Comparison of three diagnostic methods in the diagnosis of cryptosporidiosis and gp60 subtyping of Cryptosporidium parvum in diarrheic calves in Central Anatolia Region of Turkey

Abstract

The aim of this study was to compare three diagnostic methods for the diagnosis of cryptosporidiosis and to detect subtypes of Cryptosporidium parvum by sequences analyses of gp60 gene in diarrheic calves in several herds in Konya province located in Central Anatolia Region of Turkey. Fecal samples were collected from a total of 194 pre-weaned calves (n=158, .15 days old, and n=36, 15 to 40 days old), with diarrhoea. For comparative diagnosis, all samples were examined by modified Ziehl-Neelsen staining of fecal smears for the presence of oocyst, nested PCR-RFLP of SSU rRNA and TaqMan qPCR for the detection of Cryptosporidium DNA. A total of 92 (47.4%) and 104 (53.6%) out of the examined samples were found positive by microscopic examination and molecular tools, respectively. The diagnostic sensitivity and specificity of microscopic identification were determined as 88.5% and 100.0%, respectively compared to molecular assays. Cryptosporidium parvum was the only detected species in all positive samples by species-specific qPCR and nested PCR-RFLP assays. Species identifications were further confirmed by sequence analyses of the SSU rRNA PCR products. There was no statistically significant difference in C. parvum prevalence between early pre-weaned calves and calves older than 15 days. The sequence analyses of the gp60 gene of C. parvum isolates revealed a one subtype IIaA13G2R1 belonging to zoonotic family IIa in diarrheic calves.