[Identification of species of Cyclospora isolates from patients by the PCR-based diagnostic methods]

Cyclospora cayetanensis is an intestinal coccidian parasite and known as a human pathogen causing watery diarrhea. Recently, Cyclospora organisms, morphologically indistinguishable from C. cayetanensis, were detected from the several species of primates, and three new species named, C. cercopitheci, C. colobi, and C. papionis, have been proposed to the isolates on the basis of the genetic differences. The infectivity of these species to humans is strictly unknown, and there is a possibility of infection with not only C. cayetanensis but also the species from primates among patients. Therefore, it is necessary for the accurate diagnosis of Cyclospora infection to distinguish among the Cyclospora species. In the present study, we identified species of Cyclospora isolates from patients by the PCR-based diagnostic method. Since the sequence of Cyclospora 18S ribosomal RNA gene generated with the primary pair CYC1 FE and CYC4RB is found to be variable among Cyclospora species, we applied the PCR-direct sequencing using above primers to identify species of the isolates. Consequently, the diagnostic fragment was amplified by the PCR in all isolates, and the sequence of the PCR product obtained from each isolate was completely identical to that of C. cayetanensis. Therefore, we identified the isolates from patients as C. cayetanensis. On the basis of the results obtained in the present study, it is supposed that the PCR-direct sequencing using the primer pair CYC1FE and CYC4RB is a useful tool for the distinction among Cyclospora species.