Sarcocystis cruzi infection in free-living European bison (Bison bonasus bonasus L.) from the Białowieża Forest, Poland - A molecular analysis based on the cox1 gene

Abstract

European bison are susceptible to a range of pathogens which may influence their health, and hence, to ensure their protection, it is essential to provide effective monitoring of potential exposure. This study presents the first molecular confirmation of Sarcocystis cruzi infection in European bison based on PCR amplification of the cytochrome c oxidase subunit I (cox1) gene. A sample of heart tissue taken from one fifteen-year-old European bison cow was examined by light microscopy for the presence of heart sarcocysts. The genomic DNA isolated from any identified sarcocysts was subjected to PCR to amplify cox1 gene sequences, and the obtained amplicons were sequenced by Sanger dideoxy sequencing. Two partial cox1 sequences were obtained; they were identified as S. cruzi and deposited in the GenBank™ database under the accession numbers MW490605 and MW490606. BLAST analysis found them to demonstrate the closest similarity to S. levinei (MH255771-MH255779 and KU247874-KU247884), sharing an identity of 93.14-93.8 %. This is the first report to identify sarcocysts isolated from heart tissue of infected European bison living in the Białowieża forest to species level using cox1 analysis. Our findings confirm that the European bison is a natural intermediate host for S. cruzi. As such, coordinators of future conservation programmes should consider the impact of these diseases on reintroduced animals.

Keywords: European bison; Sarcocystis cruzi; cox1 gene.

Graphical abstract

Fig. 1

Overview of consistent nucleotide differences between two cox1 gene sequences of S. cruzi MW490605 and MW490606. Numbers above and below the sequences refer to nucleotide positions in the two GenBank sequences used in the comparison. Positions where the two sequences were identical are signified with Dots (.).

Fig. 2

Phylogenetic tree for selected Sarcocystis cruzi isolates and selected members of Sarcocystis species found in Bovidae based on partial sequences of cox1 using the maximum parsimony method. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) are shown next to the branches. GenBank accession numbers are listed next to the taxon names. The country of origin of S. cruzi isolates are listed in brackets.