Genetic investigation of GPI anchored Bd37 orthologs inBabesia divergensgroup and use of recombinant protein for ecological survey in deer.

The Glycosylphosphatidylinositol (GPI) anchored protein group has great potential as an excellent immunodiagnostic marker, because of its high expression and necessity for parasite survival.Babesia divergens /B. capreoligroup includes parasites with confirmed or possible zoonotic potential to cause human babesiosis. In this study, we investigated ortholog ofBd37, a GPI-anchored major merozoite surface protein ofB. divergenssensu stricto, in the Asia lineage of theB. divergens /B. capreoligroup. From two genomic isolates from sporozoites/sporoblasts, threeBd37gene variants, namely Bd37 JP-A, JP-B, and JP-C, were isolated with 62.3% -64.1% amino acid sequences identity. Discriminative blood direct PCR revealed that JP-A was exclusively encoded in all parasites infecting wild sika deer examined (n=22). While JP-B and JP-C genes were randomly detected in 12 and 11 specimens, respectively. Recombinant JP-A-based ELISA showed an overall positive rate of 13.9% in deer in Japan from north (Hokkaido) to south (Kyushu islands) (24 prefectures, n=360). This positive rate was twice as high as that examined by 18S rRNA-based PCR (6.8%). Antibodies against recombinant JP-B and JP-C were also evident in the deer. This study demonstrated that the presence of three orthologs in theBd37gene family in Asia lineage and identified JP-A as an informative marker for serological surveys in Japan. This is the first report that diagnostic antigen ofBabesiaparasite was identified by a comprehensive analysis of genetic polymorphisms from a various developmental stage in host and vector.