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Between 2013 and 2018, autochthonous infections by <jats:italic>T. cruzi<\/jats:italic> were suspected in three European wolves (<jats:italic>Canis lupus<\/jats:italic>) and an orange dwarf porcupine (<jats:italic>Coendou spinosus<\/jats:italic>) from <jats:italic>Funda\u00e7\u00e3o Parque Zool\u00f3gico de S\u00e3o Paulo<\/jats:italic> (FPZSP), Brazil, current <jats:italic>Coordenadoria de Fauna Silvestre<\/jats:italic> (CFS), which is inserted in a remnant of the Atlantic rainforest inside one of the biggest and most populous municipality of the world. This study aims to detect <jats:italic>T. cruzi<\/jats:italic> infections in captive mammals, triatomines and free-living opossums from surrounding areas of FPZSP\/CFS.<\/jats:p>\n<\/jats:sec>\n<jats:sec>\n<jats:title>Methodology\/Principal Findings<\/jats:title>\n<jats:p>Blood samples from captive and free-living mammals from surrounding areas were collected for parasitological (direct examination and culture), serological (IFAT) and molecular diagnosis using Nested-PCR <jats:italic>18SrDNA<\/jats:italic> followed by DNA sequence analysis. Triatomines (<jats:italic>Panstrongylus megistus<\/jats:italic>) found in FPZSP\/CFS were also examined by culture of the digestive tract and PCR. <jats:italic>Trypanosoma cruzi<\/jats:italic> infection was detected in 35.7% (n\u2009=\u200960\/168 \u2013 106 captive and 62 free-living) of the mammals that belonged to nine different families. From captive mammals, positive <jats:italic>T. cruzi<\/jats:italic> serology was observed in 29.6% (n\u2009=\u200927\/91). Twenty-six positive hemocultures were obtained, from which parasite isolation was achieved in 69.2%, while positive PCR was observed in 40% of them, including nine free-living individuals that were also positive in hemoculture. Of 28 individuals in which <jats:italic>T. cruzi<\/jats:italic> characterization was successful, 89.3% were genotyped as DTU TcI, 7.2% as TcII and 3.5% as TcI\/TcII mixed infection. Besides, 29 of 30 collected triatomines were infected, and infection by <jats:italic>T. cruzi<\/jats:italic> DTU TcI was confirmed in 16 of them.<\/jats:p>\n<\/jats:sec>\n<jats:sec>\n<jats:title>Conclusions\/Significance<\/jats:title>\n<jats:p>The confirmed autochthonicity of at least 68.9% of cases demonstrates that captive mammals from FPZSP are immersed in the <jats:italic>T. cruzi<\/jats:italic> enzootic cycle that involves the vector species <jats:italic>P. megistus,<\/jats:italic> and the reservoir hosts <jats:italic>C. spinosus<\/jats:italic> and <jats:italic>Didelphis aurita<\/jats:italic> from the wild surrounding areas.<\/jats:p>\n<\/jats:sec>","DOI":"10.1371\/journal.pntd.0013055","type":"journal-article","created":{"date-parts":[[2025,4,28]],"date-time":"2025-04-28T17:47:34Z","timestamp":1745862454000},"page":"e0013055","update-policy":"https:\/\/doi.org\/10.1371\/journal.pntd.corrections_policy","source":"Crossref","is-referenced-by-count":3,"title":["Trypanosoma cruzi (Chagas, 1909) transmission among captive wild mammals, triatomines and free-living opossums from surrounding areas in the S\u00e3o Paulo Zoological Park, Brazil"],"prefix":"10.1371","volume":"19","author":[{"given":"Suelen Sanches","family":"Ferreira","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marina Galv\u00e3o","family":"Bueno","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Carolina V. 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