A recent study by scientists at the Cummings School of Veterinary Medicine at Tufts University has revealed that ferrets are not susceptible to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection even if they are directly exposed to symptomatic coronavirus disease 2019 (COVID-19) patients for a prolonged time. The study is currently available on the bioRxiv* preprint server.
The recent outbreak of SARS-CoV-2 is considered to be one of the deadliest pandemic situations, which already has infected more than 23.5 million people and claimed over 810,000 lives worldwide. Given rapid viral transmission, it is still uncertain whether natural SARS-CoV-2 infection of animals plays a role in augmenting the pandemic or threatening wildlife.
Analysis of angiotensin-converting enzyme-2 (ACE2), which is responsible for viral entry to host cells, has shown that amino acid residues responsible for viral binding are moderately conserved between humans and domestic animals, and reverse-zoonosis (human to animal transmission) has been noticed in case of SARS-CoV-2 infection. So far, human to animal transmission of SARS-CoV-2 has been observed in European mink, resulting in outbreaks in several mink farms in the United States and European countries.
Current study design
In the current study, the researchers selected ferrets as an animal model to study SARS-CoV-2 transmission, because these domestic mammals are widely used in laboratories to study experimental infection and clinical outcomes of human diseases caused by respiratory viruses.
During the initial period of COVID-19 pandemic in the New England area (March 2020), the researchers performed a rapid response study to evaluate possibilities of human to animal spillover and onward transmission in many domestic, farm, and wild animals. Their main aim was to understand the possible nature of SARS-CoV-2 transmission at these interfaces to refine public health guidelines as well as assess possible animal or human health risks related to spillover. They also aimed at investigating whether infected animals can potentially act as endemic reservoirs.
The researchers conducted the experiments with 29 ferrets living in one household with two human participants suffering from symptomatic (fever, fatigue, sore throat, anosmia, and migraine) COVID-19. Both individuals cared for the ferrets throughout their course of infection.
Absence of natural human to ferret transmission of SARS-CoV-2
To check whether there is human to animal transmission, oral swabs were collected from each ferret over two weeks using home sampling kits. All collected samples were tested for active SARS-CoV-2 infection via polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA). Surprisingly, the researchers found that all ferrets tested negative for active or recent SARS-CoV-2 infection. Moreover, they found that none of the experimental ferrets developed antibodies against the receptor-binding domain of SARS-CoV-2, indicating the absence of seroconversion.
Identification of three mutations in the surface glycoprotein of mustelid-derived SARS-CoV-2
Because previous studies have shown that ferrets are susceptible to experimentally executed SARS-CoV-2 infection, the researchers hypothesized that there might be certain factors that protect ferrets from natural SARS-CoV-2 infection. To confirm their hypothesis, the researchers analyzed available genetic sequences of SARS-CoV-2 that are known to naturally infect European minks and then experimentally infect ferrets. Three mutations (N501T, D614G, and S686G) in the viral spike protein were identified. Of these mutations, N501T was in the receptor-binding domain of the viral spike protein, which interacts with host ACE2 at Y41, K353, G354, and D355. Mustelids and humans differ from each other only at ACE2 G354R. Because both N501T and G354R are present in minks and ferrets, ACE2 is expected to provide a certain level of protection against SARS-CoV-2 infection. According to the researchers, ACE2 provides only a weak host barrier against viral entry to mustelids, given the fact that SARS-CoV-2 without N501T mutation has been known to infect minks initially. As described by the researchers, N501T mutation may have increased viral pathogenicity in mustelids; however, this mutation is not required for species to species transmission.
Because D614G mutation has become highly prevalent in humans, the researchers concluded that the observed D614G mutation in ferrets may be due to variation in human donors and is not related to ferret infection.
Interestingly, the researchers observed that S686G mutation, which was present only in ferrets, is located directly adjacent to the novel S1/S2 polybasic cleavage site that is associated with virulence. Using computational prediction analysis, the researchers found that this specific mutation is predicted to reduce furin activity, which in turn may reduce viral transmission and pathogenicity. These findings suggest that S686G mutation may provide a strong host barrier, specifically to ferrets. As described by the researchers, the current study findings indicate that natural infection in ferrets is limited by virus and host genetic barriers, which can only be overcome by experimental infections using concentrated viral preparations obtained from humans.
bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.