Characterization of Cryptosporidium parvum in human and animal feces by single-tube nested polymerase chain reaction and restriction analysis.

A DNA isolation and purification method is described that produced DNA free of inhibitory substances in 148 of the 159 analyzed fecal samples. The polymerase chain reaction (PCR) product from a sensitive single-tube nested PCR that amplifies a part of an oocyst protein was used to characterize Cryptosporidium parvum genotypes by a simple restriction analysis. Genotype 1 was solely detected in human-derived oocysts, genotype 2 was present in both animal and human-derived oocysts. The ratio between both genotypes in humans in The Netherlands varied markedly between samples obtained during a period of augmented cases of cryptosporidiosis in the western part of the country and randomly selected samples from gastroenteritis patients. Sequence analysis of a 581-bp fragment from the nested PCR product revealed 12 nucleotide substitutions between the two genotypes. Sequences from isolates in each genotype group were identical.