【摘要】 目的对来源于哈尔滨某奶牛场一头3月龄奶牛的一个隐孢子虫分离株进行了分子鉴定。选择隐孢子虫基因分型常用的18SrRNA和actin基因位点,设计引物,用巢氏PCR方法分别扩增出约840bp和1066bp目的片段。PCR产物经双向测序后用ClustlX、DNAstar和PHLIP软件进行序列分析。结果表明在18SrRNA基因位点上,此分离株与报道的牛源C.ryanae分离株(DQ871345、AY587166、EU410344)同源性最高,达到100%。在actin位点,此分离株和美国牛源C.ryanae分离株(EU410345)和隐孢子虫鹿样基因型(AY741309)同源性也为100%。种系发育分析显示所获得的分离株在两个基因位点上均和C.ryanae处于同一进化分支。这些分析表明此次分离获得的隐孢子虫分离株为C.ryanae。鉴定结果为研究我国牛源隐孢子虫种类分布以及评价其公共卫生意义提供了重要参考。更多还原

【Abstract】 One Cryptopsoridium isolate from a 3 month-old calf in a dairy cow farm of Harbin was identified by molecular method,in which the common genes of 18S rRNA and actin used in Cryptosporidium genotyping were selected,the primers were designed,and the approximate 840 bp and 1066 bp fragments of these two genes were amplified respectively by nested PCR.The PCR products were sequenced by two directions,and then the sequences were analyzed by using the soft wares of ClustlX,DNAstar,and PHLIP.The experimental result showed that in the 18SrRNA gene,there was the highest identities compared to other reported isolates of cow-derived C.ryanae(DQ871345,AY587166,and EU410344),with 100% of similarity.Likewise,at actin locus,the present isolate also had 100% similarity with an American cow-derived C.ryanae isolate(EU410345) and a deer-like genotype isolate(AY741309).Phylogenetic analysis indicated that the present isolate shared the same evolution branch in the two gene loci.These analyses demonstrated that the present isolate belonged to C.ryanae.These results provide a significant reference for the study on the species distribution and assessment in the public health significance of Cryptosporidium in cow milk in China.更多还原