Sracocystis cruzi in cattle (Bos taurus) and Sarcocystis poephagicanis in yaks (Bos grunniens) are morphologically indistinguishable. However, the relationship of the two parasites is still un-clear. Here, muscular tissues of the two domestic animals collected from abattoirs in China were examined for sarcocysts of S. cruzi and S. poephagicanis. The sarcocysts isolated from the samples were processed for light microscopy (LM), transmission electron microscopy (TEM), and DNA analysis. Sarcocysts of S. cruzi and S. poephagicanis were found in 405 of 950 (42.6%) cattle and 304 of 320 (95.0%) yaks, respectively. By LM and TEM, the sarcocysts of the two parasites showed similar morphologically characteristics. The thin-walled sarcocysts had hair-like protrusions on the surface. Ultrastructures exhibited the primary cyst wall contained irregularly folded, hirsute or bone-like protrusions. Four genetic markers, 18S rDNA, 28S rDNA, mitochondrial cox1, and apicoplast rpl6 of the two parasites were sequenced and analyzed. The sequences of the four loci presented an interspecific identity of 97.9−98.6%, 97.2−98.1%, 89.5−90.4%, and 96.9−97.2%, respec-tively. Phylogenetic analysis using 28S rDNA and cox1 sequences indicated that both species were placed into a group encompassing Sarcocystis spp. in ruminants with canid as known or putative definitive hosts. Sarcocystis cruzi and S. poephagicanis should represent separated species, and cox1 and rpl6 was suitable for distinguishing between them.