Molecular identification of Cycospora spp. using multiplex PCR from diarrheic children compared to others conventional methods

ABSTRACT

Cyclospora organisms named C. cercopitheci, C. colobi and C. papionis were identified in stool samples from several primates. They were morphologically indistinguishable from C. cayetanensis but genetically different. In the present work, Cyclospora infection was diagnosed among 140 diarrheic children with three conventional diagnostic methods and was confirmed using nested PCR. The possibility of infection with not only C. cayetanensis but the three Cyclospora species of primates was identified by multiplex PCR among all cyclosporiasis patients diagnosed by different methods. The results showed that Cyclospora was detected in 25 [17.8%], 31 [22.2%], 32 [22.9%] and 35 [25%] patients using modified Kinyoun stain, auto-fluorescent characteristics, sporulation process of the oocysts and nested PCR respectively. The sensitivity, specificity, accuracy, positive predictive value, negative predictive value and Kappa test were calculated in relation to nested PCR. The single nucleotide polymorphisms [SNPs] in the 18S rRNA gene of C. cayetanensis were identified in 35 cyclosporiasis patients and only one patient had the possibility of human co-infection with primates Cyclospora species [C. cercopitheci, C. colobi and C. papionis] and C. cayetanensis by appearance of a 361-bp. Scanning electron microscopy proved no morphological differences could be detected among Cyclospora oocysts isolated from this patient