key: cord-0823683-foolgmqj
authors: Gao, Hainv; Yao, Hangping; Yang, Shigui; Li, Lanjuan
title: From SARS to MERS: evidence and speculation
date: 2016-12-23
journal: Front Med
DOI: 10.1007/s11684-016-0466-7
sha: 53bc05147511693d56aa5cfe1ba38931da0a7ecb
doc_id: 823683
cord_uid: foolgmqj

The Middle East respiratory syndrome coronavirus (MERS-CoV) is a novel zoonotic pathogen. In 2012, the infectious outbreak caused by MERS-CoV in Saudi Arabia has spread to more than 1600 patients in 26 countries, resulting in over 600 deaths.Without a travel history, few clinical and radiological features can reliably differentiate MERS from SARS. But in real world, comparing with SARS, MERS presents more vaguely defined epidemiology, more severe symptoms, and higher case fatality rate. In this review, we summarize the recent findings in the field of MERS-CoV, especially its molecular virology, interspecies mechanisms, clinical features, antiviral therapies, and the further investigation into this disease. As a newly emerging virus, many questions are not fully answered, including the exact mode of transmission chain, geographical distribution, and animal origins. Furthermore, a new protocol needs to be launched to rapidly evaluate the effects of unproven antiviral drugs and vaccine to fasten the clinical application of new drugs.

The world witnessed the devastating outbreak of severe acute respiratory syndrome coronavirus (SARS-CoV) in 2003, resulting in up to 8000 cases and 700 deaths worldwide [1] . SARS-CoV is a novel zoonotic pathogen, causing typical fever and respiratory symptoms. No specific antiviral drugs and vaccines were available for SARS-CoV. Fortunately, SARS-CoV remained in the population for only 8 months and vanished without a trace. In 2012, a new interspecies coronavirus outbreak occurred in Saudi Arabia, and this infection has spread to 26 countries across the globe, infecting 1698 patients, and resulting in 609 deaths as of March 23, 2016 [2] . The virus was initially designated HCoV-EMC [3] . All individuals diagnosed with Middle East respiratory syndrome (MERS) have been linked directly or indirectly to one of four countries in the Middle East. Therefore, the virus was renamed Middle East respiratory syndrome coronavirus (MERS-CoV). Compared with SARS, the novel coronavirus emerged with a more vaguely defined epidemiology, more severe symptoms, higher case fatality rate, absence of prophylactic or therapeutic measures, and most importantly, circulation in humans with mixed features of both epidemic and sporadic nature [4] . The largest outbreak of MERS-CoV outside of Saudi Arabia occurred in South Korea with 185 confirmed cases and 36 deaths [5] , including the fourth-generation descendants of MERS cases, which emerged sporadically but later as an epidemic. In this review, we summarize the recent findings in the field of MERS-CoV, especially its molecular virology, interspecies mechanism, clinical features, and antiviral therapies, as well as our speculations. contains a 5′ cap structure along with a 3′ poly (A) tail and comprises four main structural proteins, namely, spike (S), membrane (M), envelope (E), and nucleocapsid (N) proteins; all these proteins are encoded within the 3′ end of the viral genome. The viral genome is organized in the following order: 5′UTR-ORF1a/1b-S-E-M-N-3′UTR-poly (A) tail. The 5′ cap structure and 3′ poly (A) tail play a vital role in the replication and transcription of the viral genome. The ORF1a/1b occupies 2/3 of the genome and encodes a series of non-structural proteins. Most of these proteins are involved in the formation of the viral replication/ transcription complex. The S glycoprotein mediates receptor recognition and membrane fusion and is also the focus of most immunization strategies against MERS-CoV. The S glycoprotein is cleaved into two subunits: S1 and S2. The S1 subunit contains the receptor-binding domain (RBD), which mediates viral attachment to its host receptor. When S1 binds to its receptor, the S2 subunit changes its conformation and partially inserts into the target cell, drawing viral and host cell surfaces closer to facilitate plasma membrane fusion. This conformational change requires the formation of a fusion core by two heptad repeats (HR1 and HR2) of the S2 subunit. Interference with either HR1 or HR2 blocks the viral entry. The three other structural proteins relate to virion assembly and release. MERS-CoV contains five accessory proteins, namely, 3, 4a, 4b, 5, and 8b ( Fig. 1 ) [8] , which present no homology with other coronaviruses. The accessory protein 4a exhibits a potent antagonistic activity against interferon response through both cytoplasmic and nuclear targets. The functions of the other accessory proteins have yet to be clarified.

Accumulating evidence suggests that the dromedary camel is the animal reservoir of MERS-CoV. Serologic studies have shown the presence of cross-reactive antibodies to MERS-CoV in dromedary camels in Oman, Qatar, and Mongolia [9] [10] [11] . Live MERS-CoV was isolated directly from infected camels and confirmed in camel-to-human transmission. However, the exact mode of transmission remains largely under investigation. The effects of raw meat consumption, intake of camel milk, or exposure to other infections remain unclear. To date, studies have not found any regularity in MERS-CoV transmission from the existing cases. A comparative analysis indicated that the virus isolated from camels shows more genetic variation, with few genomic variants not infectious to humans [12] ; this finding partially explained the lower infection rate in humans than in camels. Therefore, camels serve as an important reservoir for the maintenance and diversification of MERS-CoVs.

The existence of an intermediate host between camels and humans has yet to be established. During the SARS epidemic, virus transmission from bats to humans was intermediate via civet (Paguma larvata) and spread by raccoon. We speculated whether MERS-CoV was transmitted similarly, given the evidence that MERS-CoV replicated in cell lines of other animals, including goat, pig, rabbit, horse, and civet [13] [14] [15] . However, no evidence of other intermediate hosts was found mediating MERS transmission from camels to humans.

The origin of MERS-CoV infection in camel remains unclear. The transmission from bats is supported by the following evidence: (1) MERS-CoV is phylogenetically and closely related to Tylonycteris bat CoV HKU4 (Ty-BatCoV-HKU4) and Pipistrellus bat CoV HKU5 (Pi-BatCoV-HKU5), which were discovered in Tylonycteris pachypus and Pipistrellus abramus, respectively, in Hong Kong in 2006, revealing the MERS-CoV genomic origins in bats [16] ; (2) the MERS-CoV receptor is also the receptor for HKU4 but not HKU5, hence showing functional identity [17] ; and (3) viral gene fragments identical or quite similar to those of MERS-CoV have also been recovered in bats. However, an infectious virus was not isolated directly from bats [18] . If MERS-CoV originated directly from bats, the spread from the bats to humans cannot be explained. The exact mode of transmission, geographical distribution, and origins cannot also be explained. 

The initial entry of the virus into human cells is mediated by specific receptors. Scientists have identified dipeptidyl peptidase 4 (DPP4; also known as CD26) as a functional receptor for MERS-CoV, whereas SARS enters the target cells via angiotensin converting enzyme 2 (ACE2). DPP4 is a membrane-bound peptidase with a type II topology and forms homodimers on the cell surface. DPP4 contributes critically to our understanding of the pathogenesis and epidemiology of this newly emerging human coronavirus and may facilitate the development of antiviral therapies and vaccines [19] . Patients with chronic obstructive pulmonary disease and cystic fibrosis demonstrate increased DPP4 immunostaining in alveolar epithelia and alveolar macrophages. This finding suggests that a preexisting pulmonary disease increases MERS-CoV receptor abundance and predisposes individuals to MERS morbidity and mortality, which is consistent with current clinical observations [20] .

When MERS-CoV enters the body, it will impair the innate immune response of cells, including the RIG-I-like receptor signaling and MDA-5, which are associated with the expression of interferon α. Additionally, type I and II major histocompatibility (MHC) genes are affected, thereby decreasing the expression levels of the major cytokines involved in the activation of lymphocytes [21] [22] [23] .

The host-viral interaction suggests that the interspecies transmission of MERS-CoV might be associated with the following two elements for efficient infection and replication: (1) vira l adsorption capacity on the surface of human cells, particularly respiratory epithelial cells; and (2) inhibition of innate immunity of subsequent viral entry into the human cells and attenuation of the activation of adaptive immune response to take over the host metabolic apparatus and replicate efficiently.

The severity and outcome of MERS are related to gender, older age, and underlying diseases. Majority of the MERS patients are elderly males with median age of 56 years. A study involving 47 patients showed an increase in casefatality rates with age, from 39% (seven of 18) in those younger than 50 years, to 48% (13 of 27) in the group aged under 60 years, and 75% (15 of 20) in patients aged 60 years or older [24] . A study involving 70 consecutive patients found the age of 65 years associated with increased mortality (OR 4.39, 95% CI 2.13-9.05; P < 0.001) [25] . Most patients present underlying comorbid medical disorders, such as diabetes and hypertension [26] .

Patients with MERS present with symptoms of influ-enza-like illness (ILI). Clinically, MERS and SARS show few similar features, including fever, cough (predominantly dry), and even renal failure. The majority of MERS patients are reported to suffer from multiple co-morbid conditions. However, few MERS patients show no fever or cough at all and present with walking pneumonia at early stages, which can progress rapidly. A quarter to a third of all the patients shows digestive tract symptoms. More than half of the patients develop acute renal impairment at a median time of around 11 days after symptom onset, with most cases requiring renal replacement therapy [24, 27, [28] [29] [30] [31] . However, about 6.6% of patients with SARS show acute renal failure occurring at 20 days after the onset of symptoms, and only 5% need replacement therapy [24] . The possible factors contributing to the common presentation of acute renal failure in MERS patients include (1) increased number of chronic renal diseases in MERS patients, with progressive respiratory failure; and (2) direct renal injury; DPP4 is present in the renal cells, and MERS-CoV was detected in urine. Without a travel history, linking the patient to the Arabian Peninsula, or a known MERS case, few clinical and radiological features can reliably differentiate MERS from acute pneumonia caused by other microbial agents. Asymptomatic cases have been reported among female healthcare workers and children [28] . MERS-CoV incidence in children is less frequent and seems to be associated with less mortality in patients without underlying comorbidities [32] . Low mortality in children with MERS-CoV was also reported in SARS-CoV infections, in which symptoms were milder, without any mortality, and with few hospitalizations [33] .

MERS-CoV shedding was further clarified during the new epidemic in Korea, at the hospital from the first week up to 18 days to 25 days after the onset of symptoms. The viable virus can shed through respiratory secretion from clinically fully recovered patients [34] . The SARS "viral load" in upper respiratory tract secretions was low in the first 5 days of illness, and then increased progressively, peaking early in the second week [35] . These results emphasize the need for sufficient isolation based on laboratory results rather than solely on clinical symptoms in patients with coronavirus infection.

To date, no approved antiviral therapy is available for MERS-CoV, indicating a much slower response to this potential pandemic than to the avian H7N9 flu. Candidate antiviral agents are identified using three general approaches. First, broad-spectrum antiviral drugs, such as interferon, ribavirin, and cyclophilin inhibitors, which were effective in SARS patients, were determined to test the activity to MERS-CoV [36] [37] [38] . In vitro studies suggest that the type I interferon exerted an antiviral effect against several cell lines. MERS-CoV was 50 times to 100 times more sensitive to interferon α (IFN α) treatment than SARS-CoV [36, 37] . In vitro studies also showed significant antiviral effects of ribavirin and interferon α-2b combination therapy [37] . In a retrospective study, ribavirin and interferon α-2a therapy were significantly associated with improved survival at 14 days but not at 28 days [38] . The clinical significance of cyclosporine for MERS treatment is likely limited, as the drug's peak serum level at clinical dosages was below its EC50 for MERS-CoV [36] .

The second approach to identify candidate antivirals targeting MERS involves screening of chemical libraries comprising numerous existing drugs. The advantages of this approach include the commercial availability, known pharmacokinetics, and established safety reports. The first drug identified by this approach was mycophenolic acid, which is an antirejection drug used in transplantation and a broad-spectrum antiviral drug. The EC50 of this drug against MERS-CoV was very low. Therefore, a low dosage may be effective without inducing significant immunosuppression, but this assumption warrants further preclinical evaluation in animal studies [39] . The other drugs, including lopinavir and chloroquine, still remain in the in vitro stage.

The third approach to antiviral drug developments is based on the knowledge of genome and structural biology of MERS-CoV. Compared with the drugs identified by the first and second approaches, those identified by this approach were most effective against MERS-CoV. However, this novel drug development needs time and investment. The therapeutic potential of antibodies targeting coronaviruses was well recognized during the SARS outbreak [40] [41] [42] . To date, monoclonal antibodies targeting different epitopes on the RBD in the S1 subunit of the MERS-CoV S protein have been identified. These monoclonal antibodies bind to RBD with 10-fold to 450fold higher affinity than the RBD binding affinity to the human DPP4, conferring broader and higher neutralizing activity [43] [44] [45] [46] . One of the antibodies, m336, neutralizes the virus with exceptional potency and therefore represents a potential drug and vaccine candidate [47] . HR2P, a synthetic peptide derived from the HR2 domain of MERS-CoV spike protein, specifically binds to the HR1 domain of the viral spike protein and blocks viral fusion, resulting in the inhibition of MERS-CoV replication and its spike protein-mediated cell fusion [48] . Nevertheless, a major obstacle relates to the difficulty in generating highly potent neutralizing mAbs in a relatively short time during an epidemic. The other challenge relates to the emergence of possible escape mutants.

The outbreak of MERS-CoV poses a serious threat to global public health and highlights the imperative for further investigation into the viral epidemiology and pathogenesis, as well as the development of effective therapeutic and prophylactic agents against MERS-CoV infection. First, well-designed large-scale case-control studies are needed to define the transmission chain of MERS-CoV and to enable appropriate intervention by the government. Second, retrospective serum antibody detection should be continued to define its true distribution in humans and animals. Third, systematic surveillance for signs of host adaptation and viral genome variations is very important. Fourth, a randomized control study should be conducted to evaluate the effects of the available antiviral drugs. Finally, animal vaccines limit the animal-to-human transmission [49] , which will alter our approach to drug development against newly emerging infectious diseases. However, with regard to clinical trials, we should note that a traditional sequence of studies in animals followed by phased clinical trials may be very slow during public health emergencies. A common protocol should be launched to rapidly evaluate the promising but unproven therapies in the current fatal emerging infectious disease outbreaks and future epidemics [50] .

World Health Organization. Summary of probable SARS cases with onset of illness from 1

Middel East respiratory syndrome coronavirus (MERS-CoV) Saudi Arabia

Isolation of a novel coronavirus from a man with pneumonia in Saudi Arabia

Background and summary of novel coronavirus infection-as of 22

Middle East respiratory syndrome coronavirus (MERS-CoV)-Republic of Korea

Middle East respiratory syndrome

Genetic characterization of Betacoronavirus lineage C viruses in bats reveals marked sequence divergence in the spike protein of pipistrellus bat coronavirus HKU5 in Japanese pipistrelle: implications for the origin of the novel Middle East respiratory syndrome coronavirus

Coronaviruses: an overview of their replication and pathogenesis

Evidence for camel-to-human transmission of MERS coronavirus

High proportion of MERS-CoV shedding dromedaries at slaughterhouse with a potential epidemiological link to human cases

Absence of MERS-Coronavirus in Bactrian Camels, Southern Mongolia

Middle East respiratory syndrome coronavirus quasispecies that include homologues of human isolates revealed through whole-genome analysis and virus cultured from dromedary camels in Saudi Arabia

Asymptomatic Middle East respiratory syndrome coronavirus infection in rabbits

Host species restriction of Middle East respiratory syndrome coronavirus through its receptor, dipeptidyl peptidase 4

Receptor variation and susceptibility to Middle East respiratory syndrome coronavirus infection

Human betacoronavirus 2c EMC/2012-related viruses in bats, Ghana and Europe

Receptor usage and cell entry of bat coronavirus HKU4 provide insight into bat-to-human transmission of MERS coronavirus

Middle East respiratory syndrome coronavirus in bats, Saudi Arabia

Dipeptidyl peptidase 4 is a functional receptor for the emerging human coronavirus-EMC

Dipeptidyl peptidase 4 distribution in the human respiratory tract: implications for the middle east respiratory syndrome

Middle East respiratory syndrome coronavirus 4a protein is a double-stranded RNA-binding protein that suppresses PACT-induced activation of RIG-I and MDA5 in the innate antiviral response

Distinct immune response in two MERS-CoVinfected patients: can we go from bench to bedside?

Cell host response to infection with novel human coronavirus EMC predicts potential antivirals and important differences with SARS coronavirus

Epidemiological, demographic, and clinical characteristics of 47 cases of Middle East respiratory syndrome coronavirus disease from Saudi Arabia: a descriptive study

In-vitro renal epithelial cell infection reveals a viral kidney tropism as a potential mechanism for acute renal failure during Middle East respiratory syndrome (MERS) coronavirus infection

Clinical aspects and outcomes of 70 patients with Middle East respiratory syndrome coronavirus infection: a single-center experience in Saudi Arabia

Screening for Middle East respiratory syndrome coronavirus infection in hospital patients and their healthcare worker and family contacts: a prospective descriptive study

Hospital-associated outbreak of Middle East respiratory syndrome coronavirus: a serologic, epidemiologic, and clinical description

KSA MERS-CoV Investigation Team. Hospital outbreak of Middle East respiratory syndrome coronavirus

Middle East respiratory syndrome coronavirus: a case-control study of hospitalized patients

Clinical course and outcomes of critically ill patients with Middle East respiratory syndrome coronavirus infection

Middle East respiratory syndrome coronavirus in children

Severe acute respiratory syndrome coronavirus pathogenesis, disease and vaccines: an update

Environmental contamination and viral shedding in MERS patients during MERS-CoV Outbreak in South Korea

Severe acute respiratory syndrome vs. the Middle East respiratory syndrome

MERS-coronavirus replication induces severe in vitro cytopathology and is strongly inhibited by cyclosporin A or interferon-α treatment

Inhibition of novel β coronavirus replication by a combination of interferon-α2b and ribavirin

Ribavirin and interferon α-2a for severe Middle East respiratory syndrome coronavirus infection: a retrospective cohort study

Broad-spectrum antivirals for the emerging Middle East respiratory syndrome coronavirus

Potent neutralization of severe acute respiratory syndrome (SARS) coronavirus by a human mAb to S1 protein that blocks receptor association

Human monoclonal antibody as prophylaxis for SARS coronavirus infection in ferrets

An efficient method to make human monoclonal antibodies from memory B cells: potent neutralization of SARS coronavirus

A conformation-dependent neutralizing monoclonal antibody specifically targeting receptorbinding domain in Middle East respiratory syndrome coronavirus spike protein

Exceptionally potent neutralization of Middle East respiratory syndrome coronavirus by human monoclonal antibodies

Potent neutralization of MERS-CoV by human neutralizing monoclonal antibodies to the viral spike glycoprotein

Identification of human neutralizing antibodies against MERS-CoV and their role in virus adaptive evolution

Development of human neutralizing monoclonal antibodies for prevention and therapy of MERS-CoV infections

Structure-based discovery of Middle East respiratory syndrome coronavirus fusion inhibitor

Combating emerging threats-accelerating the availability of medical therapies

An orthopoxvirus-based vaccine reduces virus excretion after MERS-CoV infection in dromedary camels

This work was partially supported by the National Natural Science 

Hainv Gao, Hangping Yao, Shigui Yang, and Lanjuan Li declare that they have no conflict of interest. This manuscript is a review article and does not involve a research protocol requiring approval by the relevant institutional review board or ethics committee.