Accurate analysis of prevalence of coccidiosis in individually identified wild cranes in inhabiting and migrating populations in Japan.

Eimeria gruis and E. reichenowi cause coccidiosis, a major parasitic disease of cranes. By non-invasive molecular approaches, we investigated the prevalence and genetic characterization of pathogens in two Japanese crane habitats; one is Hokkaido inhabited by the endangered red-crowned crane, and the other is Izumi in Kyushu where populations that consist mainly of vulnerable hooded and white-naped cranes migrate in winter. The non-invasively collected faecal samples from each wintering population were first subjected to host genomic DNA-targeted analyses to determine the sample origin and avoid sample redundancy. Extremely high prevalence was observed in the Izumi populations (> 90%) compared with the Hokkaido population (18-30%) by examining 470 specimens by microscopy and PCR-based capillary electrophoresis (PCR-CE), using genetic markers in the second internal transcribed spacer (ITS2). Correspondence analysis of PCR-CE data revealed differences in community composition of coccidia between hooded and white-naped cranes. 18S rRNA and ITS2 sequences were determined from single oocysts excreted by red-crowned and hooded cranes. Phylogenetic analysis of 18S rRNA suggested that E. reichenowi was polyphyletic while E. gruis was monophyletic. Together with PCR-CE data, these results indicate different host specificity among the E. reichenowi type. Our data suggest that E. reichenowi comprises multiple species.