β-defensin 2 (HBD-2) in the respiratory epithelium, which supports a role for HBD-2 in host defense to HRV infection. 88

HRV can be detected in respiratory secretions by isolation in cultures of susceptible cell lines. 78 HRV shedding peaks around 48 hours after infection and declines rapidly, but may remain at low levels for up to 3 weeks. 77 Cultures should be kept at 33°C to 35°C in a roller drum and examined for 10 to 14 days. The presence of HRV, indicated by the typical CPE, is confirmed by the acid sensitivity of the isolate. Rapid immunocytochemical methods are not available because of the large number of serotypes. RT-PCR in clinical samples is more sensitive and less tedious than HRV isolation, 83 and the recently introduced real-time PCR-based assay is more sensitive than conventional RT-PCR. 89 PCR-based assays have been useful in studies to assess the impact of HRV in different settings. The homotypic nature of HRV antibodies restricts serology to experimental settings. 77

Trials of antiviral agents for HRV have been conducted, but no specific treatment suitable for routine use has yet been identified, mainly because of lack of potency, untoward side effects, and drug delivery problems. 90 Ruprintrivir, a selective inhibitor of HRV 3C protease, has potent, broad-spectrum anti-HRV activity in vitro. A double-blind, placebo-controlled clinical trial of intranasal ruprintrivir in experimental HRV infection reduced symptoms by 33% and also decreased viral titers and nasal discharge. 91 Symptomatic relief from cold symptoms can be obtained with a broad variety of nonprescription medications. Systemic sympathomimetic decongestants, such as pseudoephedrine, may reduce nasal obstruction, first-generation antihistamines may reduce sneezing and rhinorrhea, and nonsteroidal antiinflammatory drugs such as naproxen or ibuprofen may reduce headache, cough, and systemic symptoms. 77

The large number of HRV serotypes with minimal crossantigenicity has hampered the development of an HRV vaccine.

It may be possible to reduce exposure to HRV by hand washing after contact with a cold sufferer or after handling objects that may have been contaminated with respiratory secretions. 77 Studies in experimentally infected volunteers show that application of the virucidal agents salicylic acid or pyroglutamic acid to the hands reduced recovery of rhinovirus from the hand skin of treated persons as compared with controls. 92 This result suggests that rhinovirus transmission can be prevented by virucidal hand treatments.

Short-term, postexposure prophylaxis by intranasal IFN-α significantly reduced the incidence of HRV colds in household contacts of an index case. 93 However, the cost and difficulty of making the drug available to homes in a timely fashion reduce the utility of this approach for extended use by populations, especially in tropical countries.

Ruprintrivir has also been evaluated for prophylaxis of HRV colds starting 6 hours prior to inoculation of human volunteers. This approach reduced the proportion of subjects with positive viral cultures and viral titers but did not affect the frequency of colds. 91 

Respiratory infections caused by adenoviruses are among the most frequent illnesses that these viruses cause, particularly in children under age 5 years. 94 Adenoviruses have been frequently isolated in ARI studies in tropical countries. 5 In the south cone of South America, adenoviruses were the second most frequent virus recovered from children hospitalized for ARI. 95

Adenoviruses are nonenveloped, icosahedral DNA viruses of the genus Mastadenovirus in the family Adenoviridae. 94 Adenoviruses are distinguished antigenically by group-specific (A through F) and type-specific (1 through 49) antigens and by genomic subtypes identified by restriction site mapping. 93, 94 The adenovirus capsid consists of three morphologically, antigenically, and functionally distinct types of capsomers: hexons, penton bases, and fibers that project from the penton bases. The hexon and penton bases contain complementfixing, group-specific antigens common to all human adenoviruses, whereas the fibers have primarily neutralizing and hemagglutination-inhibiting, type-specific antigens. Adenoviruses are commonly accompanied by small, singlestranded DNA parvoviruses known as adenoassociated viruses, which do not seem to cause any specific disease. Most people have antibodies to at least one of the four serotypes of adenoassociated virus by age 10 years. 96 The fiber protein binds to the host cell, through the protein coxsackie and adenovirus receptor (CAR) of the immunoglobulin superfamily, which serves as a high-affinity receptor for adenoviruses. The class I major histocompatibility complex (MHC) also may serve as receptor for adenovirus 5. 94 Ligand-receptor interaction facilitates interaction of the penton base with cell surface integrins, which triggers entry. After endocytosis, the double-stranded linear genomic DNA is transported to the nucleus, where "early" and "late" sets of viral genes are transcribed, resulting in mRNAs coding for structural and nonstructural proteins. Virus assembly takes place in the nucleus, and the infectious cycle is completed by the release of up to 1 million virions upon cell lysis. 94 Adenoviruses replicate well in continuous cell lines of epithelial origin, such as HEp-2, HeLa, and A549, and can be adapted to grow in human embryonic lung fibroblasts. They are stable over a wide pH range (5 to 9), resistant to isopropyl alcohol, ether, and chloroform, stable for weeks at room temperature and for years at approximately 20°C or colder, and can be lyophilized. They are inactivated by sodium hypochlorite and a temperature of 60°C for 2 minutes. 97

Respiratory transmission of adenoviruses occurs at all ages but is of prime importance during epidemics among military recruits. Ocular transmission has been associated with swimming pools and physician offices where sterilization or hand washing has been inadequate. Asymptomatic infection and a prolonged carrier state are common. 94 Low-number adenovirus serotypes (1, 2, 3, and 5) are more frequent before age 5 years and account for 5% to 20% of cases of URI and approximately 5% of cases of LRI in children. 98 In adults, adenoviruses occur sporadically and cause mostly URI. Infections by adenoviruses types 4 and 7 are usually epidemic, with attack rates of 6% to 16% per week in newly assembled military recruits, whose adenovirus carriage rate may be as high as 18%. 94 In this group, the adenoviral syndromes vary from mild colds to severe LRI, but overall attack rates may reach 80%, with 20% to 40% of the individuals needing hospitalization. 94 In temperate climates, adenoviral infections are more frequent in late winter, spring, and early summer, whereas in northeast Brazil they seem to occur year-round. 5 In Salvador, also in northeast Brazil, adenoviruses were detected in 11% of children younger than age 2 years with ARI in a day-care center. 11 In tropical areas the incidence of adenovirus infections in military recruits is lower, and different serotypes may be involved. 98 Pharyngoconjunctival fever, commonly caused by adenoviruses types 3 and 7, may be epidemic or endemic among children during the summer in temperate climates. Inadequate chlorination or filtration of swimming pools and lakes has been associated with epidemics. 99 The incubation period of adenovirus infections averages 10 days. 98 

Adenovirus respiratory diseases may involve all parts of the respiratory tract, and up to 50% of nonepidemic infections are asymptomatic. In fact, adenoviruses were discovered because of their propensity for latency in adenoidal tissue. 94, 98 In southeast Brazil, adenoviruses were detected with equal frequency in wheezing young children and asymptomatic controls. 60 Most adenoviral illnesses consist of febrile colds, and in children the fever may be high and long-lasting. Pharyngitis is common and may be associated with fever, pharyngeal exudate, granular appearance of the mucosa, and anterior cervical adenopathy, similarly to streptococcal pharyngitis. 94 Adenoviruses can be recovered from up to 20% of cases of pharyngitis in small children. Pharyngitis may be concurrent with pharyngoconjunctival fever, a syndrome caused by adenovirus types 3 and 7 and characterized by conjunctivitis, frequently unilateral, which may last for 1 to 2 weeks, preauricular adenopathy, cough, rhinitis, malaise, and fever. 99 The most frequent complication of adenoviral colds is acute otitis media, which occurs in up to 30% of cases. 100 Adenovirus LRIs consist mainly of bronchitis and pneumonia, and may make up over 10% of childhood LRIs in temperate areas. 101 Adenoviruses may cause permanent lung parenchymal damage, especially when concurrent with measles. 101 Epidemic adenoviral infections in military recruits have a spectrum of clinical manifestation ranging from colds to severe pneumonia. Typically, however, the manifestations are fever, pharyngeal symptoms, cough, chest pain, headache, and malaise. 98 Overwhelming pneumonitis may be part of disseminated adenoviral infections in newborn infants and patients with immunodeficiencies, including acquired immunodeficiency syndrome (AIDS). However, the frequent concomitance of other respiratory pathogens in AIDS patients and the high prevalence of asymptomatic adenovirus infection shed doubt on the causal role of the adenovirus in these patients. 102 Adenoviruses are also an important cause of epidemic keratoconjunctivitis. 94, 98 While most adenoviral ARIs are self-limited and uncommonly associated with death or permanent sequelae, 94 adenoviruses alone or associated with other pathogens have been recovered from 20% of fatal cases of LRI in Argentina. 103 

Adenoviral respiratory disease results from necrosis of cells of airway epithelia, and viremia may result in disseminated infection in immunocompromised persons. Bronchiolitis, interstitial pneumonitis, and mononuclear cell infiltrates are part of the inflammatory process in the lungs. It remains unclear why certain strains are more virulent than others. For example, the genomic variant B7h was associated with the majority of fatal lower respiratory disease in South America. 95 In addition to lytic infection, adenoviruses may become latent in epithelial and lymphoid cells, which is probably important to maintaining the virus in populations. 94 A possible role of latent adenovirus in the pathogenesis of chronic airway inflammation has been suggested. 94, 96, 104 Protection from adenovirus infection and disease is mainly due to type-specific neutralizing antibody, but reinfections, mostly asymptomatic, may occur. A long-lived T-cell immune response develops in most infected immunocompetent persons and is not only responsible for recovery but also is involved in tissue pathologic changes. 94

Adenoviruses can be detected in respiratory, ocular, or ear secretions, but clinical correlation is required, because asymptomatic virus shedding is common. Isolation of adenoviruses in cell culture with identification by IF constitutes the standard diagnostic method, but direct detection of viral antigens or viral DNA by PCR in clinical samples is an attractive rapid alternative. 94 Rapid antigen detection by immunochromatography is around 95% sensitive in comparison with cell culture, and easily can be used in point-of-care diagnosis of adenovirus. However, both conventional and real-time PCR are more sensitive than cell culture is. 105 Positive results by PCR should be interpreted with caution, given the propensity of adenoviruses to cause latency.

Adenoviruses cause a characteristic CPE in a variety of cell lines of human origin, and maintenance of cultures for 2 weeks combined with blind passage (i.e., passage of cells even without obvious CPE to see if it develops after passage) may increase adenovirus recovery. 3, 94 Inoculation of cells by centrifugation followed by immunostaining may shorten the detection time. 106 Several serologic tests can detect antibodies to the common hexon antigen. 94 However, their clinical utility is restricted.

At present, there is no routine effective antiviral treatment for adenovirus infections. Successful therapy of severe adenoviral infections in immunocompromised patients with IV ribavirin has been reported. 107 Cidofovir has shown some efficacy in the rabbit ocular model of adenoviral infection. Iododeoxyuridine and adenine arabinoside were unsuccessful in the treatment of adenoviral keratoconjunctivitis. 94

A live vaccine consisting of wild-type adenovirus packaged in enteric-coated capsules induces immunity by ensuring enteric infection without infection of the respiratory tree. This approach has been used successfully to vaccinate military recruits against adenoviruses types 4 and 7. 98 Proper sterilization, hand washing, and chlorination can prevent adenovirus spread via tonometers, hands, and swimming pools.

Coronaviruses are enveloped viruses with distinct virion morphology, displaying widely spaced, long petal-shaped spikes at the surface, that confer a crownlike appearance, the origin of the name corona. The envelope contains a long helical nucleocapsid with single, positive-stranded RNA, 27 to 32 kb in size, which is the largest known viral RNA genome. 108 Until very recently, only three human coronaviruses (HCoVs) were known to exist: HCoV-229E, HCoV-OC43, and the CoV associated with severe acute respiratory syndrome (SARS-CoV). Recently, two groups in The Netherlands almost simultaneously published studies that resulted in the identification of two new strains of HCoV: HCoV-NL63 109 and HCoV-NL. 110 In addition, PCR primers directed to conserved replicase 1a sequences of animal CoVs led to the identification of yet another HCoV detected in 8.8% of children from New Haven, Connecticut with symptoms of ARI. This agent was designated HCoV-NH and is likely to represent the same species of HCoV-NL and -NL63. 111 On the basis of antigenic and genetic studies, the known human coronaviruses are distributed in three of the four coronavirus groups so far identified. HCoV-229E, -NH, -NL, and -NL63 belong to group I, HCoV-OC43 belongs to group II, and SARS-CoV is the only known constituent of group IV, while group III contains no known human viruses and consists only of the avian infectious bronchitis virus.

Coronavirus RNA synthesis occurs in the cytoplasm via a negative-strand RNA intermediate. The viral RNA possesses a 5′ cap followed by a leader sequence and an untranslated region, with another 3′ terminal untranslated region followed by a poly(A) tail. The genome is polycistronic and the synthesis of subgenomic negative-sense RNAs is done by discontinuous transcription to originate a nested set of subgenomic mRNAs that share the 5′ leader sequence and overlap at the 3′ end. The envelope contains the structural proteins S (spike), M (membrane), E (envelope), and only in the case of some group II coronaviruses, HA (hemagglutinin). The S glycoprotein contains neutralizing and T-cell epitopes and functions as the cell receptor ligand, thereby determining tissue tropism. The M protein is embedded in the envelope and interacts with the N (nucleocapsid) protein during maturation. In addition to the nucleocapsid and envelope proteins, a replicase is present in cells infected by all coronaviruses. New virions assemble by budding through intracellular membranes and are released through vesicles of the secretory pathway. 108

HCoV-229E and -OC43 are considered to be second only to rhinoviruses as agents of common colds, causing infections with variable frequency, depending mainly on the detection method and season of the study. Up to 35% of mild upper respiratory tract infections in adults have been attributed to HCoV-229E. 112 While HCoV-229E and -OC43 are documented causes of colds in temperate regions, their impact as causes of respiratory infections in tropical regions has not been defined.

Human coronaviruses were first isolated in England, almost 40 years ago, in human organ cultures of tracheal and nasal tissues. There have been relatively few field studies based on HCoV isolation in cell culture, likely because these viruses are too fastidious to be propagated, but most respiratory isolates obtained so far have been antigenically similar to either HCoV-229E or -OC43. 112 These agents have the same structural features as the other members of the family. The S protein of HCoV-229E binds to the metalloprotease human aminopeptidase N at the cell surface, and entry is independent of enzymatic activity of the receptor. The hemagglutinin of HCoV-OC43 binds to sialic acid present in glycoproteins on the cell surface and this interaction facilitates infection, but to the best of our knowledge, a specific receptor has not been identified for this agent. 108 

HCoVs have been found throughout the world and are considered to be the second most frequent cause of common cold, accounting for an average rate of 15% of respiratory illnesses in the general population in the United States. However, the rates may be quite variable from year to year, ranging from 1% to 35% in years of peak activity. HCoV infections occur mainly in the winter and spring months, but summer activity has also been documented. During the autumn peak of rhinovirus activity, 8% of the adults with a cold negative for rhinovirus were positive for HCoV by RT-PCR in Charlottesville, VA. 80 HCoV-229E has caused well-documented winter outbreaks at 2-to 4-year intervals in temperate regions. 112, 113 Similarly, winter outbreak of HCoV-OC43 has also been detected in Europe. 114 In contrast, little is known about the prevalence of HCoV-229E and -OC43 in tropical countries. In Brazil, the activity of HCoV-229E as cause of respiratory infections in nonhospitalized children was first documented by serology in the early 1970s, with a seropositivity rate of 26% in adults by complement fixation assay. 115 

The usual manifestations of HCoV infection are typical common colds. The incubation period tends to be 1 day longer that that for rhinovirus colds, with illness duration of 6 to 7 days. Low-grade fever may occur in up to 20% of the patients, and in addition to nasal symptoms, cough and sore throat occur frequently. More serious infections of the lower respiratory tract caused by HCoV have also been documented, either sporadically in infants with pneumonia and immunocompromised patients, or in up to 33% of previously healthy Marine Corps recruits with pneumonia. 112, 113 In addition, HCoV-229E and -OC43 have been recognized in association with influenza-like illnesses in frail elderly patients. Eight of 100 (8%) nasopharyngeal swabs from older patients hospitalized for cardiopulmonary illnesses during the influenza seasonal outbreak in Rochester, N.Y., were positive for HCoV (five for HCoV-229E). 116 Respiratory HCoV infections have been associated with exacerbations of asthma, chronic bronchitis, and recurrent wheezing in children. 112, 113 HCoV was detected by RT-PCR in 38 of 292 (13%) episodes of asthma in children 9 to 11 years old in England. 117 In Brazil, HCoV was detected in respiratory samples from 3 (2 OC43 and 1 229E) of 73 (4%) children younger than 2 years of age who came to the ER with wheezing. 60 Similarly to HRV, HCoV infections have been frequently recognized in association with otitis media and maxillary sinusitis in children and adults. HCoV was detected by RT-PCR in the middle ear effusion or nasopharyngeal aspirate from 16 of 92 (17%) children with acute otitis media in Finland 84 and in nasal swabs from 3 of 20 adults with acute maxillary sinusitis. 83

There is no convenient small animal model to study the pathogenesis of HCoV, and humans naturally or experimentally infected are the only source of information obtained in vivo.

HCoVs are transmitted by the respiratory route, and experimentally infected volunteers shed virus for approximately 5 days, beginning 48 hours after infection, which is approximately the time of onset of symptoms. 112, 113 The peak of symptoms occurs 2 to 4 days postinoculation. 112 HCoV-229E is known to infect airway epithelial cells from the apical surface, where the receptor is constitutively expressed, and to exit productively infected cells through the same route. 118 Ultrastructural studies of nasal epithelium of volunteers experimentally infected with HCoV-229E revealed significantly greater epithelial cell damage, ciliary loss, and cytolysis in virus-inoculated subjects than in sham-inoculated ones on day 3 postinfection. 119 In the United States, seropositivity to HCoV-OC43 and -229E rises during the first 5 years of life, and around 40% of adults are seropositive. Symptomatic reinfections are possible, despite the presence of antibodies, suggesting rapidly waning immune response or circulation of closely related but antigenically different viruses. 113, 120 Several studies indicate that respiratory HCoVs are able to reach the central nervous system. 112, 113, 120 The recently reported temporal association between HCoV-NH infection and Kawaski disease 121 awaits confirmation.

Laboratory diagnosis in clinical samples by isolation is tedious, because the two best characterized strains of HCoV are difficult to grow in routine cell cultures. Since primers can be developed for relatively constant parts of the genome, RT-PCR-based assays for HCoV-229E and -OC43 have recently become the best alternative to other methods of detection. 80 More recently, a quantitative real-time PCR-based assay for HCoVs has been developed, providing a faster means for detection and determination of viral load with potential applications in clinical studies. 122 Serologic diagnosis of HCoV by EIA is sensitive and specific and has been useful in epidemiologic surveys. 113

Intranasal interferon protects against experimental infection with HCoV-229E, 123 but no specific antiviral therapy is available, and treatment of HCoV-induced colds remains largely symptomatic. No vaccines are currently available for HCoV. 

The HCoV that fulfills Koch`s postulates as the causative agent of SARS 127 shares structural features and genome organization of the family Coronaviridae ( Fig. 59-1) . The prompt recognition of the peculiar morphology of a coronavirus in the electron microscopic studies of Vero E6 cells inoculated with oropharyngeal material from a patient was the initial finding that resulted in the identification of SARS-CoV. 128 The viral genome is 29,727 nucleotides in length, with more than 11 open reading frames coding for 23 putative proteins, some of which have unknown functions. SARS-CoV is phylogenetically different and equidistant from all previously known coronaviruses, but isolates from different origins are relatively homogeneous genetically. Genome analysis reveals that SARS-CoV is neither a host-range mutant nor a recombinant of Respiratory Tract Viral Infections ■ 651 previously known coronaviruses but rather an independently emerged virus. SARS-CoV seems to have evolved from an animal SARS-like virus, acquiring greater fitness in humans during the course of the outbreaks, probably through the appearance of nucleotide deletions in open reading frame 8. 129 It is also noteworthy that genetic signatures present in the genomes allow for differentiation of isolates obtained from different clusters. 129 The replicative cycle of SARS-CoV is thought to follow the same main steps as other coronaviruses.

SARS coronavirus (SARS-CoV) probably emerged around November 2002 in the province of Guangdong, China, where there was no serologic evidence of infection caused by this virus in sera of healthy humans sampled prior to that time. 130 At the beginning of the outbreak, many affected individuals in Guangdong were directly or indirectly involved with game trade, and indeed, palm civets and raccoon dogs from wildgame markets in the area were later found to harbor a CoV 99% homologous to SARS-CoV at the nucleotide level. This suggests that animal-to-human interspecies transmission was involved in the outbreak, providing the source of an agent that later adapted to efficient human-to-human transmission. 131, 132 Interestingly, shortly after the lifting of a wildlife trade ban that originally had been imposed to control the SARS outbreak, new cases were again detected in Guangdong, all of them caused by viruses newly introduced from animals. Since the ban was reinstalled, there have been no further naturally acquired human cases of SARS in Guangdong. 131 Remarkably, 1.8% of 938 serum samples from adults recruited in 2001 in Hong Kong tested positive for SARS-CoV antibodies, suggesting that a small proportion of healthy people from Hong Kong, as opposed to Guangdong, China, had been exposed to SARS-related viruses at least 2 years before the outbreak. 133 It is probable that SARS-CoV precursors previously crossed the species barrier and may even have caused subclinical human infection, but perhaps only occasionally this event generated strains adapted to successful human-to-human transmission. 131 SARS-CoV is mainly transmitted between humans by the deposition of infected droplets or aerosols on the respiratory epithelium. The number of confirmed secondary cases generated by one index case of SARS is relatively low, ranging from 2.2 to 3.7, suggesting relatively inefficient transmission. In addition, transmission is infrequent during the first 5 days of illness, partly because of the low viral load in respiratory secretions during that phase. For reasons not completely understood, some SARS patients, identified as superspreaders, disproportionately contribute to the generation of a high number of secondary cases. 131 Excretion of SARS-CoV in sputa and stools may average 21 and 27 days, respectively, after symptom onset, but an excretion period as prolonged as 126 days has been documented in stools. 134 Such prolonged shedding of virus in feces raises the possibility of oral-fecal transmission and, in fact, one outbreak of SARS was attributed to a faulty sewage system. 131 Case-fatality rates estimated based on cases admitted to hospital have been around 13% for patients younger than age 60 and 43% for those older than age 60 years. However, it is likely that case-fatality rates based on all infections occurring in the community would be lower. 135 Transmission of SARS-CoV among health-care workers and between patients in the hospital setting played a pivotal role in outbreak propagation. Analysis of data from initial outbreaks indicates that close contact is the most important factor leading to nosocomial transmission of this agent. Despite the lack of complete studies on the sensitivity of SARS-CoV to different environmental conditions, there have been reports of SARS-CoV persisting for up to 2 days on environmental surfaces and 4 days in diarrheal stools. 136 

The median incubation period of SARS is 4 to 6 days. Clinical symptoms and signs of SARS appear 2 to 10 days after exposure, and systemic symptoms, such as fever, chills, myalgia, and malaise, usually appear first. Respiratory symptoms appear 2 to 7 days later, represented most frequently by nonproductive cough, dyspnea, chest pain, headache, and sore throat. Diarrhea and vomiting may occur. Chest radiograms frequently reveal infiltrates consistent with viral pneumonitis, consisting mostly of consolidations and ground-glass opacifications. Computed tomography (CT) scans in patients with normal or equivocal chest radiograms may show unilobar or multilobar abnormalities. Fever generally subsides in 48 hours, but one or two relapses within 8 to 15 days are frequently observed. Lymphopenia with reduction of both CD4+ and CD8+ cells, slight decrease in platelet counts, prolonged coagulation profile, and elevated serum enzymes (lactic dehydrogenase [LDH], creatinine kinase [CK], and C-reactive protein [CRP]) are often observed. Around one third of patients may have CD4+ lymphocyte counts below 200 cells/mm 3 and higher susceptibility to secondary infections. Watery diarrhea with an average of six evacuations per day is common. 130, 131, 137 Radiologic worsening of the pulmonary lesions seen at admission, with or without appearance of new lesions, is a frequent observation, with development of diffuse groundglass changes frequently heralding the development of acute respiratory distress syndrome (ARDS). Hypoxemia is noted in approximately half of the patients at around 9 days after the onset of symptoms, and a high proportion of those admitted to the intensive care unit (ICU), especially older males, require mechanical ventilation around day 13. Development of spontaneous pneumomediastinum during follow-up is not uncommon, probably as a consequence of ruptured peripheral lung lesions into the pleural space. 130 Prognosis is related to the level of viral replication in tissues, and patients with high viral loads in serum, nasopharyngeal aspirates, or feces, as well as those in whom virus can be detected from multiple sites, tend to have poor clinical outcome. 131 In addition to old age and severe underlying diseases, CK and CRP levels have been identified as predictors of poor outcome. 138

The N-terminal portion of the spike glycoprotein is needed for virus attachment to the virus receptor, identified as the metallopeptidase angiotensin-converting enzyme homolog (ACE-2), 139 but it is unclear whether the mechanism of entry is contingent on pH-dependent endocytosis. 131 Some inconsistencies between ACE-2 and SARS-CoV tissue distribution suggest that ACE-2 may not be the only receptor, or that a coreceptor molecule may be needed for cell infection.

SARS-CoV spike protein can also bind the dendritic cell-specific C-type lectin intercellular adhesion molecule 3-grabbing nonintegrin (DC-SIGN), which does not result in dendritic cell infection by the agent but allows for SARS-CoV to be transported to susceptible target cells elsewhere. 140 SARS-CoV has been detected in studies using different combinations of immunohistochemistry, in situ hybridization, and electron microscopy in pneumocytes and on the apical surface of enterocytes. Marked inflammatory infiltrates and mucosal atrophy have not been observed in the intestine, and the pathogenesis of the SARS-CoV-related diarrhea remains largely unknown. 131 SARS-CoV viral load in the upper airways is low in the 4 initial days, with the peak at day 10 of illness. Quantitative RT-PCR for SARS-CoV in nasopharyngeal aspirates from patients who tested positive at admission revealed viral loads around 10 5 copies/mL on days 5 and 15 after clinical onset, and peak 10 7 copies/mL on day 10. 130 Higher viral loads can be detected in the lower respiratory tract than in the upper airways. Pulmonary tissue shows diffuse alveolar damage, mixed infiltrate, lung edema, hyaline membrane, abundant macrophages in alveoli and interstitium, and syncytia formation.

Besides respiratory secretions and stools, SARS-CoV can be detected in urine in up to 30% of patients, with titers averaging 10 4.4 copies/mL, in association with abnormal urinalysis results. 141 The effect of SARS-CoV infection on the immune system is highlighted by pronounced T-cell lymphopenia and elevation of several inflammatory cytokines (IL-1β, IL-6, and IL-12) and chemokines (MCP-1 and IP-10) observed in SARS patients. While MCP-1 is likely to be involved in the lung monocytic/macrophagic infiltrate, its role is not firmly established, since other viral diseases that are associated with elevated MCP-1, such as influenza, do not include such prominent histologic features. In addition, since immunologic markers in the peripheral blood may not reflect what happens in the microenvironment of the lung, the pathogenic importance of these findings is not clear. Co-inheritance of HLA-B*0703 and -B60 is higher among SARS patients than in the general population, favoring a role for the genetic background in susceptibility to SARS-CoV. 131 The pathogenesis of the T-cell lymphopenia remains unknown.

Seroconversion has been documented in 93% of the patients at around 20 days and the rise in IgG titers correlates with decrease in viral load. 130 Paradoxically, clinical worsening also occurs during this phase, suggesting that, rather than unchecked viral replication, immunopathologic factors may be responsible for the lung lesions. 130 While infection in experimental animals, such as cynomolgus macaques, ferrets, cats, golden Syrian hamsters, mice, and African green monkeys, does not induce disease that mimics that in humans, these models are important for studies of pathogenesis and development of vaccines and therapy. 129 In addition, the development of an infectious cDNA clone of SARS-CoV should permit reverse genetics experiments and may help elucidate determinants of viral pathogenesis. 142

Low viral loads in the upper respiratory tract in the first few days of illness account for the relatively poor sensitivity (35% to 65%) of first-generation RT-PCR for diagnosis in that period. SARS-CoV is detectable by RT-PCR in nasopharyngeal aspirates in only one third of patients at presentation and in two thirds at day 14. RT-PCR may be positive for SARS-CoV in stools from as much as 97% of patients at day 14, and in urine in 42% of samples at day 15. 130 Testing multiple nasopharyngeal, serum, and fecal samples increases the sensitivity of the diagnosis by RT-PCR. 143, 144 To overcome the low sensitivity of conventional RT-PCR, quantitative real-time PCR-based assays for SARS-CoV have been developed that improve sensitivity and turnaround time, allow for amplification and analysis to be done in a closed system, and thus reduce cross-contamination. In addition, the capability of the assay to quantitate viral load has contributed not only to understanding viral pathogenesis but also to predicting outcome, since high viral loads are associated with poor prognosis. 143 The ability to grow SARS-CoV in Vero E6 cell cultures was critical to identifying the agent. SARS-CoV can be recovered by isolation from respiratory secretions, feces, and urine in the first 3 weeks of illness, but the overall sensitivity is relatively low and recovery is more likely to be successful from respiratory secretions than from stools and urine. 143 Recent small outbreaks of SARS-CoV originating in laboratories 143 have heightened concern about laboratory safety issues regarding SARS specimens. The WHO guidelines for biosafety in the diagnosis of SARS (updates available at the WHO web site) recommend that propagation of SARS-CoV in cell culture for isolation or for preparation of viral stocks and cell slides be performed in biosafety level 3 (BSL3) laboratories, whereas handling serum and blood specimens for routine tests and serology can be performed in BSL2 laboratories. Nucleic acid extraction procedures, inoculation of bacterial or mycologic cultures, and preparation of sample smears can be done in BSL2 laboratories, observing BSL3 work practices (use of safety cabinets, sealed centrifuges, protective equipment, 5% bleach spillage decontamination, and proper waste disposal).

Although not useful for early diagnosis, seroconversion determined by IFA or EIA remains the gold standard for confirming SARS diagnosis. IgG seroconversion is detectable in over 90% of patients at around day 28. 130 Antibody crossreaction with other human coronaviruses, however rare, remains a possibility; therefore, confirmation of positive serology by an independent neutralization assay should be performed if available. 143 

The main component of treatment of SARS patients is supportive therapy, chiefly the management of hypoxemia and ARDS. During the 2003 outbreak, treatment included a broadspectrum antiviral agent (ribavirin) and immunosuppressive doses of corticosteroids, aimed at reducing the immunopathologic damage to the lungs. The use of high-dose steroid therapy is controversial and for the most part supported by anecdotal evidence, whereas the use of ribavirin is based on the broad antiviral spectrum of the drug. However, SARS-CoV is only modestly susceptible to ribavirin in vitro, and therapeutic doses are difficult to achieve clinically. Since it became possible to grow SARS-CoV in culture, many potential antiviral compounds have been evaluated in vitro, but just a few have been tested in animal models and even fewer are in clinical testing. 131 Interferons (IFN-αn1/n3, leukocytic IFN-α, IFN-β) and HIV protease inhibitors were consistently active in vitro and may be considered for animal testing and clinical trials. 131 The resolution of the structure of SARS-CoV principal protease has prompted studies of the inhibitory capacity of known anti-HIV protease inhibitors for treatment of SARS. In one open-label study, a combination of HIV protease inhibitor (lopinavir plus pharmacokinetic booster ritonavir) and ribavirin was used to treat SARS patients and the outcomes were compared with historical controls treated with ribavirin alone. At day 21 after onset of symptoms, development of ARDS or death was significantly less frequent in the group treated with the combination (2.3%) than in historical controls (28.8%). In addition, peak viral loads in respiratory samples and stools were reduced in the group treated with the combination as compared with controls. 145 However, since there were differences in outcome predictors, such as sex, platelet counts, and LDH levels, between the two groups, these results should be interpreted with caution.

A preliminary open-label study found that a restricted number of patients treated with subcutaneous interferon alfacon-1 in association with corticosteroids showed reduced oxygen-saturation impairment and faster resolution of radiographic chest findings than those treated with corticosteroids alone. 146 Convalescent plasma has also been tested in the treatment of SARS patients. In one preliminary uncontrolled study, convalescent plasma may have reduced the frequency of poor outcome when given before 14 days of illness. 147

It is impossible to predict whether naturally reemerging SARS-CoV would be likely to cause a global outbreak. Nevertheless, a vaccine for this agent would be relevant for high-risk individuals, such as workers in laboratories, hospitals, and game-animal farming. Therefore, considerable effort has been directed at developing such a vaccine. It has been shown that SARS-CoV spike protein produced in bacteria and expressed on chimeric parainfluenza virus, as well as spike protein-encoding DNA, induced neutralizing antibodies and protected experimental animals from challenge with live virus. At present, no SARS-CoV vaccine is available for human use. Therefore, in the absence of person-to-person transmission of SARS-CoV worldwide, prevention of future outbreaks of SARS requires careful surveillance. The goal is to maximize early detection of new cases of SARS to implement control measures, thereby minimizing social disruption. 131 To reach this goal, the CDC recommends testing for SARS-CoV in patients who require hospitalization for radiographically confirmed pneumonia or ARDS without identifiable etiology and who have one of the following risk factors in the 10 days before the onset of illness: (1) travel to mainland China, Hong Kong, or Taiwan, or close contact with an ill person with a history of recent travel to one of these areas, or (2) employment in an occupation associated with a risk for SARS-CoV exposure (e.g., health-care worker with direct patient contact; worker in a laboratory that contains live SARS-CoV), or (3) belonging to a cluster of cases of atypical pneumonia without an alternative diagnosis (updates on these recommendations are made available at the CDC web site http://www.cdc.gov/ncidod/sars).

During times of overt SARS activity, prevention of humanto-human transmission is pivotal to curtailing outbreaks. Although SARS infectiousness relative to the onset and termination of clinical symptoms has not been accurately determined, it is clear that shortening the time from onset to hospital admission and isolation reduces the risk of transmission, thus contributing substantially to curtailing of outbreaks. Identification of new cases through contact tracing played an important role in the control of the outbreaks registered so far. Stringent isolation procedures must be adopted for confirmed and suspected cases, which require a high level of alertness among health-care workers for early identification of SARS cases. The scenario may be further complicated in situations in which other diseases such as influenza and Hantavirus pulmonary infections may occur simultaneously. 135 Rates of transmission of SARS-CoV among health-care workers vary, depending on stringency of control measures adopted, presence of so-called superspreaders in the hospital, and kind of activities carried out by personnel, especially as related to proximity to the index case. Assisting during intubation, suctioning, and manipulating ventilatory apparatuses seem to be high-risk activities. While studies conducted in different settings have produced conflicting results, one study in Toronto found that up to 25% of the nurses who cared for SARS patients in critical care units became infected. 148 The presence of severe watery diarrhea may add to the challenge for the infection control team. 130 An updated set of recommendations for health-care and laboratory personnel is available at the CDC web site (http://www.cdc.gov/ncidod/sars).

A new paramyxovirus was described in The Netherlands in 2001, in association with respiratory illness in children. The agent was first detected by analysis of previously unidentifiable viral isolates that induced cytopathic effect in LLC-MK2 cell cultures. The isolates were recovered over a 10-year period in respiratory secretions from 28 children with ARI occurring in the winter time. Electron microscopy of cell culture isolates revealed paramyxovirus-like particles, and RNA sequencing revealed genome sequences and organization consistent with a paramyxovirus of the subfamily Pneumovirinae, most closely related to avian pneumovirus of the genus Metapneumovirus. Rather than an avian virus that can also infect humans, this agent is now recognized as a primarily human pathogen, and thus has been named human metapneumovirus (HMPV). 156,157 HMPV antibodies detected in sera collected in 1958 in The Netherlands indicate that this agent has been in circulation for at least 4 to 5 decades. 156 AGENT HMPV particles are enveloped, pleomorphic, spherical, and filamentous particles, with a mean diameter of about 209 nm. 156,157 Complete genome sequences of HMPV are available and, in contrast to the genomic organization of pneumoviruses, metapneumoviruses have different positioning of the genes between M and L and lack NS1 and NS2 genes. 156,158 Similar to HRSV, genetic and antigenic studies indicate that HMPV isolates cluster into two main serotype named A and B, with N gene sequences 83% to 85% similar at the nucleotide level, each subgroup including two genetic lineages (A1, A2, B1, and B2). 156, 159, 160 Both are globally distributed. There have been no detailed studies of the HMPV replication cycle, but it is likely to be similar to that of other human paramyxoviruses.

HMPV is a frequent cause of community-acquired ARI in children and adults in all continents, although with variable incidence in different settings. 157, 159, [161] [162] [163] [164] [165] [166] [167] [168] [169] In the United States, HMPV has been reported in up to 20% of lower respiratory tract illnesses whose etiology would have been unidentifiable prior to the development of assays for the detection of HMPV. 170 In Canada, during the 2001-2002 winter season, HMPV was detected in 15% of patients of all age groups from four different provinces. 167 Similar to HRSV, HMPV infections are more frequent in the colder months in temperate regions, and different strains of both subgroups A and B cocirculate during the same year. 157, 162 However, only limited knowledge is available about HMPV seasonality in more tropical climates. Peaks of HMPV activity have been documented in the spring/summer in Hong Kong, 171 while in South Africa HMPV has been detected in 6% to 9% of children with ARI admitted to hospitals in the winter season. 161 ,165 HMPV was detected alone or simultaneously with RSV in 24% of children younger than 3 years of age admitted to health-care facilities in Aracaju, northeast Brazil, in the months of April and May, 2002. 164 Interestingly, HMPV was not detected by the same methods in that same city, in the following year. 166 This apparent variability in HMPV incidence from year to year has also been observed in studies conducted in Argentina 168 and Italy, where HMPV frequencies varied from 7% to 43% in three consecutive annual respiratory virus seasons. 172 Long-term prospective studies will be needed to establish whether there is a seasonal pattern in HMPV circulation in tropical regions of the world.

Clinically, HMPV infections resemble closely those caused by HRSV, ranging from mild upper ARI to severe bronchiolitis and pneumonia. The median age of children hospitalized with HMPV infection is older than those with HRSV. HRSV in hospitalized infants and young children may require intensive care and mechanical ventilation, 156,173,174 and dual infection with HMPV and HRSV appears to increase the likelihood of severe illness. 175, 176 The most frequent symptoms in all age groups are fever, dyspnea, cough, wheezing/stridor, rhinitis, and sore throat. 162, 173 All infected children in one study had either pneumonia or bronchiolitis, frequently accompanied by otitis media. 162 HMPV may cause more serious infections in patients with comorbid or immunosuppressive conditions, as well as in the very young and the elderly. 162, 167 In one study, all individuals older than 65 with lower respiratory infection caused by HMPV had at least one underlying chronic or debilitating condition, including lymphoma, leukemia, or neurologic or cardiovascular diseases. 162 HMPV infection in adults may present as influenza-like illness, acute bronchitis, or common cold. 162 In England, in the winter of 2000-2001, HMPV was detected by RT-PCR in association with 2.2% of samples taken from patients in all age groups with influenza-like illnesses negative for HRSV and influenza viruses. 159 HMPV has been increasingly recognized as cause of acute wheezing in children. One study conducted in Finland found HMPV in 8% of wheezing children, who presented significantly higher levels of IL-8 in nasal secretions as compared to children with HRSV-associated wheezing. 174 A study conducted in Brazil found that 47% of the children with HMPV had wheezing and 31% had chest indrawing. 164 Previous history of asthma has been more frequently associated with HMPV than with HRSV infection and HMPV-infected patients are more often treated with bronchodilators and corticosteroids than HRVS-infected patients. 173

Little is known about specific mechanisms of pathogenesis and host immune response in HMPV infections. HMPV is a pathogen of both the upper and lower respiratory tracts. 162 HMPV replicates efficiently in the respiratory tract of monkeys, with virus shedding peaking between days 2 and 8 following infection. 156 Serologic data indicates that HMPV infects young individuals, and by the age of 5 virtually all children have become seropositive for the agent; reinfections at later ages are common. 156, 157, 159 Interestingly, coinfection with HMPV has been reported to correlate with increased severity of HRSV infections. A study conducted in the United Kingdom found that this coinfection caused a tenfold increase in the relative risk of admission to the ICU for mechanical ventilation in children under 2 with HRSV bronchiolitis. 173 A similar finding was also reported in Germany. 176 

HMPV can be isolated in LLC-MK2 cells from nasal aspirates or nasopharyngeal swabs. The cytopathic effect, characteristically negative on hemadsorption testing, develops usually late after inoculation (up to 23 days). 160 Sensitive RT-PCR assays for this agent have been developed in many different laboratories and have rapidly become standard for HMPV diagnosis. 159,169 A real-time PCR assay for HMPV showed to be more sensitive than conventional RT-PCR, even when hybridization was used to increase sensitivity of the detection of amplicons generated by the conventional method. 169 Using real-time PCR, HMPV was detected in 10% of 329 samples collected from patients with ARI in Australia from March to October 2001 that were negative for other pathogens. 169 

Other than supportive measures, oxygen therapy, bronchodilators, corticosteroids and mechanical ventilation, there is no specific antiviral treatment for this agent. 173 Ribavirin is inhibitory for HMPV in vitro. 176 Although a HMPV vaccine is not available at this time, the demonstration that hamsters, ferrets, and African green monkeys are susceptible to infection by HMPV, and that hamsters vaccinated with serotype A

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Detection of influenza virus by centrifugal inoculation of MDCK cells and staining with monoclonal antibodies

Rapid detection and simultaneous subtype differentiation of influenza A viruses by real time PCR

Rapid and sensitive method using multiplex real-time PCR for diagnosis of infections by influenza A and influenza B viruses, respiratory syncytial virus, and parainfluenza viruses 1, 2, 3, and 4

Rational design of potent sialidase-based inhibitors of influenza virus replication

Efficacy and safety of the neuraminidase inhibitor zanamivir in the treatment of influenzavirus infections

Resistant influenza A viruses in children treated with oseltamivir: Descriptive study

Centers for Disease Control and Prevention: Prevention and Control of Influenza: Recommendations of the Advisory Committee on Immunization Practices (ACIP)

Influenza vaccination in 2000: Recommendations and vaccine use in 50 developed and rapidly developing countries

Immunization against influenza in the elderly: The Argentinian experience

Regional perspectives on influenza surveillance in South America

Clinical and epidemiologic importance of influenza A viruses resistant to amantadine and rimantadine

Pandemic influenza: Is an antiviral response realistic?

Respiratory syncytial virus and parainfluenza viruses

Respiratory syncytial virus

Brief report: Respiratory syncytial virus activity-United States

Seasonal trends of viral respiratory tract infections in the tropics

Occurrence and severity of infections caused by subgroup A and B respiratory syncytial virus in children in southeast Brazil

Viral etiology of acute respiratory infections among children in Porto Alegre, RS, Brazil

Respiratory syncytial virus: Changes in prevalence of subgroups A and B among Argentinian children

Clinical patterns and seasonal trends in respiratory syncytial virus hospitalizations in Sao Paulo

Antigenic characterization of respiratory syncytial virus group A and B isolates in Rio de Janeiro, Brazil

Management of acute bronchiolitis

Detection of genomic sequences of respiratory syncytial virus in otitis media with effusion in children

Acute otitis media in children with bronchiolitis

Prevalence of serious bacterial infections in febrile infants with respiratory syncytial virus infection

Etiology of acute lower respiratory tract infection in children from Alabang, Metro Manilla

Rhinovirus and respiratory syncytial virus in wheezing children requiring emergency care. IgE and eosinophil analyses

Risk factors for wheezing in a subtropical environment: Role of respiratory viruses and allergen sensitization

Microbial inciters of acute asthma in urban Nigerian children

Winter viruses: Influenza-and respiratory syncytial virus-related morbidity in chronic lung disease

Pathogenesis of respiratory syncytial virus bronchiolitis-related wheezing

Pathogenesis of respiratory syncytial virus vaccine-augmented pathology

Evaluation of quantitative and typespecific real-time RT-PCR assays for detection of respiratory syncytial virus in respiratory specimens from children

American Academy of Pediatrics Committee on Infectious Diseases: Reassessment of the indications for ribavirin therapy in respiratory virus infections

Combination therapy with aerosolized ribavirin and intravenous immunoglobulin for respiratory syncytial virus disease in adult bone marrow transplant recipients

Role of antibody and use of respiratory syncytial virus (RSV) immune globulin to prevent severe RSV disease in high-risk children

Parainfluenza viruses

Parainfluenza viral infections in pediatric outpatients: Seasonal patterns and clinical characteristics

Biology of parainfluenza viruses

Viral croup: A current perspective

Parainfluenza and influenza virus infections in pediatric organ transplant recipients

Simultaneous detection of fourteen respiratory viruses in clinical specimens by two multiplex reverse transcription nested-PCR assays

Efficacy of novel hemagglutinin-neuraminidase inhibitors BCX 2798 and BCX 2855 against human parainfluenza viruses in vitro and in vivo

Evaluation of combined live, attenuated respiratory syncytial virus and parainfluenza 3 virus vaccines in infants and young children

Clinical Virology

Comparative susceptibilities of human embryonic fibroblasts and HeLa cells for isolation of human rhinoviruses

Rhinovirus antibodies in an isolated Amazon Indian tribe

Frequency and natural history of rhinovirus infections in adults during autumn

Acute respiratory symptoms in adults in general practice

Computed tomographic study of the common cold

Detection of rhinovirus in sinus brushings of patients with acute community-acquired sinusitis by reverse transcription-PCR

Detection of rhinovirus, respiratory syncytial virus, and coronavirus infections in acute otitis media by reverse transcriptase polymerase chain reaction

Viral infections in relation to age, atopy, and season of admission among children hospitalized for wheezing

Localization of human rhinovirus replication in the upper respiratory tract by in situ hybridization

Peripheral blood mononuclear cells interleukin-2 and interferon-g production, cytotoxicity, and antigen-stimulated blastogenesis during experimental rhinovirus infection

Human rhinovirus infection induces airway epithelial cell production of human beta-defensin 2 both in vitro and in vivo

Rhinovirus detection: Comparison of real-time and conventional PCR

Clinical studies of antiviral agents for picornaviral infections

Phase II, randomized, double-blind, placebo-controlled studies of ruprintrivir nasal spray 2-percent suspension for prevention and treatment of experimentally induced rhinovirus colds in healthy volunteers

Efficacy of organic acids in hand cleansers for prevention of rhinovirus infections

Prevention of natural colds by contact prophylaxis with intranasal alpha2-interferon

Fields' Virology

Molecular epidemiology of adenovirus acute lower respiratory infections of children in the south cone of South America (1991-1994)

Latent viral infections in airway epithelium

Diagnostic Procedures for Viral, Rickettsial and Chlamydial Infections

Viral Infections of Humans

Prevention: Outbreak of pharyngoconjunctival fever at a summer camp-North Carolina

Acute otitis media and respiratory virus infection

Pneumonias in childhood: Etiology and response to antimicrobial therapy

Adenovirus infections in human immunodeficiency virus-positive patients: Clinical features and molecular epidemiology

Etiologic, clinical and pathologic analysis of 31 fatal cases of acute respiratory tract infections in Argentinian children under 5 years of age

Latent adenoviral infection in the pathogenesis of chronic airway obstruction

Evaluation of a bedside immunochromatographic test for detection of adenovirus in respiratory samples, by comparison to virus isolation, PCR and real-time PCR

Rapid diagnosis of respiratory viral infections by using a shell vial assay and monoclonal antibody pool

Intravenous ribavirin therapy for disseminated adenovirus infection

Coronaviridae: The viruses and their replication

Identification of a new human coronavirus

A previously undescribed coronavirus associated with respiratory disease in humans

Evidence of a novel human coronavirus that is associated with respiratory tract disease in infants and young children

Clinical Virology

Viral Infection of Humans

An outbreak of coronavirus OC43 respiratory infection in Normandy, France

Seroepidemiologic study of coronavirus infection in Brazilian children and civilian adults

Rhinovirus and coronavirus infection-associated hospitalizations among older adults

Community study of role of viral infections in exacerbations of asthma in 9-11 year old children

Human coronavirus 229E infects polarized airway epithelia from the apical surface

The effects of coronavirus on human nasal ciliated respiratory epithelium

Fields' Virology. Philadelphia

Association between a novel human coronavirus and Kawasaki disease

Frequent detection of human coronaviruses in clinical specimens from patients with respiratory tract infections by use of a novel real-time reversetranscriptase polymerase chain reaction

Intranasal interferon as protection against experimental respiratory coronavirus infection in volunteers

Severe acute respiratory syndrome: Identification of the etiological agent

Update: Outbreak of severe acute respiratory syndrome worldwide

A novel coronavirus associated with severe acute respiratory syndrome

Comparative full-length genome sequence analysis of 14 SARS coronavirus isolates and commom mutations associated with putative origins of infection

Clinical Progression and viral load in a community outbreak of coronavirus-associated SARS pneumonia: A prospective study

Severe acute respiratory syndrome

Isolation and characterisation of viruses related to SARS coronavirus from animals in Southern China

SARS-related virus predating SARS outbreak

Long-term SARS coronavirus excretion from patient cohort

Epidemiological determinants of spread of causal agent of severe acute respiratory syndrome in Hong Kong

SARS: Responding to an unknown virus

Clinical manifestations, laboratory findings, and treatment outcomes of SARS patients

Angiotensin-converting enzyme 2 is a functional receptor for the SARS coronavirus

pH-dependent entry of severe acute respiratory syndrome coronavirus is mediated by the spike glycoprotein and enhanced by dentritic cell transfer through DC SIGN

Viral loads in clinical specimens and SARS manifestations

Reverse genetics with a full-length infectious cDNA clone of severe acute respiratory syndrome coronavirus

The aetiology, origins, and diagnosis of severe acute respiratory syndrome

Evaluation of reverse transcription-PCR assays for repid diagnosis of severe respiratory syndrome associated with a novel coronavirus

Role of lopinavir/ritonavir in the treatment of SARS: Initial virological and clinical finings

Interferon alfacon-1 plus corticosteroids in severe acute respiratory syndrome

Use of convalescent plasma therapy in SARS patients in Hong Kong

SARS among critical care nurses

Acute respiratory tract infections among a birth cohort of children from Cali, Colombia, who were studied through 17 months of age

Etiology of acute lower respiratory tract infections in children in a rural community in The Gambia

Viral agents of acute respiratory infections in young children in Kuala Lampur

Viral etiology and epidemiology of acute respiratory infections in children in Nairobi, Kenya

Diagnoses of acute lower respiratory tract infections in children in Rawalpindi and Islamabad

A study of nonbacterial agents of acute lower respiratory tract infection in Thai children

Patterns of acute respiratory tract infection in children: A longitudinal study in a depressed community in Metro Manila

Characterization of human metapneumoviruses isolated from patients in North America

Analysis of the genome sequence of a human metapneumovirus

Human metapneumovirus as a cause of community-acquired respiratory illness

Antigenic and genetic variability of human metapneumoviruses

Human metapneumovirusassociated lower respiratory tract infections among hospitalized human immunodeficiency virus type 1 (HIV-1)-infected and HIV-1-uninfected African infants

Virological features and clinical manifestations associated with human metapneumovirus: A new paramyxovirus responsible for acute respiratory infections in all age groups

Human metapneumovirus as a causative agent of acute bronchiolitis in infants

Human metapneumovirus infection in hospital referred South African children

Respiratory syncytial virus and metapneumovirus in children over two seasons with a high incidence of respiratory infections in Brazil

Human metapneumovirus infections in the Canadian population

Evidence of human metapneumovirus in children in Argentina

Molecular assays for detection of human metapneumovirus

Human metapneumovirus in lower respiratory tract disease in otherwise healthy infants and children

Children with respiratory disease associated with metapneumovirus in Hong Kong

Human metapneumovirus associated with respiratory tract infections in a 3-year study of nasal swabs from infants in Italy

Metapneumovirus and acute wheezing in children

Dual infection of infants by human metapneumovirus and human respiratory syncytial virus is strongly associated with severe bronchiolitis

Prospective study of human metapneumovirus infection in children less than 3 years of age

Identification of small animal and primate models for evaluation of vaccine candidates for human metapneumovirus (HMPV) and implications for HMPV design

were protected from challenge with either A or B HMPV serotypes opens possibilities for HMPV vaccine design. 176 Humanized neutralizing monoclonal antibody to F protein is active in experimentally infected animals. 177