In a recent study posted to the bioRxiv* preprint server, researchers from Belgium conducted severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) surveillance among all mammal species found in two zoos in the country to determine the potential range of hosts of SARS-CoV-2 and assess the risk of conservation in captivity or in situ of wild animals.
Study: SARS-CoV-2 surveillance between 2020 and 2021 of all mammalian species in two Flemish zoos (Antwerp Zoo and Planckendael Zoo). Image Credit: ArtMediaFactory / Shutterstock.com
*Important notice: bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.
Background
Extensive research in the form of experimental and observational studies has reported that SARS-CoV-2 can infect a wide range of mammals apart from humans, including macaques, ferrets, domestic cats, North American deer mice, American minks, Egyptian fruit bats, raccoon dogs, white-tailed deer, and Syrian hamsters.
Studies that compared orthologs of the angiotensin-converting enzyme 2 (ACE-2) receptor also revealed that various species might be susceptible to SARS-CoV-2. However, similarities in the ACE-2 receptor alone are not good indicators of susceptibility.
There have been 64 events where SARS-CoV-2 infections have been reported from 17 species of zoo animals, with felines being the most commonly infected group. However, these reports have been based on external signs or symptoms such as nasal discharge and cough, changes in behavior, or death due to the coronavirus disease 2019 (COVID-19).
Zoos present an ideal opportunity to conduct COVID-19 surveillance across a vast number of species from various geographic regions.
About the study
The present study was conducted in the Planckendael and Antwerp Zoos in Mechelen and Antwerp, respectively. Samples were collected during early September, mid-October, and mid-December in 2020 and July 2021.
The 2020 sampling periods were during the dominance of the ancestral Wuhan-Hu-1 strain, whereas the July 2021 sampling was conducted during the dominance of the Delta variant, which was more contagious than other SARS-CoV-2 variants at the time.
Zookeepers collected fecal samples from 103 mammal species, with 48 and 67 samples collected from the Antwerp and Planckendael zoos, respectively. Blood samples from routine veterinary check-ups were also available for some animals spanning periods before and after 2020.
In the Antwerp zoo, primates comprised the largest group, followed by carnivores, while in the Planckendael zoo, the most commonly sampled group included even-toed ungulates and cetaceans.
Following the July 2021 sample collection, two hippopotamuses from the Antwerp zoo exhibited symptoms of SARS-CoV-2 infection, which was subsequently confirmed through immunochemistry and polymerase chain reaction (PCR) assay of nasal swabs.
Therefore, targeted surveillance was conducted in December 2021 among animals that may have encountered the infected hippos and were thought to be susceptible or of interest due to their conservation status. These included large felines and primates.
Ribonucleic acid extraction and PCR were conducted to screen the samples for SARS-CoV-2. Tests were also conducted to validate the use of the Pan-Coronavirus (Pan-CoV) Real-Time PCR system to detect SARS-CoV-2. Additionally, surrogate virus neutralization tests were used to test serum samples for anti-SARS-CoV-2 antibodies.
Results
Of the 1,523 fecal samples collected from 103 mammal species, none tested positive for SARS-CoV-2 in the Pan-CoV screening. Furthermore, all serum samples were seronegative, thus indicating that these mammals had not contracted SARS-CoV-2 around the time of the screening. Apart from the two hippos with symptomatic SARS-CoV-2 infections, no evidence of SARS-CoV-2 infections was observed in the Planckendael and Antwerp zoos.
Although studies have reported that SARS-CoV-2 could remain stable in feces for up to four days depending on certain factors like pH, the authors believe that delays between fecal excretion and collection could have influenced the sample quality.
Furthermore, studies have indicated that SARS-CoV-2 RNA can be detected in fecal samples for up to 21.8 days after the infection. With the sampling sessions in the present study being on average six weeks apart, the authors admit that some SARS-CoV-2 infections could have gone undetected.
The seronegativity of the available blood samples indicated that widespread SARS-CoV-2 infections had not occurred across species; however, a systematic collection of blood samples could have provided information on previous SARS-CoV-2 infections in these species.
Previous studies have identified asymptomatic SARS-CoV-2-infected zookeepers as the source of infection among animals in other zoos. In both these zoos in Belgium, visitors and zookeepers wore face masks while interacting with the animals, which could explain the absence of SARS-CoV-2 infection among the animals during the study period.
Conclusions
During the periods of surveillance between 2020 and 2021, no mammal species in the Planckendael and Antwerp zoos tested positive for SARS-CoV-2, other than the two hippopotamuses that had symptomatic SARS-CoV-2 infections. The authors believe that since the hippos were housed in an area where visitors entered the zoo, they had an increased risk of infection.
Overall, the study findings indicate that analysis of the ACE-2 receptor alone is not enough to determine susceptibility in mammal species.
*Important notice: 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.
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