保定地区奶牛源隐孢子虫的分离与基因鉴定  

Isolation and genotype identification of Cryptosporidium in Baoding

作　　者：曹文博[1] 刘立元[1] 任志军[1] 唐欣浩 王雪伟[1] 陈福星[1] 包永占[1] 秦建华[1] 赵月兰[1] 

机构地区：[1]河北农业大学动物医学院,河北保定071001

出　　处：《黑龙江畜牧兽医》2017年第8期35-38,286,共5页Heilongjiang Animal Science And veterinary Medicine

基　　金：河北省现代农业产业技术体系"奶牛产业创新团队建设项目奶牛疾病防控技术研发与示范"(HBCT2013080204)

摘　　要：为了解保定地区奶牛源隐孢子虫感染情况,从当地3个奶牛场随机采集145份奶牛粪便经饱和蔗糖溶液漂浮后直接镜检和抗酸染色后镜检,阳性样品采用PCR方法扩增18S rRNA基因,同时将扩增出的目的片段进行克隆和序列分析,并将分离株与其他11个隐孢子虫参考株的18S rRNA序列进行比对和遗传距离比较。结果表明:有10份粪便样本为隐孢子虫阳性,感染率为6.9%(10/145),不同奶牛场、年龄段奶牛隐孢子虫感染率有显著性差异(P<0.05)。10份粪便样品采用PCR方法扩增后有7份为18S rRNA基因阳性,扩增出的18S rRNA部分基因片段长528 bp。保定地区隐孢子虫分离株扩增的基因序列与牛源隐孢子虫18S rRNA基因序列(Gen Bank登录号为HQ179571)同源性最高,确定保定地区分离到的奶牛源隐孢子虫为牛隐孢子虫。In order to identify the ~nfectious situation of Cryptosporidium, 145 fresh fecal samples collected from cow of 3 dairy farms in Baoding were examined as subjects. All the samples were microscopically examined after floating saturated sucrose solution and acid - fast staining. The microscopy - positive samples were amplified by PCR based on 18S rRNA, and the amplified fragments were cloned and sequenced. The results showed that Cryptosporidium oocysts were preliminarily detected from the fecal samples of 10 dairy tattles with the infective rate of 6.9% （ 10/ 145 ）, the significant differences were observed between the infection rate of different dairy farms and ages（ P 〈 0.05 ）. 7 samples were ampli- fied by PCR based on 18S rRNA fromten microscopy - positive samples and the amplified fragments were 528 bp. The results of 18S rRNA se- quences comparisons demonstrated the highest identity between the two isolates and the Cryptosporidium bovis （HQ179571）. It was concluded that the iffolates of bovine Cryptosporidium from Baeding were identified as Cryptosporidium bovis.

关 键 词：奶牛 牛隐孢子虫 18S RRNA基因 PCR 序列 分子进化分析 

分 类 号：Q78[生物学—分子生物学] S823.91[农业科学—畜牧学]