One Health genomic and sero-surveillance for SARS-CoV-2 in captive wildlife in South Africa suggests continued risk for reverse zoonoses post pandemic (2021–2025)

Abstract Background SARS-CoV-2 infection has been reported in susceptible domestic, farmed, and wildlife species following exposure to infected humans. In South Africa, ecotourism and close human–animal contact may increase the risk for reverse zoonotic transmission, but animal surveillance remains limited. Methods We conducted case-based investigations of 92 animals from Gauteng, Limpopo, Western Cape and Mpumalanga Provinces in South Africa between 2021 and 2025 for SARS-CoV-2 infections. Cases were selected based on respiratory signs, unexplained death, or opportunistic sampling of high-risk species. Respiratory specimens were tested for SARS-CoV-2 RNA by RT‒PCR, and serum for SARS-CoV-2 IgG antibodies. Specimens were additionally screened for other coronaviruses, influenza A virus, and paramyxoviruses. RT‒PCR-positive specimens were subjected to genomic sequencing and phylogenetic analysis. Humans with close contact with positive cases were also tested. Results SARS-CoV-2 RNA was detected in 4/92 animals (4.3%), including three lions from Gauteng in 2021 and an exotic jaguar from a private zoo in Limpopo in 2023. The jaguar developed severe systemic and respiratory disease and subsequently died. Serological evidence of exposure was found in 6/45 animals (13.3%), including three lions, one lynx, one puma, and one mongoose. The lynx and puma were asymptomatic and originated from the same zoo as the three lions, whereas the mongoose presented respiratory signs and close human contact at a wildlife reserve in Mpumalanga. A One Health investigation identified one RT‒PCR-positive veterinary student among zoo and veterinary personnel with potential contact with the animal. Partial genome sequencing classified SARS-CoV-2 from the jaguar as Omicron BA.2, which clustered with lineages circulating in the United States and was distinct from locally circulating variants. No additional respiratory pathogens were detected. Conclusion These findings indicate that captive and free-ranging wildlife with close human contact remain at risk for SARS-CoV-2 reverse zoonosis. The detection of Omicron BA.2 in a jaguar with severe disease and a fatal outcome highlights the ongoing threat to susceptible species, possibly through ecotourism. Serological evidence suggests that infections may be more widespread than those detected by RT‒PCR alone. Integrated One Health surveillance is essential for detecting spillover events, monitoring variants, and reducing risk at human–animal interfaces.