Blastocystis spp. is a parasite that causes intestinal infection in humans and other animals. A few studies have been performed in Turkey on the distribution of Blastocystis in cattle. In this study, fecal samples were collected from 100 calves and subjected to analysis based on an SSU rRNA gene fragment. The overall prevalence of the disease was determined as 15% (15/100). This rate was 14.04% for females and 16.28% for males. In addition, three Blastocystis subtypes were identified: ST10, ST14, and novel subtypes ST25. To our knowledge, the ST25 subtype was reported with this study for the first time in Turkey. The nucleotide sequences (OM920832-OM920839) obtained in this study were deposited in GenBank. The results obtained will be useful for a better understanding of the epidemiology of Blastocystis spp., and its effects on public health.