Diagnostic biomarkers in murine Cryptosporidiosis: dose- and age-related infection

Abstract

Increasing prevalence of Cryptosporidium raises the importance to explore different aspects of its infection. In the absence of reproducible in vitro culturing, animal model is the only experimental method to study Cryptosporidium. Our study evaluated Cryptosporidium infection using coproscopy, copro-antigen and copro-DNA for early detection of murine cryptosporidiosis. Hundred and forty albino mice (neonates and adult) were divided into two groups, control group received sterile PBS solution, and infected groups were inoculated with molecularly characterized Cryptosporidium parvum oocysts and further subdivided into three subgroups for infectious dose response detection. Mice fecal samples were collected every 4h on the first day and then daily and examined for fecal oocysts, copro-antigen and copro-DNA. Four mice from each subgroup were killed at 12, 24 and 48h post-infection (P-I), and their intestines were examined for cryptosporidial mucosal DNA. Cryptosporidium copro-antigen and copro-DNA were detected 4 and 8h P-I in infected neonatal and adult mice, respectively, and intestinal mucosal DNA was detected after 12h in both. Microscopy was able to detect oocysts 48h P-I. Inoculated C. parvum oocysts were recovered in feces of infected mice without genotypic changes. Neonate mice showed higher susceptibility for cryptosporidial infection than adults without statistical differences for the given infectious doses. Both copro-immunoassay and copro-nPCR assays can early detect Cryptosporidium infection; however, nPCR was able to identify Cryptosporidium species, making nPCR a reliable biomarker for early detection in murine model.