为了解感染不同灵长类动物的隐孢子虫种类以及与感染人的人隐孢子虫(C.hominis)之间的遗传差异,本研究采用形态学和分子生物学方法对猴源隐孢子虫进行分离和鉴定。利用常规方法分离猴粪便中的隐孢子虫卵囊,通过改良抗酸染色和荧光显微镜观察对其进行形态学鉴定;并采用PCR方法扩增其卵囊壁蛋白(COWP)基因和18SrRNA基因,扩增产物克隆至pMD-18-T载体中,对阳性克隆进行测序并作进化树分析。结果表明:抗酸染色和荧光检查结果与所报道的人隐孢子虫的结果一致。PCR扩增产物经电泳检测,明显地出现554bp和370bp大小的片段,与预期结果一致;两种基因的序列分析结果显示该猴源隐孢子虫与C.hominis的相似性均为100%。由此可认为本次分离的隐孢子虫为C.hominis。
In this study,Cryptosporidium sp. was isolated from feces of rhesus (Macaca mulatta) and characterized by modified anti-acid fast staining and fluorescence observation. The Oocysts were shown to possess same morphological characteristics as that reported for C.hominis. Two target genes of COWP and 18S rRNA were amplified by PCR,cloned and sequenced,and the evolution tree was generated by Mega software. Both fragments shared 100 % sequence identities with C. hominis and their heredity distances was the nearest to that of C.hominis in evolution tree. It is concluded that the Cryptosporidium isolated from rhesus belong to C.hominis.