Molecular Identification of Parasitic Protozoa Sarcocystis in Water Samples

Abstract

Sarcocystis parasites are among the most common parasitic protozoa in farm animals. So far, the diversity of these parasites has been mainly studied in animal carcasses by morphological or molecular methods. Research on parasitic protozoa in environmental samples is scarce due to the lack of an appropriate methodology and low concentrations of parasites. For these reasons, there is a paucity of validated methods for Sarcocystis identification from environmental samples. Therefore, the present study aims to investigate various molecular methods for Sarcocystis parasite identification in water samples. In the present study, the sample volume, sporocysts isolation, and various conventional PCR were evaluated, and species-specific primers for the identification of different Sarcocystis species have been developed. Of the methods studied, based on data the most appropriate method for the identification of analyzed Sarcocystis spp. in water bodies is nested PCR, using species-specific primers targeting the cox1 gene. Sarcocystis DNA was detected in 111 out of 114 (97.4%) samples. This paper represents the first identification of S. bovifelis, S. cruzi, S. hirsuta, S. arieticanis, S. tenella, S. capracanis, S. bertrami, and S. miescheriana by PCR and sequencing in environmental water samples. Our pilot study is useful in developing techniques for the identification of Sarcocystis species from water samples.

Molecular Identification of Protozoan Sarcocystis in Different Types of Water Bodies in Lithuania

“…It was observed that, having selected adequate PCR primer pairs, detection rates of farm animals infecting Sarcocystis spp. in water samples were associated with those observed in the muscles of the intermediate host [ 10 , 14 , 15 , 16 , 17 ]. However, in our earlier research, the prevalence of S .…”

“…tenella ) using farm animals (cattle, sheep, goats, horses, and pigs) as intermediate hosts. Since the cox1 gene is considered the most appropriate for the identification of selected Sarcocystis parasites [ 11 , 12 , 13 ], primers targeting this gene were selected [ 10 ]. It was observed that, having selected adequate PCR primer pairs, detection rates of farm animals infecting Sarcocystis spp.…”

“…miescheriana and S . tenella identified in water samples was significantly lower [ 10 ]. Thus, 10 new primers ( Table 1 ; highlighted in bold) were designed for the identification of these species.…”

“…Even so, only a few studies were conducted to test for Sarcocystis spp. in water [ 8 , 9 , 10 ]. The aim of our previous study was to assess different sample preparation and common PCR methodologies for identifying Sarcocystis species in water.…”

“…miescheriana , and S . tenella ) was detected in 111 of 114 (97.4%) water samples using nested PCR targeting cox1 gene [ 10 ]. However, Sarcocystis species occurrence rates in different areas and types of water bodies were not compared.…”