Wild felids are thought to share parasites with domestic cats. However, little is known of the coccidian parasites of wild felids. We investigated the presence of Sarcocystis spp. in tissues of 6 species of 90 Neotropical small felids killed in road accidents in the state of Rio Grande do Sul, Brazil by using microscopic and molecular techniques. Formalin-fixed tissues from 28 felids were examined, and Sarcocystis felis-like sarcocysts were detected in 4 wild cats (2 Puma yagouaroundi and 2 Leopardus guttulus). By transmission electron microscopy, sarcocysts from a P. yagouaroundi were identical to S. felis from domestic cats in the USA. Direct sequencing of PCR amplicons resulted the unambiguous sequences of the ITS-1 region from 18 of the 31 PCR positive wild cats; 5 sequences from each P. yagouaroundi, and Leopardus geoffroyi, 4 sequences from L. guttulus, and 2 sequences from each Leopardus wiedii, and Leopardus colocolo. Sequences analysis of ITS-1 region revealed the highest identiy (97-99%) with that of previously describe isolates of S. felis from domestic cats in the USA and identified them as S. felis. Tissues of 1 Leopardus pardalis tested by PCR and histology were negative. The phylogenetic relationship indicated that S. felis is quite different to species which employ opossums as their definitive host. This is the first report of S. felis infection in small wild felids from Brazil.