Parasites are key players in ecosystems, influencing population sizes and food webs, yet the impact of environmental factors on their diversity is not well understood. The Atlantic rainforest in Brazil, particularly the Parque Estadual das Fontes do Ipiranga (PEFI), exemplifies a biodiversity hotspot facing significant deforestation, housing diverse animal species such as the synanthropic Brazilian common opossum (Didelphis aurita), which serves as a reservoir for multiple zoonotic pathogens. In this study, we investigated parasite diversity, abundance, prevalence, and richness in free-living D. aurita in the PEFI, São Paulo, Brazil. From January 2015 to January 2017, 101 fecal samples of D. aurita were collected in two areas of PEFI, at the Instituto de Pesquisas Ambientais (IPA) and the Parque de Ciência e Tecnologia (Cientec), and analyzed using three different parasitological methods. In total, 99% of the samples were positive for at least one parasite. The most prevalent parasite belonged to the order Strongylida (82%), followed by Cruzia sp. (77%), the latter having a significantly higher prevalence at IPA. In contrast, Acanthocephala showed greater prevalence at Cientec. Co-infections were common, with some individuals harboring up to seven different parasites. Our findings reveal significant parasite diversity in the D. aurita population at PEFI, including both helminths and protozoan trophozoites, some of which are reported for the first time in this host species. Further research is essential for accurate species identification of the observed parasites.