Cross-sectional; Prospective cohort; Ecological
Study population and setting
To validate the accuracy of the serological assay developed in the study, the authors tested serum samples from 35 species of domestic and wild animals. The ability of the assay to detect coronavirus-negative animals (specificity) was evaluated using 116 serum samples from pathogen-free chickens, ducks, mice, rats, and pigs; additional testing on 232 samples from experimental animals with no known exposure to coronavirus (guinea pigs, dog, and rhesus monkeys) was also performed. To evaluate whether the serological test cross-reacts with other animal coronaviruses, the authors immunized pigs, chickens, mice, and rats with viruses that commonly infect each corresponding species. The ability of the test to detect coronavirus-positive animals with known infection (sensitivity) was evaluated on five ferrets that were infected with SARS-CoV-2, and sampled up to 22 days after infection; five other ferrets were given placebo injections. Finally, the authors used the serological assay to test clinical samples from domesticated and wild animals with unknown exposure to SARS-CoV-2, some of which included cats and dogs sampled after the outbreak of COVID-19.
Summary of Main Findings
None of the pathogen-free animals, experimental animals, or animals infected with other coronaviruses tested positive with the assay, demonstrating that the test is specific and does not cross-react with other coronavirus antibodies. All five of the infected ferrets had detectable antibodies 22 days after infection, while none of the placebo ferrets were positive, indicating that ferrets are capable of being infected with SARS-CoV-2 and that the test is sensitive to known coronavirus infections. None of the clinical serum samples from 35 species of domestic or wild animals tested positive, including stray dogs and cats in Wuhan, China and pet dogs that may have had contact with an owner with SARS-CoV-2.
The authors made an extensive effort to test the sensitivity, specificity, and cross-reactivity of their serological assay. The advantage of using serological tests for SARS-CoV-2 rather than approaches that detect viral RNA is that antibodies to infection typically last longer than the actual viral infection, so there is a larger time window for detection. The antibody tests can also be performed in laboratories with less stringent biosafety regulations than sequencing approaches. Finally, the particular serological test used in the study is not limited to detecting antibodies in a particular species, and could be used broadly for surveys of many animals.
The most significant limitation of this study is that the authors do not indicate when and where many of the domestic and wild animal samples were collected. With the exception of the stray and pet cats and dogs, some of which were from the COVID-19 outbreak, including from Wuhan, most of the remaining animal samples are of totally unknown origin. Furthermore, only limited samples (<10) were collected from most of the wild animal species in the study. Thus, the authors cannot conclude from these results that these animals cannot be infected with SARS-CoV-2 and serve as potential intermediate hosts, only that the limited samples from the sampled populations were negative at the time of sampling.
This study expands the number of potential domestic and wild animal hosts that have been tested for evidence of SARS-CoV-2 infection.