Occurrence of Leishmania spp. in Roadkilled Wild Mammals on Highways in the State of São Paulo, Brazil

BACKGROUND: Leishmaniasis undergoes geographic expansion in the Neotropical regions, driven by ecological and socioeconomic factors that contribute to urban and peri-urban outbreaks. Roadkilled wild mammals represent a potential source of eco-epidemiological data, and polymerase chain reaction (PCR)-based detection provides a sensitive tool for the surveillance of Leishmania spp. METHODS: This study investigated the occurrence of Leishmania spp. in 40 roadkilled wild mammals collected along highways in the central-western and northern coastal regions of São Paulo State, Brazil between 2020 and 2022. Necropsies were performed on all specimens, and ear skin samples were collected for molecular detection of Leishmania spp. using primers targeting kinetoplast DNA (kDNA). Positive samples were further analyzed for species identification by PCR-restriction fragment length polymorphism (RFLP) targeting the hsp70 gene. Restriction patterns were compared with reference strains to confirm Leishmania species identity. RESULTS: Leishmania spp. kDNA was detected by molecular diagnosis in skin tissue samples from several roadkilled wild mammals collected in the state of São Paulo. Subsequent species-level identification using hsp70 PCR-RFLP detected Leishmania infantum chagasi in a maned wolf (Chrysocyon brachyurus) and Leishmania amazonensis in a raccoon (Procyon cancrivorus). CONCLUSIONS: Wild mammals could be exposed to distinct Leishmania species across diverse ecological contexts, and examining roadkilled animals, combined with the molecular detection of parasites, proved to be an effective tool for passive Leishmania surveillance, highlighting the need for integrated investigations within a One Health framework.