Trypanosomatids are obligate flagellated parasites, with the genera Leishmania and Trypanosoma acting as etiological agents of significant diseases such as leishmaniasis and Chagas disease. Although ecological studies have increasingly highlighted the role of bats as potential reservoirs of these parasites, the diversity of trypanosomatids in urban bat populations remains poorly understood. This study investigates the occurrence and diversity of Trypanosomatidae in bats from Mangabeiras Municipal Park (MMP), an urban park in Belo Horizonte, Minas Gerais, Brazil, a region of ecological interest due to the prior detection of Leishmania in sand flies. A total of 56 bats representing seven species were captured, and 149 biological samples (blood, tissues, and organs) were analyzed using NNN/LIT culture medium. Contamination was reported in 32.2% of the samples, while 67.8% yielded negative results with no growth of trypanosomatids. Detection of trypanosomatids was achieved using the V7V8 Nested-PCR technique, revealing positive results in nine bats: Artibeus lituratus (Leishmania infantum, Trypanosoma sp. Neobat 3), Anoura caudifer (Trypanosoma sp. Neobat 4), Carollia perspicillata and Glossophaga soricina (Leishmania infantum), Sturnira lilium (Trypanosoma sp. Neobat 3), and Platyrrhinus lineatus (mixed infection with Leishmania infantum and Leishmania braziliensis). The integrity of the extracted DNA was confirmed through the amplification of cytb and gamma-actin genes. By expanding knowledge of trypanosomatid diversity in urban bats, this study highlights the ecological and epidemiological relevance of bats as hosts and underscores the need for targeted surveillance to assess their role in pathogen transmission dynamics.