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Environmental changes may expose primates to new pathogens. We investigated two zoonotic protozoa, Trypanosoma cruzi and Leishmania infantum, to which free-living primates may be exposed in Rio Grande do Sul State, Brazil. We evaluated risk factors associated with species, sex, age group, and biome in howler monkeys and assessed whether eight landscape metrics of forest fragments influence the monkeys' exposure to the pathogens. To do this, we tested serum from 96 howler monkeys (61 Alouatta caraya and 35 Alouatta guariba clamitans) by using an indirect hemagglutination assay (IHA) for T. cruzi and an indirect fluorescence antibody test (IFAT) for L. infantum. In the IHA for T. cruzi, 51% (49/96) were seropositive, with titers from 1:16 to 1:256. We found that A. caraya in the Pampa biome is more susceptible to exposure to T. cruzi than A. g. clamitans in the Atlantic Forest biome, but none of the landscape metrics, sex, or age were significantly related to exposure. No samples were positive for L. infantum. This may suggest that the conditions of the Pampa biome, where A. caraya occurs, are more conducive to the presence of Trypanosoma cruzi vectors, exposing the howler monkeys to infection. However, further serological tests are needed confirm our results. We hope our findings encourage future studies focused on identifying and isolating T. cruzi in primates and understanding the eco-epidemiological role that wildlife plays in transmitting and maintaining protozoa in the wild environment. [ABSTRACT FROM AUTHOR]