Molecular characterization of bovine Cryptosporidium isolated from diarrheic calves in the Sudan

Cryptosporidiosis is a common protozoan infection causing morbidity and mortality in young cattle and may be zoonotically transmitted to humans. So far, there is no data available on the presence of Cryptosporidium spp. in the Sudan. The aim of this study was to isolate, identify, and genotype Cryptosporidium oocysts sampled from diarrheic calves housed at different farms in three different municipalities in Khartoum State (Khartoum, Khartoum North, Omdurman). A total of 149 fecal samples were evaluated microscopically for the presence of Cryptosporidium oocysts using the modified Ziehl-Neelsen staining method and 87 (58.3%) samples tested positive. Positive and negative samples were further analyzed by nested PCR targeting the SSU rRNA region. Positive samples were subjected to restriction enzyme analysis of PCR amplicons (PCR-RFLP). Nested PCR identified Cryptosporidium DNA in 53 samples (35.5%); restriction digestion of the PCR products revealed the presence of C. parvum (73.5%), C. ryanae (13.2%), C. andersoni (7.5%), and C. bovis (1.8%). Species distribution was clearly related to age with C. parvum being the predominant species in dysenteric pre-weaned calves. Sequencing of three genes (SSU rRNA, COWP, and GP60) for three C. parvum isolates originating from the three different municipalities showed that all belong to C. parvum subtype family IId. Based on data obtained by GP60, sequencing the two C. parvum isolates from Khartoum and Omdurman represent subtype IIdA18G1, whereas oocysts isolated in Khartoum North belong to subtype IIdA19G1. The observed genotypes are zoonotic and thus C. parvum in calves is potentially a health risk to humans in Khartoum State, Sudan. To the best of our knowledge, this is the first reported attempt to characterize Cryptosporidium isolated from cattle in the Sudan.