Canine piroplasmoses, caused by Babesia spp., Theileria spp. and Rangelia vitalii, are emerging vector-borne diseases with a worldwide distribution, transmitted by ticks. The aim of this study was to determine the prevalence and perform molecular characterization of piroplasmids in domestic dogs from Asuncion city, Paraguay. Blood samples were taken from 384 domestic dogs from Asuncion city, Paraguay. DNA was purified from dog blood samples and submitted to nested PCR assays for piroplasmids (18S rRNA) and sequenced for identification and phylogenetic analysis. Overall piroplasmid prevalence in dogs from Paraguay was 6% (23/384 [CI 95% = 3.6-8.4%]). Phylogenetic studies showed that Babesia vogeli was the most prevalent species (91% [21/ 23]), followed by Theileria equi (4% [1/23]) and Rangelia sp. closely related to R. vitalii (4% [1/23]). Babesia vogeli, T. equi and Rangelia sp. circulate among domestic dogs from Asuncion city, and are described for the first time in Paraguay.