Molecular identification of four Sarcocystis species in cattle from Lithuania, including S. hominis, and development of a rapid molecular detection method

Abstract

Background Six Sarcocystis species are known to use cattle (Bos taurus) as the intermediate host, two of which, S. hominis and S. heydorni, are zoonotic. There is a need for a method that will enable rapid identification of the Sarcocystis species in cattle. Methods The diaphragm muscles of 102 cattle from Lithuania were examined for the presence of Sarcocystis spp., using two different methods for species identification. Individual sarcocysts were isolated from squash preparations of the diaphragm muscle under the light microscope, followed by genetic characterisation of excised cysts using sequence analysis of the 18S rRNA (18S rRNA) and cytochrome c oxidase subunit I (cox1) genes. The same cattle muscle samples were digested and species-specific PCR analyses targeting cox1 were developed to identify the Sarcocystis isolates to the species level. Results Under the light microscope, sarcocysts were detected in 87.3% of animals, and Sarcocystis infection was verified in all digested samples. Three species, namely S. cruzi (n = 20), S. bovifelis (n = 23) and S. hirsuta (n = 6), were identified by DNA sequence analysis of isolated sarcocysts. Based on sequence analysis of cox1, the level of genetic variability depended on Sarcocystis species and geographical location. Four Sarcocystis species, S. cruzi (96.1%), S. bovifelis (71.6%), S. hirsuta (30.4%) and S. hominis (13.7%), were confirmed in the digested samples. In individual samples, the most common finding was two species of Sarcocystis (44.1%), followed by three species (26.5%), a single species (24.5%) and four species (4.9%). Conclusions Although examination of tissue preparations under the light microscrope did not detect any sarcocysts belonging to S. hominis, this species was identified in the digested samples subjected to a cox1-specific PCR analysis. These results demonstrate the need for effective molecular diagnosis techniques to detect Sarcocystis spp., which may be present at a lower prevalence and not detectable among the limited number of sarcocysts identified individually under the light microscope.

Molecular detection of cattle Sarcocystis spp. in North-West Italy highlights their association with bovine eosinophilic myositis

“…Considering the high detection of S. cruzi , followed by S. bovifelis and S. hominis , the prevalence data reported in our study show most resemblance to those of Hungary [ 22 ] and The Netherlands [ 26 ], while in Germany and Lithuania a higher prevalence of S. hirsuta is reported, ranging from 6.6 to 30.4%, respectively [ 27 , 28 ].…”

The Role of Mustelids in the Transmission of Sarcocystis spp. Using Cattle as Intermediate Hosts

“…bovifelis and humans are DH for S . hominis [ 19 ]. Thus, our results indicate that mustelids might be involved in the transmission of Sarcocystis spp.…”

“…hirsuta was confirmed in 30.4% of the samples and S . hominis was observed in 13.7% of the samples [ 19 ]. Canids are DH for S .…”

“…hirsuta and S . bovifelis are transmitted via felids [ 19 ]. The Eurasian lynx ( Lynx lynx ) is the only wild member of the felids in Lithuania [ 9 ].…”

“…in cattle from Lithuania has been recorded [18]. By performing trypsinization of the diaphragm muscles and species-specific PCR targeting the cox1 (mitochondrial gene encoding subunit 1 of cytochrome c oxidase), S. cruzi was identified in 96.1% of the samples, S. bovifelis was detected in 71.6% of the samples, S. hirsuta was confirmed in 30.4% of the samples and S. hominis was observed in 13.7% of the samples [19]. Canids are DH for S. cruzi, humans are DH for S. hominis, whereas S. hirsuta and S. bovifelis are transmitted via felids [19].…”