Molecular and Microscopic Investigation of Sarcocystis Species Isolated from Sheep Muscles in Iran

Abstract

Sarcocystis species is a genus of cyst-forming parasites infecting both humans and animals globally. Some of these species cause clinical and subclinical diseases in the host and may lead to economic losses. This study was carried out to identify the distribution patterns of Sarcocystis spp. in slaughtered sheep based on the digestion method and PCR-RFLP in Isfahan, the center of Iran. In total, 150 fresh muscle samples (30 hearts, 60 esophagi, and 60 diaphragms) were investigated by naked eye observation and then scrutinized based on the digestion method. To this end, pepsin and HCl were used to observe the Sarcocystis parasite via a light microscope. The PCR was carried out to intensify a fragment of the 18S rRNA gene. Afterward, the PCR products were exposed to digestion by endonuclease TaqI, HindII, EcoRI, and AvaI. Consequently, the results of RFLP were confirmed by sequencing, and the phylogenetic placement of all species was analyzed. Through the examination by the naked eye, 5/150 (3.33%) macroscopic cysts were found in the samples. With the tissue digestion and microscopic examination, 116 (77.33%) samples were positive for Sarcocystis spp.; however, 125 (83.33%) samples were positive with PCR. Moreover, the results of sequence analysis on macrocysts and microcysts showed that 4% and 96% of the species belonged to S. gigantea and S. tenella, respectively. According to the results of the current study, sarcocystosis caused by S. tenella are highly prevalent among sheep in the Isfahan region. Due to the high prevalence of Sarcocystis infection in the world and Iran, the development of disease control and prevention policies in sheep would be essential, and changing attitudes in the way of keeping livestock from the traditional type to the industrial method is recommended in this regard.