Human Coronavirus Infections Research Articles

Clinical characteristics of human coronavirus in children with acute lower respiratory tract infection

Objective To describe the clinical characteristics of acute lower respiratory tract infection (ALRTI) caused by human coronavirus (HCoV) in children. Methods Three thousand five hundred and three hospitalized children diagnosed with ALRTI in Beijing Children's Hospital from March 2007 to February 2013 were reviewed.Nasopharyngeal aspirate(NPA) specimen was collected from each patient.Reverse transcription (RT)-polymerase chain reaction(PCR) methods were applied to detect common respiratory viruses including respiratory syncytial virus (RSV), rhinovirus (RV), parainfluenza virus (PIV) type 1-4, influenza virus type A and B (IFA, IFB), adenovirus (AdV), enterovirus (EV), HCoV, human metapneumovirus (hMPV) and human bocavirus (HBoV). Serum antibodies of mycoplasma and sputum bacterial culture were also detected.Only HCoV positive patients were analyzed in this study. Results Eleven of 3 503 patients were proved as HCoV-positive in NPA specimens.Of the 11 children, 8 cases were male and 3 cases were female (2.7︰1.0). The median age was 3 months.The clinical symptoms of HCoV infection included cough (11/11 cases, 100.0%), wheezing (10/11 cases, 90.9%), fever (6/11 cases, 54.5%) and poor appetite (7/11 cases, 63.6%). Wheezing (8/11 cases, 72.7%) and moist rale in inspiratory phase (5/11 cases, 45.4%) could be heard.Most patient's chest X-ray showed bronchopneumonia. Full blood count displayed that leukocyte was in the normal range. Conclusions Respiratory tract infection with HCoV-positive will be easier to spread to ALRTI, especially in infants less than 1 year old.The symptoms include fever, cough and wheezing, but poor appetite and diarrhea can also be detected. Key words: Human coronavirus; Lower respiratory tract infection; Child

Coronaviruses and the human airway: a universal system for virus-host interaction studies.

Human coronaviruses (HCoVs) are large RNA viruses that infect the human respiratory tract. The emergence of both Severe Acute Respiratory Syndrome and Middle East Respiratory syndrome CoVs as well as the yearly circulation of four common CoVs highlights the importance of elucidating the different mechanisms employed by these viruses to evade the host immune response, determine their tropism and identify antiviral compounds. Various animal models have been established to investigate HCoV infection, including mice and non-human primates. To establish a link between the research conducted in animal models and humans, an organotypic human airway culture system, that recapitulates the human airway epithelium, has been developed. Currently, different cell culture systems are available to recapitulate the human airways, including the Air-Liquid Interface (ALI) human airway epithelium (HAE) model. Tracheobronchial HAE cultures recapitulate the primary entry point of human respiratory viruses while the alveolar model allows for elucidation of mechanisms involved in viral infection and pathogenesis in the alveoli. These organotypic human airway cultures represent a universal platform to study respiratory virus-host interaction by offering more detailed insights compared to cell lines. Additionally, the epidemic potential of this virus family highlights the need for both vaccines and antivirals. No commercial vaccine is available but various effective antivirals have been identified, some with potential for human treatment. These morphological airway cultures are also well suited for the identification of antivirals, evaluation of compound toxicity and viral inhibition.

A six-year descriptive epidemiological study of human coronavirus infections in hospitalized patients in Hong Kong.

We conducted a six-year epidemiological study on human coronaviruses (HCoVs) circulating in Hong Kong, using 8275 nasopharyngeal samples from patients with acute respiratory tract infections. HCoVs were detected in 77 (0.93%) of the samples by a pan-HCoV RT-PCR assay. The most frequently detected HCoV species was HCoV-OC43 (0.58%), followed by HCoV-229E (0.15%), HCoV-HKU1 (0.13%) and HCoV-NL63 (0.07%). HCoVs were detected throughout the study period (September 2008–August 2014), with the highest detection rate from September 2010 to August 2011 (22/1500, 1.47%). Different seasonal patterns of each HCoV species in Hong Kong were noted. HCoV-OC43 was predominant in the fall and winter, whereas HCoV-HKU1 showed peak activity in winter, with a few cases occurred in spring and summer. HCoV-229E mainly occurred in winter and spring, while HCoV-NL63 was predominant in summer and autumn. HCoVs most commonly infect the elderly and young children, with median age of 79.5 years (range, 22 days to 95 years). Intriguingly, the detection rate of HCoV-OC43 in the age group of > 80 years (26/2380, 1.09%) was significantly higher than that in the age group of 0–10 years (12/2529, 0.47%) (P < 0.05). These data provides new insight into the epidemiology of coronaviruses.

Development of a SARS Coronavirus Vaccine from Recombinant Spike Protein Plus Delta Inulin Adjuvant.

Given periodic outbreaks of fatal human infections caused by coronaviruses, development of an optimal coronavirus vaccine platform capable of rapid production is an ongoing priority. This chapter describes the use of an insect cell expression system for rapid production of a recombinant vaccine against severe acute respiratory syndrome coronavirus (SARS). Detailed methods are presented for expression, purification, and release testing of SARS recombinant spike protein antigen, followed by adjuvant formulation and animal testing. The methods herein described for rapid development of a highly protective SARS vaccine are equally suited to rapid development of vaccines against other fatal human coronavirus infections, e.g., the MERS coronavirus.

Camels, MERS-CoV, and other emerging infections in east Africa

Although human Middle-East respiratory syndrome coronavirus (MERS-CoV) infection seems to be associated with exposure to animals, including camels, identification of the animal reservoir remains challenging.1–3 We believe that this gap results from the fact that surveillance systems for diseases in both human beings and animals remain fragmented and fail to take into account the social and ecological contexts within which diseases emerge. Additionally, there is a paucity of data for emerging infectious diseases in animals, especially camels in east Africa.

Strain-Specific Clinical Characteristics of Human Coronavirus Infection Among Adolescents and Adults With Influenza-Like Illness

Strain-Specific Clinical Characteristics of Human Coronavirus Infection Among Adolescents and Adults With Influenza-Like Illness

Kinetics of Serologic Responses to MERS Coronavirus Infection in Humans, South Korea.

We investigated the kinetics of serologic responses to Middle East respiratory syndrome coronavirus (MERS-CoV) infection by using virus neutralization and MERS-CoV S1 IgG ELISA tests. In most patients, robust antibody responses developed by the third week of illness. Delayed antibody responses with the neutralization test were associated with more severe disease.

MERS-middleeastrespiratorysyndrome: a New Coronavirus Infection in Humans and Animals

MERS-middleeastrespiratorysyndrome: a New Coronavirus Infection in Humans and Animals

Detection and clinical analysis of acute lower respiratory tract infection with human coronaviruses in children in Beijing area 2007-2015

To investigate human coronaviruses (HCoVs) infection in children with acute lower respiratory tract infection(ALRTI)and to explore the clinical features of ALRTI caused by HCoVs in children. Totally 4 371 children with clinical diagnosis of ALRTI during the period from March 2007 to February 2015 seen in Beijing Children's Hospital were recruited into this study. Patients were divided into 4 groups by age, including 1 890 cases in < 1 year group, 788 cases in 1-3 years group, 553 cases in 3-6 years group, 1140 cases in ≥6 years group. One nasopharyngeal aspirate specimen was collected from each patient. RT-PCR methods were applied to detect 9 common respiratory viruses including HCoVs (including HCoV-OC43, HCoV-229E, HCoV-NL63 and HCoV-HKU1), respiratory syncytial virus (RSV) and so on. Clinical features of ALRTI with single HCoVs infection were analyzed and compared with hospitalized ALRTI cases with single RSV infection in the same period. (1) Totally 2 895 cases were positive for at least one virus in this study in 4 371 ALRTI patients (positive rate 66.23%), in which 147 cases were positive for HCoVs infection (positive rate 3.36%). (2) Positive rates of HCoVs in each year from 2007 to 2014 were 6.11%, 3.79%, 4.69%, 4.31%, 2.38% 2.10%, 0.77% and 2.65%, respectively. The mean positive rates of HCoVs for each month from January to December were 2.53%, 2.12%, 3.63%, 6.68%, 1.53%, 3.77%, 3.92%, 3.00%, 2.15%, 5.26%, 3.01% and 2.80%. (3) Detection results of each subtypes of HCoVs in total 4 371 pediatric ALRTI patients were: 48 cases positive for HCoV-OC43(1.10%), 32 cases positive for HCoV-229E(0.73%), 25 cases positive for HCoV-NL63 (0.57%), 27 cases positive for HCoV-HKU1 (0.62%). (4) Positive rates of HCoVs infection in <1 year group, 1-3 years group, 3-6 years group and ≥ 6 years group were 4.13%, 5.08%, 2.71% and 1.23%, respectively. There were significant differences in positive rates of HCoV among groups (χ² = 27.218, P<0.01). (5) There were 16 hospitalized cases with single infection of HCoVs in this study, of which 12 cases were diagnosed as bronchopneumonia, 3 cases developed acute laryngeal obstruction, 2 cases had acute bronchial asthma attack. Common clinical manifestations included cough (14 cases), gasping (13 cases), dyspnea (9 cases), fever (6 cases), hoarseness (4 cases), laryngeal stridor (4 cases) and abnormality on chest X-ray (including fuzzy lung texture, patchy shadow and consolidation) (12 cases). (6) There were no significant differences in the incidence of clinical manifestations (including cough, gasping, dyspnea, fever and abnormality on chest X-ray), complications (including respiratory failure, myocardial damage, and acute bronchial asthma attack) and mechanical ventilation between hospitalized ALRTI patients with single HCoV infection and 193 patients with single RSV infection in the same period. HCoVs are pathogens of ALRTI in children, The overall positive rate of HCoVs was 3.36% in this study. The clinical manifestations and severity of ALRTI caused by single HCoVs was comparable to that of ALRTI with single RSV infection in children.

Human Coronavirus-Associated Influenza-Like Illness in the Community Setting in Peru.

We present findings describing the epidemiology of non-severe acute respiratory syndrome human coronavirus-associated influenza-like illness from a population-based active follow-up study in four different regions of Peru. In 2010, the prevalence of infections by human coronaviruses 229E, OC43, NL63, or HKU1 was 6.4% in participants with influenza-like illness who tested negative for influenza viruses. Ten of 11 human coronavirus infections were identified in the fall-winter season. Human coronaviruses are present in different regions of Peru and are relatively frequently associated with influenza-like illness in Peru.

Molecular pathology of emerging coronavirus infections.

Respiratory viruses can cause a wide spectrum of pulmonary diseases, ranging from mild, upper respiratory tract infections to severe and life‐threatening lower respiratory tract infections, including the development of acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). Viral clearance and subsequent recovery from infection require activation of an effective host immune response; however, many immune effector cells may also cause injury to host tissues. Severe acute respiratory syndrome (SARS) coronavirus and Middle East respiratory syndrome (MERS) coronavirus cause severe infection of the lower respiratory tract, with 10% and 35% overall mortality rates, respectively; however, >50% mortality rates are seen in the aged and immunosuppressed populations. While these viruses are susceptible to interferon treatment in vitro, they both encode numerous genes that allow for successful evasion of the host immune system until after high virus titres have been achieved. In this review, we discuss the importance of the innate immune response and the development of lung pathology following human coronavirus infection. © 2014 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

O-020 Human Coronaviruses Infection In Acute Lower Respiratory Tract Infection In Children

<h3>Objectives</h3> To explore the effects of HCoVs (including HCoV-OC43, HCoV-229E, HCoV-NL63 and HCoV-HKU1) in acute lower respiratory tract infection (ALRTI) in children and to investigate the clinical features of paediatric ALRTI caused by HCoVs. <h3>Methods</h3> Total 3503 cases with ALRTI from March 2007 to March 2010 in Beijing Children’s Hospital Affiliated to Capital Medical University were enrolled into this study. One nasopharyngeal aspirate specimen was collected from each patient. PCR (or RT-PCR) were used to detect respiratory viruses including respiratory syncytial virus, human rhinovirus, influenza virus type A, B and C, parainfluenza virus type 1–4, adenovirus, enterovirus, human coronavirus, human metapneumovirus and human bocavirus. Clinical manifestation and laboratory findings of patients with HCoVs were analysed by using SPSS 19.0 for Windows (SPSS Inc., USA). <h3>Results</h3> The overall positive rate of HCoVs infection was 3.77%. Most cases with HCoVs infection were under 3 years old. The ratio between male and female were 2.3:1, and the rate of co-infection with other respiratory virus in patient infected HCoVs was 65.2%. The positive rate of HCoV-OC43 and HCoV-229E were higher than that of HCoV-NL63 and HCoV-HKU1. There were no significant differences of clinical manifestation, laboratory findings and severity between ALRTIs caused by HCoVs and RSV. <h3>Conclusions</h3> The overall infection rate of HCoVs in ALRTI in children was low. The clinical manifestations, laboratory findings and severity of ALRTI caused by HCoVs were comparable to that of ALRTI with RSV infection in children.

One severe acute respiratory syndrome coronavirus protein complex integrates processive RNA polymerase and exonuclease activities.

In addition to members causing milder human infections, the Coronaviridae family includes potentially lethal zoonotic agents causing severe acute respiratory syndrome (SARS) and the recently emerged Middle East respiratory syndrome. The ∼30-kb positive-stranded RNA genome of coronaviruses encodes a replication/transcription machinery that is unusually complex and composed of 16 nonstructural proteins (nsps). SARS-CoV nsp12, the canonical RNA-dependent RNA polymerase (RdRp), exhibits poorly processive RNA synthesis in vitro, at odds with the efficient replication of a very large RNA genome in vivo. Here, we report that SARS-CoV nsp7 and nsp8 activate and confer processivity to the RNA-synthesizing activity of nsp12. Using biochemical assays and reverse genetics, the importance of conserved nsp7 and nsp8 residues was probed. Whereas several nsp7 mutations affected virus replication to a limited extent, the replacement of two nsp8 residues (P183 and R190) essential for interaction with nsp12 and a third (K58) critical for the interaction of the polymerase complex with RNA were all lethal to the virus. Without a loss of processivity, the nsp7/nsp8/nsp12 complex can associate with nsp14, a bifunctional enzyme bearing 3'-5' exoribonuclease and RNA cap N7-guanine methyltransferase activities involved in replication fidelity and 5'-RNA capping, respectively. The identification of this tripartite polymerase complex that in turn associates with the nsp14 proofreading enzyme sheds light on how coronaviruses assemble an RNA-synthesizing machinery to replicate the largest known RNA genomes. This protein complex is a fascinating example of the functional integration of RNA polymerase, capping, and proofreading activities.

Targeting membrane-bound viral RNA synthesis reveals potent inhibition of diverse coronaviruses including the middle East respiratory syndrome virus.

Coronaviruses raise serious concerns as emerging zoonotic viruses without specific antiviral drugs available. Here we screened a collection of 16671 diverse compounds for anti-human coronavirus 229E activity and identified an inhibitor, designated K22, that specifically targets membrane-bound coronaviral RNA synthesis. K22 exerts most potent antiviral activity after virus entry during an early step of the viral life cycle. Specifically, the formation of double membrane vesicles (DMVs), a hallmark of coronavirus replication, was greatly impaired upon K22 treatment accompanied by near-complete inhibition of viral RNA synthesis. K22-resistant viruses contained substitutions in non-structural protein 6 (nsp6), a membrane-spanning integral component of the viral replication complex implicated in DMV formation, corroborating that K22 targets membrane bound viral RNA synthesis. Besides K22 resistance, the nsp6 mutants induced a reduced number of DMVs, displayed decreased specific infectivity, while RNA synthesis was not affected. Importantly, K22 inhibits a broad range of coronaviruses, including Middle East respiratory syndrome coronavirus (MERS–CoV), and efficient inhibition was achieved in primary human epithelia cultures representing the entry port of human coronavirus infection. Collectively, this study proposes an evolutionary conserved step in the life cycle of positive-stranded RNA viruses, the recruitment of cellular membranes for viral replication, as vulnerable and, most importantly, druggable target for antiviral intervention. We expect this mode of action to serve as a paradigm for the development of potent antiviral drugs to combat many animal and human virus infections.

Epidemiological and clinical features of human coronavirus in hospitalized children with respiratory infections in Chongqing

Objective To detect the existence of Middle East respiratory syndrome coronavirus (MERS-CoV) in the hospitalized children with respiratory infection in Chongqing area and study the clinical symptoms associated with the detection of 4 human coronaviruses (HCoV): HCoV-229E, HCoV-OC43, HCoV-NL63 and HCoV-HKU1 types. Methods Nasopharyngeal aspirates collected over a 4-year period (from Jul.2009 to Jun.2013) from pediatric inpatients with respiratory infection were analyzed for the presence of 5 coronavirus subtypes by using multi-nested polymerase chain reaction and sensitive real-time polymerase chain reaction assays.The clinical characteristics associated with coronavirus infection were examined. Results MERS-CoV was not found and other 4 coronaviruses were positive in 74(3.38%) of 2 188 children, in which 31(1.42%), 21(0.96%), 6(0.27%) and 15 (0.69%) specimens were positive for subtypes HCoV-229E, HCoV-NL63, HCoV-OC43, and HCoV-HKU1, respectively, and 1 case was co-detected with HCoV-229E and HCoV-NL63.HCoV infection was sporadic in the whole year, of which HCoV-229E and HCoV-NL63 infection mainly occurred in summer and autumn, while HCoV-OC43 in summer and HCoV-HKU1 in winter.Of the 74 children, 56(75.7%) were male, 36(48.6%)were at ages of 0-6 months and 19(25.7%)at 7-12 months, and 20(27.0%) had underlying diseases .The clinical symptoms of HCoV infection included cough(66/74 cases, 89.2%), sputum(53/74 cases, 71.6%), wheezing(39/74 cases, 52.7%), fever(33/74 cases, 44.6%)and diarrhea(26/74 cases, 35.1%). Most patients were diagnosed as pneumonia(54/74 cases, 73.0%), including 10 cases of severe pneumonia.Fifty patients were co-infected with another respiratory virus, in which respiratory syncytial virus was the mostly detected.HKU1 was the most likely to detect in the co-infected group.Patients who were co-infected with another respiratory virus were more likely to have cough and wheezing than sole-infected patients (P<0.05). Conclusions No MERS-CoV existed in the hospitalized children with respiratory infection in Chongqing area and HCoV-229E, HCoV-OC43, HCoV-NL63 and HCoV-HKU1 are of low prevalent.HCoV that primarily infects male infants less than 1 year old and lead to lower respiratory tract infection, especially in children with underlying diseases and plays a role in severe pneumonia disease. Key words: Middle East respiratory syndrome coronavirus; Coronavirus; Molecular epidemiology; Respiratory infection; Child

Screening of an FDA-approved compound library identifies four small-molecule inhibitors of Middle East respiratory syndrome coronavirus replication in cell culture.

Coronaviruses can cause respiratory and enteric disease in a wide variety of human and animal hosts. The 2003 outbreak of severe acute respiratory syndrome (SARS) first demonstrated the potentially lethal consequences of zoonotic coronavirus infections in humans. In 2012, a similar previously unknown coronavirus emerged, Middle East respiratory syndrome coronavirus (MERS-CoV), thus far causing over 650 laboratory-confirmed infections, with an unexplained steep rise in the number of cases being recorded over recent months. The human MERS fatality rate of ∼ 30% is alarmingly high, even though many deaths were associated with underlying medical conditions. Registered therapeutics for the treatment of coronavirus infections are not available. Moreover, the pace of drug development and registration for human use is generally incompatible with strategies to combat emerging infectious diseases. Therefore, we have screened a library of 348 FDA-approved drugs for anti-MERS-CoV activity in cell culture. If such compounds proved sufficiently potent, their efficacy might be directly assessed in MERS patients. We identified four compounds (chloroquine, chlorpromazine, loperamide, and lopinavir) inhibiting MERS-CoV replication in the low-micromolar range (50% effective concentrations [EC(50)s], 3 to 8 μM). Moreover, these compounds also inhibit the replication of SARS coronavirus and human coronavirus 229E. Although their protective activity (alone or in combination) remains to be assessed in animal models, our findings may offer a starting point for treatment of patients infected with zoonotic coronaviruses like MERS-CoV. Although they may not necessarily reduce viral replication to very low levels, a moderate viral load reduction may create a window during which to mount a protective immune response.

Possible involvement of infection with human coronavirus 229E, but not NL63, in Kawasaki disease.

Although human coronavirus (HCoV)‐NL63 was once considered a possible causative agent of Kawasaki disease based on RT‐PCR analyses, subsequent studies could not confirm the result. In this study, this possibility was explored using serological tests. To evaluate the role of HCoV infection in patients with Kawasaki disease, immunofluorescence assays and virus neutralizing tests were performed. Paired serum samples were obtained from patients with Kawasaki disease who had not been treated with γ‐globulin. HCoV‐NL63 and two antigenically different isolates of HCoV‐229E (ATCC‐VR740 and a new isolate, Sendai‐H) were examined as controls. Immunofluorescence assays detected no difference in HCoV‐NL63 antibody positivity between the patients with Kawasaki disease and controls, whereas the rate of HCoV‐229E antibody positivity was higher in the patients with Kawasaki disease than that in controls. The neutralizing tests revealed no difference in seropositivity between the acute and recovery phases of patients with Kawasaki disease for the two HCoV‐229Es. However, the Kawasaki disease specimens obtained from patients in recovery phase displayed significantly higher positivity for Sendai‐H, but not for ATCC‐VR740, as compared to the controls. The serological test supported no involvement of HCoV‐NL63 but suggested the possible involvement of HCoV‐229E in the development of Kawasaki disease. J. Med. Virol. 86:2146–2153, 2014. © 2014 Wiley Periodicals, Inc.

Severe acute respiratory syndrome-coronavirus infection in aged nonhuman primates is associated with modulated pulmonary and systemic immune responses

BackgroundMany respiratory viruses disproportionately impact the elderly. Likewise, advanced age correlated with more adverse disease outcomes following severe acute respiratory syndrome coronavirus (SARS-CoV) infection in humans. We used an aged African green monkey SARS-CoV infection model to better understand age-related mechanisms of increased susceptibility to viral respiratory infections. Nonhuman primates are critical translational models for such research given their similarities to humans in immune-ageing as well as lung structure.ResultsSignificant age- and infection-dependent differences were observed in both systemic and mucosal immune compartments. Peripheral lymphocytes, specifically CD8 T and B cells were significantly lower in aged monkeys pre- and post- SARS-CoV infection, while neutrophil and monocyte numbers were not impacted by age or infection status. Serum proinflammatory cytokines were similar in both age groups, whereas significantly lower levels of IL-1beta, IL-18, IL-6, IL-12 and IL-15 were detected in the lungs of SARS-CoV-infected aged monkeys at either 5 or 10 days post infection. Total lung leukocyte numbers and relative frequency of CD8 T cells, B cells, macrophages and dendritic cells were greatly reduced in the aged host during SARS-CoV infection, despite high levels of chemoattractants for many of these cells in the aged lung. Dendritic cells and monocytes/macrophages showed age-dependent differences in activation and chemokine receptor profiles, while the CD8 T cell and B cell responses were significantly reduced in the aged host. In examination of viral titers, significantly higher levels of SARS-CoV were detected in the nasal swabs early, at day 1 post infection, in aged as compared to juvenile monkeys, but virus levels were only slightly higher in aged animals by day 3. Although there was a trend of higher titers in respiratory tissues at day 5 post infection, this did not reach statistical significance and virus was cleared from all animals by day 10, regardless of age.ConclusionsThis study provides unique insight into how several parameters of the systemic and mucosal immune response to SARS-CoV infection are significantly modulated by age. These immune differences may contribute to deficient immune function and the observed trend of higher SARS-CoV replication in aged nonhuman primates.

Human coronavirus NL63 replication is cyclophilin A-dependent and inhibited by non-immunosuppressive cyclosporine A-derivatives including Alisporivir

Until recently, there were no effective drugs available blocking coronavirus (CoV) infection in humans and animals. We have shown before that CsA and FK506 inhibit coronavirus replication (Carbajo-Lozoya, J., Müller, M.A., Kallies, S., Thiel, V., Drosten, C., von Brunn, A. Replication of human coronaviruses SARS-CoV, HCoV-NL63 and HCoV-229E is inhibited by the drug FK506. Virus Res. 2012; Pfefferle, S., Schöpf, J., Kögl, M., Friedel, C., Müller, M.A., Stellberger, T., von Dall’Armi, E., Herzog, P., Kallies, S., Niemeyer, D., Ditt, V., Kuri, T., Züst, R., Schwarz, F., Zimmer, R., Steffen, I., Weber, F., Thiel, V., Herrler, G., Thiel, H.-J., Schwegmann-Weßels, C., Pöhlmann, S., Haas, J., Drosten, C. and von Brunn, A. The SARS-Coronavirus-host interactome: identification of cyclophilins as target for pan-Coronavirus inhibitors. PLoS Pathog., 2011). Here we demonstrate that CsD Alisporivir, NIM811 as well as novel non-immunosuppressive derivatives of CsA and FK506 strongly inhibit the growth of human coronavirus HCoV-NL63 at low micromolar, non-cytotoxic concentrations in cell culture. We show by qPCR analysis that virus replication is diminished up to four orders of magnitude to background levels. Knockdown of the cellular Cyclophilin A (CypA/PPIA) gene in Caco-2 cells prevents replication of HCoV-NL63, suggesting that CypA is required for virus replication. Collectively, our results uncover Cyclophilin A as a host target for CoV infection and provide new strategies for urgently needed therapeutic approaches.

Human Respiratory Coronaviruses Detected In Patients with Influenza-Like Illness in Arkansas, USA.

Acute respiratory viruses often result in significant morbidity and mortality. The potential impact of human respiratory coronavirus (CoV) infections was underestimated until the severe acute respiratory syndrome (SARS-CoV) outbreak in 2003, which showed that new, highly pathogenic coronaviruses could be introduced to humans, highlighting the importance of monitoring the circulating coronaviruses. The use of sensitive molecular methods has contributed to the differential diagnosis of viruses circulating in humans. Our study aim was to investigate the molecular epidemiology of human CoV strains circulating in Arkansas, their genetic variability and their association with reported influenza-like symptoms. We analyzed 200 nasal swab samples, collected by the Arkansas Department of Health in 2010, for influenza diagnosis. All samples were from patients showing acute respiratory symptoms while testing negative for influenza. Samples were pre-screened, using a quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) multiprobe for coronavirus, and subjected to confirmatory pancoronavirus and/or strain-specific reverse transcriptase (RT)-PCR followed by sequence analysis. Seventy-nine samples (39.5%) were positive by qRT-PCR and 35 samples (17.5%) were confirmed by conventional RT-PCR. Twenty-three of the confirmed samples (59%) were sequenced. The most frequent strain detected was HCoV-OC43-like followed by NL63-like; only one sample was positive for HCoV-229E and one for HCoV-HKU1. Feline-like CoV strains were detected in three samples, representing possible evidence of interspecies transmission or a new human strain. Seventeen percent of the coronavirus positive samples were also positive for other respiratory viruses, such as Respiratory Syncytial Virus (RSV), Parainfluenza 2 and 3, and Rhinovirus. Thus, HCoV-OC43, NL63, HKU1 and new feline-like strains were circulating in Arkansas in 2010. HCoV was the sole respiratory virus detected in 16% of the patients who showed acute respiratory symptoms with negative diagnoses for influenza virus.