Structural basis and analysis of hamster ACE2 binding to different SARS-CoV-2 spike RBDs.

Pet golden hamsters were first identified being infected with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) delta variant of concern (VOC) and transmitted the virus back to humans in Hong Kong in January 2022. Here, we studied the binding of two hamster (golden hamster and Chinese hamster) angiotensin-converting enzyme 2 (ACE2) proteins to the spike protein receptor-binding domains (RBDs) of SARS-CoV-2 prototype and eight variants, including alpha, beta, gamma, delta, and four omicron sub-variants (BA.1, BA.2, BA.3, and BA.4/BA.5). We found that the two hamster ACE2s present slightly lower affinity for the RBDs of all nine SARS-CoV-2 viruses tested than human ACE2 (hACE2). Furthermore, the similar infectivity to host cells expressing hamster ACE2s and hACE2 was confirmed with the nine pseudotyped SARS-CoV-2 viruses. Additionally, we determined two cryo-electron microscopy (EM) complex structures of golden hamster ACE2 (ghACE2)/delta RBD and ghACE2/omicron BA.3 RBD. The residues Q34 and N82, which exist in many rodent ACE2s, are responsible for the lower binding affinity of ghACE2 compared to hACE2. These findings suggest that all SARS-CoV-2 VOCs may infect hamsters, highlighting the necessity of further surveillance of SARS-CoV-2 in these animals.IMPORTANCESARS-CoV-2 can infect many domestic animals, including hamsters. There is an urgent need to understand the binding mechanism of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants to hamster receptors. Herein, we showed that two hamster angiotensin-converting enzyme 2s (ACE2s) (golden hamster ACE2 and Chinese hamster ACE2) can bind to the spike protein receptor-binding domains (RBDs) of SARS-CoV-2 prototype and eight variants and that pseudotyped SARS-CoV-2 viruses can infect hamster ACE2-expressing cells. The binding pattern of golden hamster ACE2 to SARS-CoV-2 RBDs is similar to that of Chinese hamster ACE2. The two hamster ACE2s present slightly lower affinity for the RBDs of all nine SARS-CoV-2 viruses tested than human ACE2. We solved the cryo-electron microscopy (EM) structures of golden hamster ACE2 in complex with delta RBD and omicron BA.3 RBD and found that residues Q34 and N82 are responsible for the lower binding affinity of ghACE2 compared to hACE2. Our work provides valuable information for understanding the cross-species transmission mechanism of SARS-CoV-2.

Prototype and BA.5 protein nanoparticle vaccines protect against Omicron BA.5 variant in Syrian hamsters.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants with greater transmissibility or immune evasion properties has jeopardized the existing vaccine and antibody-based countermeasures. Here, we evaluated the efficacy of boosting pre-immune hamsters with protein nanoparticle vaccines (Novavax, Inc.) containing recombinant Prototype (Wuhan-1) or BA.5 S proteins against a challenge with the Omicron BA.5 variant of SARS-CoV-2. Serum antibody binding and neutralization titers were quantified before challenge, and viral loads were measured 3 days after challenge. Boosting with Prototype or BA.5 vaccine induced similar antibody binding responses against ancestral Wuhan-1 or BA.5 S proteins, and neutralizing activity of Omicron BA.1 and BA.5 variants. One and three months after vaccine boosting, hamsters were challenged with the Omicron BA.5 variant. Prototype and BA.5 vaccine-boosted hamsters had reduced viral infection in the nasal washes, nasal turbinates, and lungs compared to unvaccinated animals. Although no significant differences in virus load were detected between the Prototype and BA.5 vaccine-boosted animals, fewer breakthrough infections were detected in the BA.5-vaccinated hamsters. Thus, immunity induced by Prototype or BA.5 S protein nanoparticle vaccine boosting can protect against the Omicron BA.5 variant in the Syrian hamster model. IMPORTANCE: As SARS-CoV-2 continues to evolve, there may be a need to update the vaccines to match the newly emerging variants. Here, we compared the protective efficacy of the updated BA.5 and the original Wuhan-1 COVID-19 vaccine against a challenge with the BA.5 Omicron variant of SARS-CoV-2 in hamsters. Both vaccines induced similar levels of neutralizing antibodies against multiple variants of SARS-CoV-2. One and three months after the final immunization, hamsters were challenged with BA.5. No differences in protection against the BA.5 variant virus were observed between the two vaccines, although fewer breakthrough infections were detected in the BA.5-vaccinated hamsters. Together, our data show that both protein nanoparticle vaccines are effective against the BA.5 variant of SARS-CoV-2 but given the increased number of breakthrough infections and continued evolution, it is important to update the COVID-19 vaccine for long-term protection.

Characterization of the SARS-CoV-2 BA.5.5 and BQ.1.1 Omicron variants in mice and hamsters.

The continued evolution and emergence of novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants have resulted in challenges to vaccine and antibody efficacy. The emergence of each new variant necessitates the need to re-evaluate and refine animal models used for countermeasure testing. Here, we tested a recently circulating SARS-CoV-2 Omicron lineage variant, BQ.1.1, in multiple rodent models including K18-human ACE2 (hACE2) transgenic, C57BL/6J, and 129S2 mice, and Syrian golden hamsters. In contrast to a previously dominant BA.5.5 Omicron variant, inoculation of K18-hACE2 mice with BQ.1.1 resulted in substantial weight loss, a characteristic seen in pre-Omicron variants. BQ.1.1 also replicated to higher levels in the lungs of K18-hACE2 mice and caused greater lung pathology than the BA.5.5 variant. However, in C57BL/6J mice, 129S2 mice, and Syrian hamsters, BQ.1.1 did not cause increased respiratory tract infection or disease compared to animals administered BA.5.5. Moreover, the rates of direct contact or airborne transmission in hamsters were not significantly different after BQ.1.1 and BA.5.5 infections. Taken together, these data suggest that the BQ.1.1 Omicron variant has increased virulence in rodent species that express hACE2, possibly due to the acquisition of unique spike mutations relative to earlier Omicron variants. IMPORTANCE As severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to evolve, there is a need to rapidly assess the efficacy of vaccines and antiviral therapeutics against newly emergent variants. To do so, the commonly used animal models must also be re-evaluated. Here, we determined the pathogenicity of the BQ.1.1 SARS-CoV-2 variant in multiple SARS-CoV-2 animal models including transgenic mice expressing human ACE2 (hACE2), two strains of conventional laboratory mice, and Syrian hamsters. While BQ.1.1 and BA.5.5 infection resulted in similar levels of viral burden and clinical disease in hamsters and the conventional strains of laboratory mice tested, increases in lung infection were detected in hACE2-expressing transgenic mice, which corresponded with greater levels of pro-inflammatory cytokines and lung pathology. Taken together, our data highlight important differences in two closely related Omicron SARS-CoV-2 variant strains and provide a foundation for evaluating countermeasures.

Characterization of SARS-CoV-2 Omicron BA.4 and BA.5 isolates in rodents.

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.

The Host Response to Influenza A Virus Interferes with SARS-CoV-2 Replication during Coinfection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza A virus (IAV) represent two highly transmissible airborne pathogens with pandemic capabilities. Although these viruses belong to separate virus families-SARS-CoV-2 is a member of the family Coronaviridae, while IAV is a member of the family Orthomyxoviridae-both have shown zoonotic potential, with significant animal reservoirs in species in close contact with humans. The two viruses are similar in their capacity to infect human airways, and coinfections resulting in significant morbidity and mortality have been documented. Here, we investigate the interaction between SARS-CoV-2 USA-WA1/2020 and influenza H1N1 A/California/04/2009 virus during coinfection. Competition assays in vitro were performed in susceptible cells that were either interferon type I/III (IFN-I/-III) nonresponsive or IFN-I/-III responsive, in addition to an in vivo golden hamster model. We find that SARS-CoV-2 infection does not interfere with IAV biology in vivo, regardless of timing between the infections. In contrast, we observe a significant loss of SARS-CoV-2 replication following IAV infection. The latter phenotype correlates with increased levels of IFN-I/-III and immune priming that interferes with the kinetics of SARS-CoV-2 replication. Together, these data suggest that cocirculation of SARS-CoV-2 and IAV is unlikely to result in increased severity of disease. IMPORTANCE The human population now has two circulating respiratory RNA viruses with high pandemic potential, namely, SARS-CoV-2 and influenza A virus. As both viruses infect the airways and can result in significant morbidity and mortality, it is imperative that we also understand the consequences of getting coinfected. Here, we demonstrate that the host response to influenza A virus uniquely interferes with SARS-CoV-2 biology although the inverse relationship is not evident. Overall, we find that the host response to both viruses is comparable to that to SARS-CoV-2 infection alone.

[Specific features of the pathology of the respiratory system in SARS-CoV-2 (Coronaviridae: Coronavirinae: Betacoronavirus: Sarbecovirus) infected Syrian hamsters (Mesocricetus auratus)].

INTRODUCTION: Verification of histological changes in respiratory system using Syrian (golden) hamsters (Mesocricetus auratus) as experimental model is an important task for preclinical studies of drugs intended for prevention and treatment of the novel coronavirus infection COVID-19.The aim of this work was to study pathological changes of pulmonary tissue in SARS-CoV-2 (Coronaviridae: Coronavirinae: Betacoronavirus; Sarbecovirus) experimental infection in Syrian hamsters. MATERIAL AND METHODS: Male Syrian hamsters weighting 80-100 g were infected by intranasal administration of culture SARS-CoV-2 at dose 4 × 104 TCID50/ml (TCID is tissue culture infectious dose). Animals were euthanatized on 3, 7 and 14 days after infection, with gravimetric registration. The viral load in lungs was measured using the polymerase chain reaction (PCR). Right lung and trachea tissues were stained with hematoxylin-eosin and according to Mallory. RESULTS AND DISCUSSION: The highest viral replicative activity in lungs was determined 3 days after the infection. After 7 days, on a background of the decrease of the viral load in lungs, a pathologically significant increase of the organ's gravimetric parameters was observed. Within 3 to 14 days post-infection, the lung histologic pattern had been showing the development of inflammation with a succession of infiltrative-proliferative, edematousmacrophagal and fibroblastic changes. It was found that initial changes in respiratory epithelium can proceed without paranecrotic interstitial inflammation, while in the formation of multiple lung parenchyma lesions, damage to the epithelium of bronchioles and acinar ducts can be secondary. The appearance of epithelioid large-cell metaplastic epithelium, forming pseudoacinar structures, was noted as a pathomorphological feature specific to SARS-CoV-2 infection in Syrian hamsters. CONCLUSION: As a result of the study, the specific features of the pathology of the respiratory system in SARSCoV-2 infected Syrian hamsters were described. These findings are of practical importance as reference data that can be used for preclinical studies to assess the effectiveness of vaccines and potential drugs.

The omicron (B.1.1.529) SARS-CoV-2 variant of concern does not readily infect Syrian hamsters.

The emergence of SARS-CoV-2 variants of concern (VoCs) has exacerbated the COVID-19 pandemic. End of November 2021, a new SARS-CoV-2 variant namely the omicron (B.1.1.529) emerged. Since this omicron variant is heavily mutated in the spike protein, WHO classified this variant as the 5th variant of concern (VoC). We previously demonstrated that the ancestral strain and the other SARS-CoV-2 VoCs replicate efficiently in and cause a COVID19-like pathology in Syrian hamsters. We here wanted to explore the infectivity of the omicron variant in comparison to the ancestral D614G strain in the hamster model. Strikingly, in hamsters that had been infected with the omicron variant, a 3 log10 lower viral RNA load was detected in the lungs as compared to animals infected with D614G and no infectious virus was detectable in this organ. Moreover, histopathological examination of the lungs from omicron-infected hamsters revealed no signs of peri-bronchial inflammation or bronchopneumonia.

Enhanced fitness of SARS-CoV-2 variant of concern Alpha but not Beta.

Emerging variants of concern (VOCs) are driving the COVID-19 pandemic1,2. Experimental assessments of replication and transmission of major VOCs and progenitors are needed to understand the mechanisms of replication and transmission of VOCs3. Here we show that the spike protein (S) from Alpha (also known as B.1.1.7) and Beta (B.1.351) VOCs had a greater affinity towards the human angiotensin-converting enzyme 2 (ACE2) receptor than that of the progenitor variant S(D614G) in vitro. Progenitor variant virus expressing S(D614G) (wt-S614G) and the Alpha variant showed similar replication kinetics in human nasal airway epithelial cultures, whereas the Beta variant was outcompeted by both. In vivo, competition experiments showed a clear fitness advantage of Alpha over wt-S614G in ferrets and two mouse models-the substitutions in S were major drivers of the fitness advantage. In hamsters, which support high viral replication levels, Alpha and wt-S614G showed similar fitness. By contrast, Beta was outcompeted by Alpha and wt-S614G in hamsters and in mice expressing human ACE2. Our study highlights the importance of using multiple models to characterize fitness of VOCs and demonstrates that Alpha is adapted for replication in the upper respiratory tract and shows enhanced transmission in vivo in restrictive models, whereas Beta does not overcome Alpha or wt-S614G in naive animals.

Abnormal Blood Coagulation and Kidney Damage in Aged Hamsters Infected with Severe Acute Respiratory Syndrome Coronavirus 2.

Systemic symptoms have often been observed in patients with coronavirus disease 2019 (COVID-19) in addition to pneumonia, however, the details are still unclear due to the lack of an appropriate animal model. In this study, we investigated and compared blood coagulation abnormalities and tissue damage between male Syrian hamsters of 9 (young) and over 36 (aged) weeks old after intranasal infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Despite similar levels of viral replication and inflammatory responses in the lungs of both age groups, aged but not young hamsters showed significant prolongation of prothrombin time and prominent acute kidney damage. Moreover, aged hamsters demonstrated increased intravascular coagulation time-dependently in the lungs, suggesting that consumption of coagulation factors causes prothrombin time prolongation. Furthermore, proximal urinary tract damage and mesangial matrix expansion were observed in the kidneys of the aged hamsters at early and later disease stages, respectively. Given that the severity and mortality of COVID-19 are higher in elderly human patients, the effect of aging on pathogenesis needs to be understood and should be considered for the selection of animal models. We, thus, propose that the aged hamster is a good small animal model for COVID-19 research.

Broad sarbecovirus neutralization by a human monoclonal antibody.

The recent emergence of SARS-CoV-2 variants of concern1-10 and the recurrent spillovers of coronaviruses11,12 into the human population highlight the need for broadly neutralizing antibodies that are not affected by the ongoing antigenic drift and that can prevent or treat future zoonotic infections. Here we describe a human monoclonal antibody designated S2X259, which recognizes a highly conserved cryptic epitope of the receptor-binding domain and cross-reacts with spikes from all clades of sarbecovirus. S2X259 broadly neutralizes spike-mediated cell entry of SARS-CoV-2, including variants of concern (B.1.1.7, B.1.351, P.1, and B.1.427/B.1.429), as well as a wide spectrum of human and potentially zoonotic sarbecoviruses through inhibition of angiotensin-converting enzyme 2 (ACE2) binding to the receptor-binding domain. Furthermore, deep-mutational scanning and in vitro escape selection experiments demonstrate that S2X259 possesses an escape profile that is limited to a single substitution, G504D. We show that prophylactic and therapeutic administration of S2X259 protects Syrian hamsters (Mesocricetus auratus) against challenge with the prototypic SARS-CoV-2 and the B.1.351 variant of concern, which suggests that this monoclonal antibody is a promising candidate for the prevention and treatment of emergent variants and zoonotic infections. Our data reveal a key antigenic site that is targeted by broadly neutralizing antibodies and will guide the design of vaccines that are effective against all sarbecoviruses.

In vivo monoclonal antibody efficacy against SARS-CoV-2 variant strains.

Rapidly emerging SARS-CoV-2 variants jeopardize antibody-based countermeasures. Although cell culture experiments have demonstrated a loss of potency of several anti-spike neutralizing antibodies against variant strains of SARS-CoV-21-3, the in vivo importance of these results remains uncertain. Here we report the in vitro and in vivo activity of a panel of monoclonal antibodies (mAbs), which correspond to many in advanced clinical development by Vir Biotechnology, AbbVie, AstraZeneca, Regeneron and Lilly, against SARS-CoV-2 variant viruses. Although some individual mAbs showed reduced or abrogated neutralizing activity in cell culture against B.1.351, B.1.1.28, B.1.617.1 and B.1.526 viruses with mutations at residue E484 of the spike protein, low prophylactic doses of mAb combinations protected against infection by many variants in K18-hACE2 transgenic mice, 129S2 immunocompetent mice and hamsters, without the emergence of resistance. Exceptions were LY-CoV555 monotherapy and LY-CoV555 and LY-CoV016 combination therapy, both of which lost all protective activity, and the combination of AbbVie 2B04 and 47D11, which showed a partial loss of activity. When administered after infection, higher doses of several mAb cocktails protected in vivo against viruses with a B.1.351 spike gene. Therefore, many-but not all-of the antibody products with Emergency Use Authorization should retain substantial efficacy against the prevailing variant strains of SARS-CoV-2.

ALG-097111, a potent and selective SARS-CoV-2 3-chymotrypsin-like cysteine protease inhibitor exhibits in vivo efficacy in a Syrian Hamster model.

There is an urgent need for antivirals targeting the SARS-CoV-2 virus to fight the current COVID-19 pandemic. The SARS-CoV-2 main protease (3CLpro) represents a promising target for antiviral therapy. The lack of selectivity for some of the reported 3CLpro inhibitors, specifically versus cathepsin L, raises potential safety and efficacy concerns. ALG-097111 potently inhibited SARS-CoV-2 3CLpro (IC50 = 7 nM) without affecting the activity of human cathepsin L (IC50 > 10 µM). When ALG-097111 was dosed in hamsters challenged with SARS-CoV-2, a robust and significant 3.5 log10 (RNA copies/mg) reduction of the viral RNA copies and 3.7 log10 (TCID50/mg) reduction in the infectious virus titers in the lungs was observed. These results provide the first in vivo validation for the SARS-CoV-2 3CLpro as a promising therapeutic target for selective small molecule inhibitors.

DNA Based Vaccine Expressing SARS-CoV-2 Spike-CD40L Fusion Protein Confers Protection Against Challenge in a Syrian Hamster Model.

SARS-CoV-2 infections present a tremendous threat to public health. Safe and efficacious vaccines are the most effective means in preventing the infections. A variety of vaccines have demonstrated excellent efficacy and safety around the globe. Yet, development of alternative forms of vaccines remains beneficial, particularly those with simpler production processes, less stringent storage conditions, and the capability of being used in heterologous prime/boost regimens which have shown improved efficacy against many diseases. Here we reported a novel DNA vaccine comprised of the SARS-CoV-2 spike protein fused with CD40 ligand (CD40L) serving as both a targeting ligand and molecular adjuvant. A single intramuscular injection in Syrian hamsters induced significant neutralizing antibodies 3-weeks after vaccination, with a boost substantially improving immune responses. Moreover, the vaccine also reduced weight loss and suppressed viral replication in the lungs and nasal turbinates of challenged animals. Finally, the incorporation of CD40L into the DNA vaccine was shown to reduce lung pathology more effectively than the DNA vaccine devoid of CD40L. These results collectively indicate that this DNA vaccine candidate could be further explored because of its efficacy and known safety profile.

Spike mutation D614G alters SARS-CoV-2 fitness.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein substitution D614G became dominant during the coronavirus disease 2019 (COVID-19) pandemic1,2. However, the effect of this variant on viral spread and vaccine efficacy remains to be defined. Here we engineered the spike D614G substitution in the USA-WA1/2020 SARS-CoV-2 strain, and found that it enhances viral replication in human lung epithelial cells and primary human airway tissues by increasing the infectivity and stability of virions. Hamsters infected with SARS-CoV-2 expressing spike(D614G) (G614 virus) produced higher infectious titres in nasal washes and the trachea, but not in the lungs, supporting clinical evidence showing that the mutation enhances viral loads in the upper respiratory tract of COVID-19 patients and may increase transmission. Sera from hamsters infected with D614 virus exhibit modestly higher neutralization titres against G614 virus than against D614 virus, suggesting that the mutation is unlikely to reduce the ability of vaccines in clinical trials to protect against COVID-19, and that therapeutic antibodies should be tested against the circulating G614 virus. Together with clinical findings, our work underscores the importance of this variant in viral spread and its implications for vaccine efficacy and antibody therapy.

A single-dose live-attenuated YF17D-vectored SARS-CoV-2 vaccine candidate.

The expanding pandemic of coronavirus disease 2019 (COVID-19) requires the development of safe, efficacious and fast-acting vaccines. Several vaccine platforms are being leveraged for a rapid emergency response1. Here we describe the development of a candidate vaccine (YF-S0) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that uses live-attenuated yellow fever 17D (YF17D) vaccine as a vector to express a noncleavable prefusion form of the SARS-CoV-2 spike antigen. We assess vaccine safety, immunogenicity and efficacy in several animal models. YF-S0 has an excellent safety profile and induces high levels of SARS-CoV-2 neutralizing antibodies in hamsters (Mesocricetus auratus), mice (Mus musculus) and cynomolgus macaques (Macaca fascicularis), and-concomitantly-protective immunity against yellow fever virus. Humoral immunity is complemented by a cellular immune response with favourable T helper 1 polarization, as profiled in mice. In a hamster model2 and in macaques, YF-S0 prevents infection with SARS-CoV-2. Moreover, a single dose conferred protection from lung disease in most of the vaccinated hamsters within as little as 10 days. Taken together, the quality of the immune responses triggered and the rapid kinetics by which protective immunity can be attained after a single dose warrant further development of this potent SARS-CoV-2 vaccine candidate.

Animal models for COVID-19.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the aetiological agent of coronavirus disease 2019 (COVID-19), an emerging respiratory infection caused by the introduction of a novel coronavirus into humans late in 2019 (first detected in Hubei province, China). As of 18 September 2020, SARS-CoV-2 has spread to 215 countries, has infected more than 30 million people and has caused more than 950,000 deaths. As humans do not have pre-existing immunity to SARS-CoV-2, there is an urgent need to develop therapeutic agents and vaccines to mitigate the current pandemic and to prevent the re-emergence of COVID-19. In February 2020, the World Health Organization (WHO) assembled an international panel to develop animal models for COVID-19 to accelerate the testing of vaccines and therapeutic agents. Here we summarize the findings to date and provides relevant information for preclinical testing of vaccine candidates and therapeutic agents for COVID-19.

Adenovirus 14p1 Immunopathogenesis during Lung Infection in the Syrian Hamster.

Adenovirus (Ad) infections are usually mild and self-limited, with minimal inflammatory responses. During worldwide outbreaks, Ad14p1, an emerging Ad14 variant, has caused severe pulmonary disease, including acute respiratory distress syndrome (ARDS). This increased pathogenicity of Ad14p1 is not completely understood. In initial studies, we observed that infection of Syrian hamsters with Ad14p1 can cause a patchy bronchopneumonia, with an increased intensity of inflammation, compared to wild type Ad14 infection. The current study compared the dynamics of the immunopathogenesis of Ad14 and Ad14p1 infection of hamster lungs through the first two weeks after infection. Little difference was seen in infection-induced inflammation at day 1. Beginning at day 3, Ad14p1-infected hamsters showed marked inflammation that continued through to day 7. The inflammation began to resolve by day 10 but was still detectable at day 14. In contrast, Ad14-infected hamsters showed little inflammation during the 14-day period of observation. Inflammatory cell type analysis revealed that, at day 1, hamsters infected with either virus had predominantly neutrophil infiltration that began to resolve by day 3. However, at day 5, Ad14p1-infected hamsters had a second wave of neutrophil infiltration that was accompanied by edema which persisted to a variable extent through to day 10. These differences were not explained by an increased Ad14p1 replication rate, compared with Ad14 in vitro, but there was prolonged persistence of Ad14p1 in hamster lungs. There were differences in lung tissue cytokine and chemokine responses to Ad14p1 vs. Ad14 infection that might account for the increased leukocyte infiltrates in Ad14p1-infected hamsters. This animal model characterization provides the basis for future translational studies of the viral genetic mechanisms that control the increased immunopathogenesis of the emergent, Ad14p1 strain.

Pathogenesis and transmission of SARS-CoV-2 in golden hamsters.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel coronavirus with high nucleotide identity to SARS-CoV and to SARS-related coronaviruses that have been detected in horseshoe bats, has spread across the world and had a global effect on healthcare systems and economies1,2. A suitable small animal model is needed to support the development of vaccines and therapies. Here we report the pathogenesis and transmissibility of SARS-CoV-2 in golden (Syrian) hamsters (Mesocricetus auratus). Immunohistochemistry assay demonstrated the presence of viral antigens in nasal mucosa, bronchial epithelial cells and areas of lung consolidation on days 2 and 5 after inoculation with SARS-CoV-2, followed by rapid viral clearance and pneumocyte hyperplasia at 7 days after inoculation. We also found viral antigens in epithelial cells of the duodenum, and detected viral RNA in faeces. Notably, SARS-CoV-2 was transmitted efficiently from inoculated hamsters to naive hamsters by direct contact and via aerosols. Transmission via fomites in soiled cages was not as efficient. Although viral RNA was continuously detected in the nasal washes of inoculated hamsters for 14 days, the communicable period was short and correlated with the detection of infectious virus but not viral RNA. Inoculated and naturally infected hamsters showed apparent weight loss on days 6-7 post-inoculation or post-contact; all hamsters returned to their original weight within 14 days and developed neutralizing antibodies. Our results suggest that features associated with SARS-CoV-2 infection in golden hamsters resemble those found in humans with mild SARS-CoV-2 infections.

Establishment of an RNA polymerase II-driven reverse genetics system for Nipah virus strains from Malaysia and Bangladesh.

Nipah virus (NiV) has emerged as a highly lethal zoonotic paramyxovirus that is capable of causing a febrile encephalitis and/or respiratory disease in humans for which no vaccines or licensed treatments are currently available. There are two genetically and geographically distinct lineages of NiV: NiV-Malaysia (NiV-M), the strain that caused the initial outbreak in Malaysia, and NiV-Bangladesh (NiV-B), the strain that has been implicated in subsequent outbreaks in India and Bangladesh. NiV-B appears to be both more lethal and have a greater propensity for person-to-person transmission than NiV-M. Here we describe the generation and characterization of stable RNA polymerase II-driven infectious cDNA clones of NiV-M and NiV-B. In vitro, reverse genetics-derived NiV-M and NiV-B were indistinguishable from a wildtype isolate of NiV-M, and both viruses were pathogenic in the Syrian hamster model of NiV infection. We also describe recombinant NiV-M and NiV-B with enhanced green fluorescent protein (EGFP) inserted between the G and L genes that enable rapid and sensitive detection of NiV infection in vitro. This panel of molecular clones will enable studies to investigate the virologic determinants of henipavirus pathogenesis, including the pathogenic differences between NiV-M and NiV-B, and the high-throughput screening of candidate therapeutics.

Protocadherin-1 is essential for cell entry by New World hantaviruses.

The zoonotic transmission of hantaviruses from their rodent hosts to humans in North and South America is associated with a severe and frequently fatal respiratory disease, hantavirus pulmonary syndrome (HPS)1,2. No specific antiviral treatments for HPS are available, and no molecular determinants of in vivo susceptibility to hantavirus infection and HPS are known. Here we identify the human asthma-associated gene protocadherin-1 (PCDH1)3-6 as an essential determinant of entry and infection in pulmonary endothelial cells by two hantaviruses that cause HPS, Andes virus (ANDV) and Sin Nombre virus (SNV). In vitro, we show that the surface glycoproteins of ANDV and SNV directly recognize the outermost extracellular repeat domain of PCDH1-a member of the cadherin superfamily7,8-to exploit PCDH1 for entry. In vivo, genetic ablation of PCDH1 renders Syrian golden hamsters highly resistant to a usually lethal ANDV challenge. Targeting PCDH1 could provide strategies to reduce infection and disease caused by New World hantaviruses.