Molecular identification of the Cryptosporidium deer genotype in the Hokkaido sika deer ( Cervus nippon yesoensis ) in Hokkaido, Japan

Abstract

The protozoan Cryptosporidium occurs in a wide range of animal species including many Cervidae species. Fecal samples collected from the Hokkaido sika deer (Cervus nippon yesoensis), a native deer of Hokkaido, in the central, western, and eastern areas of Hokkaido were examined by polymerase chain reaction (PCR) to detect infections with Cryptosporidium and for sequence analyses to reveal the molecular characteristics of the amplified DNA. DNA was extracted from 319 fecal samples and examined with PCR using primers for small-subunit ribosomal RNA (SSU-rRNA), actin, and 70-kDa heat shock protein (HSP70) gene loci. PCR-amplified fragments were sequenced and phylogenetic trees were created. In 319 fecal samples, 25 samples (7.8 %) were positive with SSU-rRNA PCR that were identified as the Cryptosporidium deer genotype. Among Cryptosporidium-positive samples, fawns showed higher prevalence (16.1 %) than yearlings (6.4 %) and adults (4.7 %). The result of Fisher's exact test showed a statistical significance in the prevalence of the Cryptosporidium deer genotype between fawn and other age groups. Sequence analyses with actin and HSP70 gene fragments confirmed the SSU-rRNA result, and there were no sequence diversities observed. The Cryptosporidium deer genotype appears to be the prevalent Cryptosporidium species in the wild sika deer in Hokkaido, Japan.

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