基于cox1基因的棕熊锡兰钩虫分子鉴定与种系发育分析

Molecular Identification and Phylogenetic Analysis of cox1 Gene of Ancylostoma ceylanicum from Boar

本次研究旨在利用线粒体细胞色素I(cox1)部分基因(pcox1)对长沙市动物园棕熊粪便分离的钩虫进行种类鉴定,并分析其基因多态性。用PCR技术扩增从棕熊粪便分离的钩虫长沙分离株的pcox1基因,并对其基因序列进行分析。结果显示7条钩虫分离株的pcox1序列片段大小均为423 bp,种内遗传差异为0.0%~0.5%,与GenBank中收录的锡兰钩虫(Ancylostoma ceylanicum)分离株同源性为93.8%~94.8%,与其他钩虫(犬钩虫、巴西钩虫、十二指肠钩虫、猫钩虫和美洲钩虫等)序列相似性低于88.7%;系统发育分析显示,本次研究获得的棕熊钩虫分离株与已知锡兰钩虫聚为一支,与其他钩虫相隔较远。结果表明,长沙市动物园棕熊粪便分离的钩虫为锡兰钩虫,线粒体pcox1基因在锡兰钩虫种内相对保守,与其他钩虫种类序列差异较大,可以作为熊锡兰钩虫分子鉴定及遗传变异研究。

The objective of this study was to identify hookworm species isolated from brown bear feces using partial sequence (pcox1) of mitochondrial cytochrome I (cox1) gene for polymorphism analysis.The mitochondrial pcox1 sequences of hookworm isolates were amplified using PCR and analyzed for their phylogenetic relationship.Results showed the sequence fragments of pcox1 gene were 423 bp in length and the genetic variations within the species was 0.0%-0.5%.The nucleotide homology the cox1 sequences was 93.8%-94.8% between our hookworm isolates and Ancylostoma ceylanicum in GenBank but lower than 88.7% with other hookworm isolates including Ancylostoma caninum,Nippostrongylus brasiliensis,Ancylostoma tubaeforme,etc).The phylogenetic analysis showed that the hookworm isolates in this study were clustered into one branch with those A.ceylanicum isolates available in GenBank,which was far away from other hookworm branches.The results showed that the hookworm isolates from brown bear feces were identified with A.ceylanicum.In addition,the mitochondrial pcox1 gene was relatively conserved within the species but the genetic difference among these species was more obvious,which could be used as useful information for molecular identification and genetic variation of A.ceylanicum.