Background: Many species/genotypes of Cryptosporidium infect a wide range of hosts including humans, domestic and wild animals and are one of the most important pathogens causing diarrhoea in children and immunocompromised hosts. The most common species include C.hominis that is anthroponotic and C.parvum that is both zoonotic and anthroponotic.
Objectives: To differentiate species/genotypes of Cryptosporidium isolates from human faecal samples by SSU rRNA based PCR-RFLP and to subtype C. hominis and C. parvum by sequence analysis of GP60 region.
Methods: Cryptosporidium oocysts were detected in stool samples by bright field microscopy (100X) of Ziehl-Neelsen stained faecal smears and further genotyped by SSU rRNA based PCR-RFLP with enzymes SspI and VspI. C. hominis and C. parvum isolates were sub-typed by sequence analysis of nested PCR amplified GP60 gene. For subtype families, sequences were searched for similarity by BLAST analysis and aligned with reference sequences retrieved from GeneBank by multiple sequence
alignment (ClustalW, BioEdit220.127.116.11). Subtypes within each subtype family were named by manual counting of tandem repeats in the poly-serine tract of GP60 gene.
Results: Out of 52 Cryptosporidium isolates, RFLP analysis revealed 39 isolates as C. hominis and 13 as C. parvum and while 6 mixed infections with C. maleagridis and C.parvum or C.hominis could be detected by sequencing. GP60 based sequencing of C.hominis and C. parvum divided them into 8 subtype families and 17 subtypes.
Conclusion: Anthroponotic C. hominis and C.parvum are more prevalent in North India suggesting the importance of human to human transmission.