Surveillance for SARS-CoV-2 in Norway Rats (Rattus norvegicus) from Southern Ontario

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from wildlife has raised concerns about spillover from humans to animals, the establishment of novel wildlife reservoirs, and the potential for future outbreaks caused by variants of wildlife origin. Norway rats (Rattus norvegicus) are abundant in urban areas and live in close proximity to humans, providing the opportunity for spillover of SARS-CoV-2. Evidence of SARS-CoV-2 infection and exposure has been reported in Norway rats. We investigated SARS-CoV-2 infection and exposure in Norway rats from Southern Ontario, Canada. From October 2019 to June 2021, 224 rats were submitted by collaborating pest control companies. The majority of samples were collected in Windsor (79.9%;n = 179), Hamilton (13.8%;n = 31), and the Greater Toronto Area (5.8%;n = 13). Overall, 50.0% (n = 112) were female and most rats were sexually mature (55.8%;n = 125). Notably, 202 samples were collected prior to the emergence of variants of concern (VOC) and 22 were collected while the Alpha variant (B.1.1.7) was the predominant circulating VOC in humans. Nasal turbinate (n = 164) and small intestinal (n = 213) tissue samples were analyzed for SARS-CoV-2 RNA by RT-PCR. Thoracic cavity fluid samples (n = 213) were tested for neutralizing antibodies using a surrogate virus neutralization test (sVNT) (GenScript cPass);confirmatory plaque reduction neutralization test (PRNT) was conducted on presumptive positive samples. We did not detect SARS-CoV-2 RNA in any samples tested. Two out of eleven samples positive on sVNT had neutralizing antibodies confirmed positive by PRNT (1 : 40 and 1 : 320 PRNT70);both were collected prior to the emergence of VOC. It is imperative that efforts to control and monitor SARS-CoV-2 include surveillance of rats and other relevant wildlife species as novel variants continue to emerge.

First detection of SARS-CoV-2 infection in Canadian wildlife identified in free-ranging white-tailed deer (Odocoileus virginianus) from southern Quebec, Canada (preprint)

White-tailed deer are susceptible to SARS-CoV-2 and represent a relevant species for surveillance. We investigated SARS-CoV-2 infection in white-tailed deer in Quebec, Canada. In November 2021, 251 nasal swabs and 104 retropharyngeal lymph nodes from 258 deer were analyzed for SARS-CoV-2 RNA, whole genome sequencing and virus isolation and 251 thoracic cavity fluid samples were tested for neutralizing antibodies. We detected SARS-CoV-2 RNA in three nasal swabs from the Estrie region and virus was isolated from two samples; evidence of past exposure was detected among deer from the same region. Viral sequences were assigned to lineage AY.44, a sublineage of B.1.617.2. All deer sequences clustered with human GISAID sequences collected in October 2021 from Vermont USA, which borders the Estrie region. Mutations in the S-gene and a deletion in ORF8 encoding a truncated protein were detected. These findings underscore the importance of ongoing surveillance of key wildlife species for SARS-CoV-2.

SARS-CoV-2 wildlife surveillance in Ontario and Quebec, Canada (preprint)

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus responsible for the COVID-19 pandemic, is capable of infecting a variety of wildlife species. Wildlife living in close contact with humans are at an increased risk of SARS-CoV-2 exposure and if infected have the potential to become a reservoir for the pathogen, making control and management more difficult. Objective: To conduct SARS-CoV-2 surveillance in urban wildlife from Ontario and Quebec, Canada, increasing our knowledge of the epidemiology of the virus and our chances of detecting spillover from humans into wildlife. Methods: Using a One Health approach, we leveraged activities of existing research, surveillance, and rehabilitation programs among multiple agencies to collect samples from 776 animals from 17 different wildlife species between June 2020 and May 2021. Samples from all animals were tested for the presence of SARS-CoV-2 viral RNA, and a subset of samples from 219 animals across 3 species (raccoons, Procyon lotor; striped skunks, Mephitis mephitis; and mink, Neovison vison) were also tested for the presence of neutralizing antibodies. Results: No evidence of SARS-CoV-2 viral RNA or neutralizing antibodies was detected in any of the tested samples. Conclusion: Although we were unable to identify positive SARS-CoV-2 cases in wildlife, continued research and surveillance activities are critical to better understand the rapidly changing landscape of susceptible animal species. Collaboration between academic, public and animal health sectors should include experts from relevant fields to build coordinated surveillance and response capacity.

Evaluation of a Commercial Culture-free Neutralization Antibody Detection Kit for Severe Acute Respiratory Syndrome-Related Coronavirus-2 and Comparison with an Anti-RBD ELISA Assay (preprint)

BackgroundSARS-CoV-2 surrogate neutralization assays that obviate the need for viral culture offer substantial advantages regarding throughput and cost. The cPass SARS-CoV-2 Neutralization Antibody Detection Kit (Genscript) is the first such commercially available assay, detecting antibodies that block RBD/ACE-2 interaction. We aimed to evaluate cPass to inform its use and assess its added value compared to anti-RBD ELISA assays. MethodsSerum reference panels comprising 205 specimens were used to compare cPass to plaque-reduction neutralization test (PRNT) and a pseudotyped lentiviral neutralization (PLV) assay for detection of neutralizing antibodies. We assessed the correlation of cPass with an ELISA detecting anti-RBD IgG, IgM, and IgA antibodies at a single timepoint and across intervals from onset of symptoms of SARS-CoV-2 infection. ResultsCompared to PRNT-50, cPass sensitivity ranged from 77% - 100% and specificity was 95% - 100%. Sensitivity was also high compared to the pseudotyped lentiviral neutralization assay (93% [95%CI 85-97]), but specificity was lower (58% [95%CI 48-67]). Highest agreement between cPass and ELISA was for anti-RBD IgG (r=0.823). Against the pseudotyped lentiviral neutralization assay, anti-RBD IgG sensitivity (99% [95%CI 94-100]) was very similar to that of cPass, but overall specificity was lower (37% [95%CI 28-47]). Against PRNT-50, results of cPass and anti-RBD IgG were nearly identical. ConclusionsThe added value of cPass compared to an IgG anti-RBD ELISA was modest.

North American deer mice are susceptible to SARS-CoV-2 (preprint)

The zoonotic spillover of the pandemic SARS-coronavirus 2 (SARS-CoV-2) from an animal reservoir, currently presumed to be the Chinese horseshoe bat, into a naive human population has rapidly resulted in a significant global public health emergency. Worldwide circulation of SARS-CoV-2 in humans raises the theoretical risk of reverse zoonosis events with wildlife, reintroductions of SARS-CoV-2 into permissive non-domesticated animals, potentially seeding new host reservoir species and geographic regions in which bat SARS-like coronaviruses have not historically been endemic. Here we report that North American deer mice (Peromyscus maniculatus) and some closely related members of the Cricetidae family of rodents possess key amino acid residues within the angiotensin-converting enzyme 2 (ACE2) receptor known to confer SARS-CoV-2 spike protein binding. Peromyscus rodent species are widely distributed across North America and are the primary host reservoirs of several emerging pathogens that repeatedly spill over into humans including Borrelia burgdorferi, the causative agent of Lyme disease, deer tick virus, and Sin Nombre orthohantavirus, the causative agent of hantavirus pulmonary syndrome (HPS). We demonstrate that adult deer mice are susceptible to SARS-CoV-2 infection following intranasal exposure to a human isolate, resulting in viral replication in the upper and lower respiratory tract with little or no signs of disease. Further, shed infectious virus is detectable in nasal washes, oropharyngeal and rectal swabs, and viral RNA is detectable in feces and occasionally urine. We further show that deer mice are capable of transmitting SARS-CoV-2 to naive deer mice through direct contact. The extent to which these observations may translate to wild deer mouse populations remains unclear, and the risk of reverse zoonosis and/or the potential for the establishment of Peromyscus rodents as a North American reservoir for SARS-CoV-2 is unknown. Nevertheless, efforts to monitor wild, peri-domestic Peromyscus rodent populations are likely warranted as the SARS-CoV-2 pandemic progresses.