Toxoplasma gondii, a globally distributed protozoan, poses risks to wildlife and public health, with domestic cats playing a significant role in its dissemination. The Valdivian temperate rainforest in southern Chile, known for its biodiversity and endemism, faces threats from human activities and from invasive species such as domestic cats. We evaluated T. gondii seroprevalence in two sympatric introduced carnivores, an intermediate host, the American mink (Neogale vison), and a definitive host, the domestic cat (Felis catus), and we determined landscape variables linked to exposure for both species and identified high T. gondii exposure using ecological niche modeling. Seroprevalence for T. gondii in 180 American minks and 140 domestic cats exceeded 60% in both species, indicating significant exposure to T. gondii within the study area. Spatial analysis highlighted a relationship between low-altitude areas, characterized by floodable forests and slower river flow, and elevated T. gondii exposure in both species. Areas with agricultural activity and higher human and cattle densities, such as croplands and grasslands, exhibited an increased risk of T. gondii exposure. Parasite exposure was higher in the Tolten and Valdivia River basins compared to the other four basins studied. These findings emphasize the importance of considering landscape factors, including altitude, land use, and human activities, when assessing T. gondii exposure patterns. The identified landscape variables associated with exposure and the mapping of high-risk areas can guide public health and conservation management plans.

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