Cryptosporidium spp. in wild, laboratory, and pet rodents in china: prevalence and molecular characterization

Abstract

To understand the prevalence of Cryptosporidium infection in rodents in China and to assess the potential role of rodents as a source for human cryptosporidiosis, 723 specimens from 18 rodent species were collected from four provinces of China and examined between August 2007 and December 2008 by microscopy after using Sheather's sugar flotation and modified acid-fast staining. Cryptosporidium oocysts were detected in 83 specimens, with an overall prevalence of 11.5%. Phodopus sungorus, Phodopus campbelli, and Rattus tanezumi were new reported hosts of Cryptosporidium. The genotypes and subtypes of Cryptosporidium strains in microscopy-positive specimens were further identified by PCR and sequence analysis of the small subunit rRNA and the 60-kDa glycoprotein (gp60) genes. In addition to Cryptosporidium parvum, C. muris, C. andersoni, C. wrairi, ferret genotype, and mouse genotype I, four new Cryptosporidium genotypes were identified, including the hamster genotype, chipmunk genotype III, and rat genotypes II and III. Mixed Cryptosporidium species/genotypes were found in 10.8% of Cryptosporidium-positive specimens. Sequence analysis of the gp60 gene showed that C. parvum strains in pet Siberian chipmunks and hamsters were all of the subtype IIdA15G1, which was found previously in a human isolate in The Netherlands and lambs in Spain. The gp60 sequences of C. wrairi and the Cryptosporidium ferret genotype and mouse genotype I were also obtained. These findings suggest that pet rodents may be potential reservoirs of zoonotic Cryptosporidium species and subtypes.

FIG. 1.

Phylogenetic relationships among various Cryptosporidium species/genotypes from rodents in China and some known Cryptosporidium species and genotypes, as inferred by a neighbor-joining analysis of the partial (∼735 positions in the final alignment) SSU rRNA sequences based on distances calculated using the Tamura-Nei 93 model and adjusted with a gamma distribution (shape parameter, 1). Bootstrap values (in percentages) above 50 from 1,000 pseudoreplicates are shown for both the neighbor-joining (the first value) and maximum-parsimony analyses (the second value). ns, node with bootstrap values lower than 50%. Sequences from brown rats in Japan (23) are not included because of the lack of data from the 5′ end of the fragment under analysis. The multiple-sequence alignment of partial SSU rRNA sequences used in the generation of the phylogenetic tree is presented elsewhere (see the supplemental material).

FIG. 2.

Phylogenetic relationship of gp60 nucleotide sequences of C. parvum, C. wrairi, and the Cryptosporidium ferret genotype and mouse genotype I obtained in this study and known Cryptosporidium subtype families, as inferred by a neighbor-joining analysis based on distance calculated using the Kimura two-parameter model. Bootstrap values greater than 50% from 1,000 pseudoreplicates are shown. The name of each subtype family starts with the Cryptosporidium species or genotype designation, with C. hominis, C. parvum, C. meleagridis, C. fayeri, rabbit genotype, horse genotype, C. wrairi, ferret genotype, and mouse genotype I represented by I, II, III, IV, V, VI, VII, VIII, and IX, respectively. The sequence of the C. parvum subtype family IIk (AB237137) is not shown because of its short length.