Sarcocystis spp. are the cyst forming protozoan parasites that prevalent in livestock all around the world. In the presented work, we examined 40 macroscopic and 40 microscopic sarcocysts from Khouzestan and Lorestan provinces, south-western Iran, utilizing PCR-RFLP based on amplification of 18S rRNA gene. Using AvaI, HindII, TaqI and EcoRI restriction enzymes the results represented Sarcocystis gigantea in both macroscopic and microscopic cysts. This result supports the importance of molecular investigations on characterization of Sarcocystis species when we aimed to assess the reliable control and preventive programs.