The order Chiroptera is the second largest group of mammals with bats being identified as reservoir of several viral zoonoses, although, little is known about their role in other groups of pathogens, including hemotropic Mycoplasma spp. To date, hemoplasma species have been found infecting several species of bats with high genetic diversity between 16S rRNA gene sequences. On this study, we aimed to identify the occurrence and characterize 16S and 23S rRNA genes of hemoplasma species in four bats species (Artibeus lituratus, Carollia perspicillata, Sturnira lilium and Sturnira tildae) from forest fragments in Paraná State, southern Brazil, using PCR-based assays. Spleen tissue samples were collected, DNA extracted and further screened by a pan‑hemoplasma PCR assay. All samples consistently amplified the mammal endogenous gapdh gene. One out of 15 (6.66%; 95% CI: 0.2-31%) bats tested positive for hemotropic Mycoplasma sp. by the PCR assay targeting the 16S rRNA gene. Sequencing of the 16S rRNA gene fragment from the hemoplasma-positive bat showed 99.14% identity with hemotropic Mycoplasma sp. detected in Sturnira parvidens from Belize. Sequencing of the 23S rRNA gene fragment from the hemoplasma-positive bat showed 86.17% identity with ‘Candidatus Mycoplasma haemosphiggurus’ detected in orange-spined hairy dwarf porcupines (Sphiggurus villosus) from Southern Brazil.