Molecular identification of four Sarcocystis species in cattle from Lithuania, including S . hominis , and development o
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RESEARCH
Molecular identification of four Sarcocystis species in cattle from Lithuania, including S. hominis, and development of a rapid molecular detection method Petras Prakas1*, Živilė Strazdaitė‑Žielienė1, Vytautas Januškevičius1,2, Francesco Chiesa3, Agnė Baranauskaitė1, Eglė Rudaitytė‑Lukošienė1, Elena Servienė1, Saulius Petkevičius2 and Dalius Butkauskas1
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 mus‑ cle 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 indi‑ vidually under the light microscope. Keywords: Cattle, Sarcocystis hominis, Trypsin digestion, Molecular identification, cox1, 18S rRNA gene
*Correspondence: [email protected] 1 Nature Research Centre, Vilnius, Lithuania Full list of author information is available at the end of the article
Background Protozoan parasites of the genus Sarcocystis (Apicomplexa: Sarcocystidae) infect mammals, birds and reptiles. The genus is characterised by an obligatory two-host
© The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License,
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