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

Nader Pestechian,Faham Khamesipour,Reza Kalantari,Mahsa Esmaeilifallah,Rasool Jafari,Mohammad Keshtkar,Hossein Ali Yousefi,Alessandro Di Cerbo

doi:10.1155/2021/5562517

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.

Highlights

Sarcocystis species is among the most prevalent cyst-forming protozoan parasites with worldwide distribution in various hosts [1, 2]
Various species of Sarcocystis are identified in sheep, such as Sarcocystis gigantea and S. medusiformis, which are nonpathogenic and cause macroscopic cysts; the cats are definitive hosts for them
Four (80%) samples out of 5 macrocysts were found in the diaphragm, and one (20%) of them was in the esophagus. e PCR reaction using the mentioned primers confirmed an expected band on the agarose gel, which is a section comprising 623–625 consensus nucleotides (Figure 1). e results of the molecular method revealed that 83.33% of the samples had Sarcocystis spp., and the highest rate of infection was detected in diaphragm samples (93.33%), followed by 83.33% and 73.33% in the heart and esophagus tissues, respectively

Summary

Introduction

Sarcocystis species is among the most prevalent cyst-forming protozoan parasites with worldwide distribution in various hosts [1, 2]. More than 200 species of Sarcocystis with different clinical symptoms were reported all over the world [5]. Livestock animals, such as sheep, are among the most infected hosts with global distribution [6]. Various species of Sarcocystis are identified in sheep, such as Sarcocystis gigantea and S. medusiformis, which are nonpathogenic and cause macroscopic cysts; the cats are definitive hosts for them. Some species, such as S. tenella and S. arieticanis should be considered pathogenic causing microscopic cysts, and canids are the definitive host for them [7, 8]. Mature sarcocysts of various Sarcocystis spp. can be differentiated by distinct phenotypic features, such as form, size, cyst wall thickness, and organization of villar

Methods

Results

Conclusion