ABSTRACT
The BA.2 sublineage of the SARS-CoV-2 Omicron variant has become dominant in most countries around the world; however, the prevalence of BA.4 and BA.5 is increasing rapidly in several regions. BA.2 is less pathogenic in animal models than previously circulating variants of concern1-4. Compared with BA.2, however, BA.4 and BA.5 possess additional substitutions in the spike protein, which play a key role in viral entry, raising concerns that the replication capacity and pathogenicity of BA.4 and BA.5 are higher than those of BA.2. Here we have evaluated the replicative ability and pathogenicity of BA.4 and BA.5 isolates in wild-type Syrian hamsters, human ACE2 (hACE2) transgenic hamsters and hACE2 transgenic mice. We have observed no obvious differences among BA.2, BA.4 and BA.5 isolates in growth ability or pathogenicity in rodent models, and less pathogenicity compared to a previously circulating Delta (B.1.617.2 lineage) isolate. In addition, in vivo competition experiments revealed that BA.5 outcompeted BA.2 in hamsters, whereas BA.4 and BA.2 exhibited similar fitness. These findings suggest that BA.4 and BA.5 clinical isolates have similar pathogenicity to BA.2 in rodents and that BA.5 possesses viral fitness superior to that of BA.2.
Subject(s)
COVID-19 , Genetic Fitness , Rodentia , SARS-CoV-2 , Animals , Cricetinae , Humans , Mice , COVID-19/virology , Mesocricetus/virology , Mice, Transgenic , Rodentia/virology , SARS-CoV-2/classification , SARS-CoV-2/genetics , SARS-CoV-2/pathogenicity , SARS-CoV-2/physiology , Animals, Genetically Modified , Genetic Fitness/genetics , Genetic Fitness/physiology , VirulenceABSTRACT
The recent emergence of SARS-CoV-2 Omicron (B.1.1.529 lineage) variants possessing numerous mutations has raised concerns of decreased effectiveness of current vaccines, therapeutic monoclonal antibodies and antiviral drugs for COVID-19 against these variants1,2. The original Omicron lineage, BA.1, prevailed in many countries, but more recently, BA.2 has become dominant in at least 68 countries3. Here we evaluated the replicative ability and pathogenicity of authentic infectious BA.2 isolates in immunocompetent and human ACE2-expressing mice and hamsters. In contrast to recent data with chimeric, recombinant SARS-CoV-2 strains expressing the spike proteins of BA.1 and BA.2 on an ancestral WK-521 backbone4, we observed similar infectivity and pathogenicity in mice and hamsters for BA.2 and BA.1, and less pathogenicity compared with early SARS-CoV-2 strains. We also observed a marked and significant reduction in the neutralizing activity of plasma from individuals who had recovered from COVID-19 and vaccine recipients against BA.2 compared to ancestral and Delta variant strains. In addition, we found that some therapeutic monoclonal antibodies (REGN10987 plus REGN10933, COV2-2196 plus COV2-2130, and S309) and antiviral drugs (molnupiravir, nirmatrelvir and S-217622) can restrict viral infection in the respiratory organs of BA.2-infected hamsters. These findings suggest that the replication and pathogenicity of BA.2 is similar to that of BA.1 in rodents and that several therapeutic monoclonal antibodies and antiviral compounds are effective against Omicron BA.2 variants.
Subject(s)
Antiviral Agents , COVID-19 Drug Treatment , SARS-CoV-2 , Animals , Antibodies, Monoclonal/pharmacology , Antibodies, Monoclonal/therapeutic use , Antibodies, Monoclonal, Humanized , Antibodies, Neutralizing/pharmacology , Antibodies, Neutralizing/therapeutic use , Antibodies, Viral/pharmacology , Antibodies, Viral/therapeutic use , Antiviral Agents/pharmacology , Antiviral Agents/therapeutic use , COVID-19/genetics , COVID-19/immunology , COVID-19/virology , Cricetinae , Cytidine/analogs & derivatives , Drug Combinations , Hydroxylamines , Indazoles , Lactams , Leucine , Mice , Nitriles , Proline , SARS-CoV-2/drug effects , SARS-CoV-2/genetics , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/genetics , Triazines , TriazolesABSTRACT
The recent emergence of B.1.1.529, the Omicron variant1,2, has raised concerns of escape from protection by vaccines and therapeutic antibodies. A key test for potential countermeasures against B.1.1.529 is their activity in preclinical rodent models of respiratory tract disease. Here, using the collaborative network of the SARS-CoV-2 Assessment of Viral Evolution (SAVE) programme of the National Institute of Allergy and Infectious Diseases (NIAID), we evaluated the ability of several B.1.1.529 isolates to cause infection and disease in immunocompetent and human ACE2 (hACE2)-expressing mice and hamsters. Despite modelling data indicating that B.1.1.529 spike can bind more avidly to mouse ACE2 (refs. 3,4), we observed less infection by B.1.1.529 in 129, C57BL/6, BALB/c and K18-hACE2 transgenic mice than by previous SARS-CoV-2 variants, with limited weight loss and lower viral burden in the upper and lower respiratory tracts. In wild-type and hACE2 transgenic hamsters, lung infection, clinical disease and pathology with B.1.1.529 were also milder than with historical isolates or other SARS-CoV-2 variants of concern. Overall, experiments from the SAVE/NIAID network with several B.1.1.529 isolates demonstrate attenuated lung disease in rodents, which parallels preliminary human clinical data.
Subject(s)
COVID-19/pathology , COVID-19/virology , Disease Models, Animal , SARS-CoV-2/pathogenicity , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Animals , Cricetinae , Female , Humans , Lung/pathology , Lung/virology , Male , Mesocricetus , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Transgenic , Viral LoadABSTRACT
The spike (S) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) plays a key role in viral infectivity. It is also the major antigen stimulating the host's protective immune response, specifically, the production of neutralizing antibodies. Recently, a new variant of SARS-CoV-2 possessing multiple mutations in the S protein, designated P.1, emerged in Brazil. Here, we characterized a P.1 variant isolated in Japan by using Syrian hamsters, a well-established small animal model for the study of SARS-CoV-2 disease (COVID-19). In hamsters, the variant showed replicative abilities and pathogenicity similar to those of early and contemporary strains (i.e., SARS-CoV-2 bearing aspartic acid [D] or glycine [G] at position 614 of the S protein). Sera and/or plasma from convalescent patients and BNT162b2 messenger RNA vaccinees showed comparable neutralization titers across the P.1 variant, S-614D, and S-614G strains. In contrast, the S-614D and S-614G strains were less well recognized than the P.1 variant by serum from a P.1-infected patient. Prior infection with S-614D or S-614G strains efficiently prevented the replication of the P.1 variant in the lower respiratory tract of hamsters upon reinfection. In addition, passive transfer of neutralizing antibodies to hamsters infected with the P.1 variant or the S-614G strain led to reduced virus replication in the lower respiratory tract. However, the effect was less pronounced against the P.1 variant than the S-614G strain. These findings suggest that the P.1 variant may be somewhat antigenically different from the early and contemporary strains of SARS-CoV-2.
Subject(s)
COVID-19/virology , SARS-CoV-2/physiology , SARS-CoV-2/pathogenicity , Virus Replication , Animals , Antibodies, Neutralizing , COVID-19/diagnostic imaging , COVID-19/pathology , Cricetinae , Humans , Immunogenicity, Vaccine , Lung/pathology , Mesocricetus , Mice , Spike Glycoprotein, Coronavirus/genetics , X-Ray MicrotomographyABSTRACT
Diagnosis by rapid antigen tests (RATs) is useful for early initiation of antiviral treatment. Because RATs are easy to use, they can be adapted for self-testing. Several kinds of RATs approved for such use by the Japanese regulatory authority are available from drug stores and websites. Most RATs for COVID-19 are based on antibody detection of the SARS-CoV-2 N protein. Since Omicron and its subvariants have accumulated several amino acid substitutions in the N protein, such amino acid changes might affect the sensitivity of RATs. Here, we investigated the sensitivity of seven RATs available in Japan, six of which are approved for public use and one of which is approved for clinical use, for the detection of BA.5, BA.2.75, BF.7, XBB.1, and BQ.1.1, as well as the delta variant (B.1.627.2). All tested RATs detected the delta variant with a detection level between 7500 and 75 000 pfu per test, and all tested RATs showed similar sensitivity to the Omicron variant and its subvariants (BA.5, BA.2.75, BF.7, XBB.1, and BQ.1.1). Human saliva did not reduce the sensitivity of the RATs tested. Espline SARS-CoV-2 N showed the highest sensitivity followed by Inspecter KOWA SARS-CoV-2 and V Trust SARS-CoV-2 Ag. Since the RATs failed to detect low levels of infectious virus, individuals whose specimens contained less infectious virus than the detection limit would be considered negative. Therefore, it is important to note that RATs may miss individuals shedding low levels of infectious virus.
Subject(s)
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/genetics , COVID-19/diagnosis , Amino Acid Substitution , Antiviral AgentsABSTRACT
At the end of 2019, a novel coronavirus (severe acute respiratory syndrome coronavirus 2; SARS-CoV-2) was detected in Wuhan, China, that spread rapidly around the world, with severe consequences for human health and the global economy. Here, we assessed the replicative ability and pathogenesis of SARS-CoV-2 isolates in Syrian hamsters. SARS-CoV-2 isolates replicated efficiently in the lungs of hamsters, causing severe pathological lung lesions following intranasal infection. In addition, microcomputed tomographic imaging revealed severe lung injury that shared characteristics with SARS-CoV-2-infected human lung, including severe, bilateral, peripherally distributed, multilobular ground glass opacity, and regions of lung consolidation. SARS-CoV-2-infected hamsters mounted neutralizing antibody responses and were protected against subsequent rechallenge with SARS-CoV-2. Moreover, passive transfer of convalescent serum to naïve hamsters efficiently suppressed the replication of the virus in the lungs even when the serum was administrated 2 d postinfection of the serum-treated hamsters. Collectively, these findings demonstrate that this Syrian hamster model will be useful for understanding SARS-CoV-2 pathogenesis and testing vaccines and antiviral drugs.
Subject(s)
Coronavirus Infections/virology , Disease Models, Animal , Lung/pathology , Pneumonia, Viral/virology , Animals , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , Betacoronavirus/pathogenicity , Betacoronavirus/physiology , COVID-19 , Cell Line , Chlorocebus aethiops , Coronavirus Infections/pathology , Coronavirus Infections/therapy , Cricetinae , Humans , Immunization, Passive , Lung/diagnostic imaging , Lung/virology , Mesocricetus , Pandemics , Pneumonia, Viral/pathology , Ribonucleoproteins/chemistry , SARS-CoV-2 , Vero Cells , Viral Proteins/chemistry , Virus Replication , COVID-19 SerotherapyABSTRACT
On influenza virus infection or vaccination, immune responses occur, including the production of antibodies with various functions that contribute to protection from seasonal influenza virus infection. In the current study, we attempted to identify the antibody functions that play a central role in preventing the onset of seasonal influenza by comparing the levels of several antibody titers for different antibody functions between 5 subclinically infected individuals and 16 patients infected with seasonal H3N2 virus. For antibody titers before influenza virus exposure, we found that the nAb titers and enzyme-linked immunosorbent assay titers against hemagglutinin and neuraminidase (NA) proteins in the subclinically infected individuals were significantly higher than those in the patients, whereas the NA inhibition titers and antibody-dependent cellular cytotoxicity activities did not significantly differ between subclinically infected individuals and infected patients. These results suggest that nAb and enzyme-linked immunosorbent assay titers against hemagglutinin and NA serve as correlates of symptomatic influenza infection.
Subject(s)
Influenza A Virus, H1N1 Subtype , Influenza Vaccines , Influenza, Human , Humans , Influenza A Virus, H3N2 Subtype , Hemagglutinins , Seasons , Antibodies, Viral , Neuraminidase , Hemagglutinin Glycoproteins, Influenza VirusABSTRACT
Surveillance of bat betacoronaviruses is crucial for understanding their spillover potential. We isolated bat sarbecoviruses from Rhinolophus cornutus bats in multiple locations in Japan. These viruses grew efficiently in cells expressing R. cornutus angiotensin converting enzyme-2, but not in cells expressing human angiotensin converting enzyme-2, suggesting a narrow host range.
Subject(s)
Chiroptera , Animals , Humans , Peptidyl-Dipeptidase A , Japan/epidemiology , Betacoronavirus , Host SpecificityABSTRACT
INTRODUCTION: Rapid antigen detection (RAD) tests are convenient tools for detecting the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in clinics, and testing using saliva samples could decrease the risk of infection during sample collection. This study aimed to assess the accuracy of the SARS-CoV-2 RAD for testing of nasopharyngeal swab specimens and saliva samples in comparison with the RT-PCR tests and viral culture for detecting viable virus. METHODS: One hundred seventeen nasopharyngeal swab specimens and 73 saliva samples with positive results on RT-PCR were used. Residual samples were assayed using a commercially available RAD test immediately, and its positivity was determined at various time points during the clinical course. The concordance between 54 nasopharyngeal swab samples and saliva samples that were collected simultaneously was determined. Viral culture was performed on 117 samples and compared with the results of the RAD test. RESULTS: The positive rate of RAD test using saliva samples was low throughout the clinical course. Poor concordance was observed between nasopharyngeal swab specimens and saliva samples (75.9%, kappa coefficient 0.310). However, a substantially high concordance between the RAD test and viral culture was observed in both nasopharyngeal swab specimens (86.8%, kappa coefficient 0.680) and saliva samples (95.1%, kappa coefficient 0.643). CONCLUSIONS: The sensitivity of the SARS-CoV-2 RAD test was insufficient, particularly for saliva samples. However, a substantially high concordance with viral culture suggests its potential utility as an auxiliary test for estimating SARS-CoV-2 viability.
Subject(s)
COVID-19 , SARS-CoV-2 , Humans , Nasopharynx , Reverse Transcriptase Polymerase Chain Reaction , SalivaSubject(s)
Antibodies, Neutralizing , Antiviral Agents , COVID-19 Drug Treatment , COVID-19 , SARS-CoV-2 , Antibodies, Neutralizing/drug effects , Antibodies, Neutralizing/immunology , Antibodies, Viral/drug effects , Antibodies, Viral/immunology , Antiviral Agents/immunology , Antiviral Agents/therapeutic use , COVID-19/genetics , COVID-19/immunology , Humans , SARS-CoV-2/drug effects , SARS-CoV-2/genetics , SARS-CoV-2/immunologySubject(s)
Antibodies, Monoclonal , Antibodies, Neutralizing , Antiviral Agents , COVID-19 Drug Treatment , COVID-19 , Immunologic Factors , SARS-CoV-2 , Antibodies, Monoclonal/therapeutic use , Antibodies, Neutralizing/therapeutic use , Antibodies, Viral/immunology , Antiviral Agents/therapeutic use , COVID-19/immunology , Humans , Immunologic Factors/therapeutic use , SARS-CoV-2/drug effects , Vaccine EfficacySubject(s)
Antibodies, Viral , Antiviral Agents , COVID-19 , SARS-CoV-2 , Vaccine Efficacy , Humans , Antibodies, Viral/immunology , Antiviral Agents/immunology , Antiviral Agents/pharmacology , In Vitro Techniques , SARS-CoV-2/drug effects , SARS-CoV-2/immunology , COVID-19/genetics , COVID-19/immunology , COVID-19/virologyABSTRACT
Long-term culture of the human lung adenocarcinoma cell line A549 promotes the differentiation of these cells toward an alveolar type II cell phenotype. Here, we evaluated the susceptibility of long-term cultured A549 cells to human influenza viruses. A549 cells were cultured continuously for 25 days (D25-A549) or 1 day (D1-A549) in Ham's F12K medium. Six human influenza A viruses grew much faster in D25-A549 cells than in D1-A549 cells; however, two influenza B viruses replicated poorly in both cell types. Two avian influenza viruses replicated efficiently in both cell types, with similar titres. Expression levels of human virus receptors were higher in D25-A549 cells than in D1-A549 cells. D25-A549 cells thus more efficiently support the replication of human influenza A viruses compared with D1-A549 cells. Our data suggest that long-term cultured A549 cells will be useful for influenza A virus research.
Subject(s)
Influenza A virus/growth & development , Influenza B virus/growth & development , Virus Replication , A549 Cells , Culture Media/chemistry , Culture Media/metabolism , Humans , Influenza A virus/physiology , Influenza B virus/physiology , Influenza, Human/virology , Time Factors , Virus CultivationABSTRACT
Ferrets and mice are frequently used as animal models for influenza research. However, ferrets are demanding in terms of housing space and handling, whereas mice are not naturally susceptible to infection with human influenza A or B viruses. Therefore, prior adaptation of human viruses is required for their use in mice. In addition, there are no mouse-adapted variants of the recent H3N2 viruses, because these viruses do not replicate well in mice. In this study, we investigated the susceptibility of Syrian hamsters to influenza viruses with a view to using the hamster model as an alternative to the mouse model. We found that hamsters are sensitive to influenza viruses, including the recent H3N2 viruses, without adaptation. Although the hamsters did not show weight loss or clinical signs of H3N2 virus infection, we observed pathogenic effects in the respiratory tracts of the infected animals. All of the H3N2 viruses tested replicated in the respiratory organs of the hamsters, and some of them were detected in the nasal washes of infected animals. Moreover, a 2009 pandemic (pdm09) virus and a seasonal H1N1 virus, as well as one of the two H3N2 viruses, but not a type B virus, were transmissible by the airborne route in these hamsters. Hamsters thus have the potential to be a small-animal model for the study of influenza virus infection, including studies of the pathogenicity of H3N2 viruses and other strains, as well as for use in H1N1 virus transmission studies.IMPORTANCE We found that Syrian hamsters are susceptible to human influenza viruses, including the recent H3N2 viruses, without adaptation. We also found that a pdm09 virus and a seasonal H1N1 virus, as well as one of the H3N2 viruses, but not a type B virus tested, are transmitted by the airborne route in these hamsters. Syrian hamsters thus have the potential to be used as a small-animal model for the study of human influenza viruses.
Subject(s)
Disease Models, Animal , Epithelium/pathology , Orthomyxoviridae Infections/pathology , Respiratory System/pathology , Animals , Antibodies, Viral/blood , Dogs , Female , Humans , Influenza A Virus, H1N1 Subtype , Influenza A Virus, H3N2 Subtype , Madin Darby Canine Kidney Cells , Mesocricetus/virology , Mice , Mice, Inbred BALB C , Mice, Inbred DBA , Respiratory System/virologyABSTRACT
Avian influenza A viruses rarely infect humans; however, when human infection and subsequent human-to-human transmission occurs, worldwide outbreaks (pandemics) can result. The recent sporadic infections of humans in China with a previously unrecognized avian influenza A virus of the H7N9 subtype (A(H7N9)) have caused concern owing to the appreciable case fatality rate associated with these infections (more than 25%), potential instances of human-to-human transmission, and the lack of pre-existing immunity among humans to viruses of this subtype. Here we characterize two early human A(H7N9) isolates, A/Anhui/1/2013 (H7N9) and A/Shanghai/1/2013 (H7N9); hereafter referred to as Anhui/1 and Shanghai/1, respectively. In mice, Anhui/1 and Shanghai/1 were more pathogenic than a control avian H7N9 virus (A/duck/Gunma/466/2011 (H7N9); Dk/GM466) and a representative pandemic 2009 H1N1 virus (A/California/4/2009 (H1N1pdm09); CA04). Anhui/1, Shanghai/1 and Dk/GM466 replicated well in the nasal turbinates of ferrets. In nonhuman primates, Anhui/1 and Dk/GM466 replicated efficiently in the upper and lower respiratory tracts, whereas the replicative ability of conventional human influenza viruses is typically restricted to the upper respiratory tract of infected primates. By contrast, Anhui/1 did not replicate well in miniature pigs after intranasal inoculation. Critically, Anhui/1 transmitted through respiratory droplets in one of three pairs of ferrets. Glycan arrays showed that Anhui/1, Shanghai/1 and A/Hangzhou/1/2013 (H7N9) (a third human A(H7N9) virus tested in this assay) bind to human virus-type receptors, a property that may be critical for virus transmissibility in ferrets. Anhui/1 was found to be less sensitive in mice to neuraminidase inhibitors than a pandemic H1N1 2009 virus, although both viruses were equally susceptible to an experimental antiviral polymerase inhibitor. The robust replicative ability in mice, ferrets and nonhuman primates and the limited transmissibility in ferrets of Anhui/1 suggest that A(H7N9) viruses have pandemic potential.
Subject(s)
Influenza A virus , Influenza, Human/virology , Orthomyxoviridae Infections/virology , Virus Replication , Animals , Antiviral Agents/pharmacology , Cells, Cultured , Chickens/virology , DNA-Directed RNA Polymerases/antagonists & inhibitors , Dogs , Enzyme Inhibitors/pharmacology , Female , Ferrets/virology , Humans , Influenza A Virus, H1N1 Subtype/drug effects , Influenza A Virus, H1N1 Subtype/enzymology , Influenza A virus/chemistry , Influenza A virus/drug effects , Influenza A virus/isolation & purification , Influenza A virus/pathogenicity , Influenza, Human/drug therapy , Macaca fascicularis/virology , Madin Darby Canine Kidney Cells , Male , Mice , Mice, Inbred BALB C , Models, Molecular , Monkey Diseases/pathology , Monkey Diseases/virology , Neuraminidase/antagonists & inhibitors , Orthomyxoviridae Infections/pathology , Orthomyxoviridae Infections/transmission , Quail/virology , Swine/virology , Swine, Miniature/virology , Virus Replication/drug effectsABSTRACT
Influenza viruses exist in each host as a collection of genetically diverse variants, which might enhance their adaptive potential. To assess the genetic and functional diversity of highly pathogenic avian influenza A(H5N1) viruses within infected humans, we used deep-sequencing methods to characterize samples obtained from infected patients in northern Vietnam during 2004-2010 on different days after infection, from different anatomic sites, or both. We detected changes in virus genes that affected receptor binding, polymerase activity, or interferon antagonism, suggesting that these factors could play roles in influenza virus adaptation to humans. However, the frequency of most of these mutations remained low in the samples tested, implying that they were not efficiently selected within these hosts. Our data suggest that adaptation of influenza A(H5N1) viruses is probably stepwise and depends on accumulating combinations of mutations that alter function while maintaining fitness.