Fecal Sample Surveillance of the Wildlife Virome in Central Italy: Insights from the Foreste Casentinesi National Park

Wildlife can act as both a reservoir and a sentinel for emerging pathogens, but surveillance is often constrained by difficulties in obtaining samples without disturbing animals. This study explored the viral diversity of wild mammals inhabiting the Foreste Casentinesi National Park (Central Italy) using non-invasive fecal sampling. From 2021 to 2022, 99 fecal samples from several species were collected and analyzed by PCR and metagenomic next-generation sequencing. Of 26 pools examined, 10 (38.5%) tested positive for at least one viral target. Astroviruses were the most frequently detected, found in deer, foxes, wolves, small mustelids, and porcupines. Several sequences showed low similarity to known strains, suggesting divergent or novel viral lineages. Metagenomic analysis also identified members of <i>Circoviridae</i>, <i>Anelloviridae</i>, and <i>Picobirnaviridae</i>. While none of these virus families are currently recognized as major zoonotic agents, their widespread occurrence in wildlife and domestic animals underscores the importance of continued surveillance to better assess their ecological roles, host range, and potential implications for both animal and human health. These results provide new insights into the virome of European wildlife, including the first reports of some viruses in certain species. Overall, our study demonstrates that non-invasive surveillance is a valuable tool for monitoring ecosystem health and supports a One Health approach to early detection of viral threats.