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

Petras Prakas,Agnė Baranauskaitė,Vytautas Januškevičius,Francesco Chiesa,Elena Servienė,Eglė Rudaitytė-Lukošienė,Živilė Strazdaitė-Žielienė,Dalius Butkauskas,Saulius Petkevičius

doi:10.1186/s13071-020-04473-9

Abstract

BackgroundSix 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.MethodsThe 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.ResultsUnder 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%).ConclusionsAlthough 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. Graphical

Highlights

Six Sarcocystis species are known to use cattle (Bos taurus) as the intermediate host, two of which, S. hominis and S. heydorni, are zoonotic
Current consensus is that that cattle may harbour up to six Sarcocystis species, namely S. bovifelis, S. bovini, S. cruzi, S. heydorni, S. hirsuta and S. hominis [4, 8,9,10,11]
Sarcocystis spp. from cattle have been molecularly characterised at the genes for 18S rRNA (18S rRNA), 28S rRNA (28S rRNA) and cytochrome c oxidase subunit I and at nuclear rDNA internal transcribed spacer 1 (ITS1) [8, 10, 11, 15]

Summary

Introduction

Six Sarcocystis species are known to use cattle (Bos taurus) as the intermediate host, two of which, S. hominis and S. heydorni, are zoonotic. Prakas et al Parasites Vectors (2020) 13:610 life-cycle and endogenous sporulation These parasites form sarcocysts mainly in the muscle tissues of the intermediate hosts and develop oocysts and sporocysts in the small intestine of the definitive hosts [1]. Three Sarcocystis species, S. suihominis, S. hominis and S. heydorni, are known to infect humans as definitive hosts [2,3,4]. The latter two species can be found in the muscles of cattle (Bos taurus), while humans may become infected by consuming raw/undercooked beef containing mature sarcocysts of S. hominis and S. heydorni. Studies have revealed that cox is the preferable target to differentiate taxonomically related Sarcocystis spp. whose intermediate hosts are such ruminants as cattle, sheep, goats, deer and others [19]

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