羊毛尾线虫凉山州分离株的线粒体cox1和Cyt b基因的序列测定及种系发育分析

Sequencing and phylogenetic analysis of mitochondrial cox1 gene and Cyt b gene of Trichuris ovis isolates from Liangshan

应用PCR技术对分离自四川省凉山州12个山羊源羊毛尾线虫分离株的线粒体cox1基因部分序列(pcox1)和Cyt b基因部分序列(pCyt b)进行扩增和分析。结果显示,所获得的羊毛尾线虫pcox1和pCyt b序列长度分别为843bp和909bp,种内变异分别为0.0%~0.5%(7个变异位点)和0.0%~0.1%(1个变异位点),分别检测到7个单倍型和2个单倍型,平均单倍型多样性分别为0.773和0.409,核苷酸多样性分别为0.001 74和0.000 45。构建的种系发育树均显示,12个山羊源羊毛尾线虫凉山州分离株聚在同一小分支,能与其他毛尾线虫相鉴别;不同宿主来源的羊毛尾线虫中国株均聚在同一分支,而羊毛尾线虫西班牙株单独聚为一支,推测不同地方羊毛尾线虫分离株可能存在隐藏种或不同的基因型。结果表明,羊毛尾线虫cox1和Cyt b基因序列种内变异小,种间差异较大,故可作为分子标记用于羊毛尾线虫的种间鉴定;羊毛尾线虫凉山州分离株存在一定的遗传变异和遗传多样性,且cox1基因的多样性水平高于Cyt b基因。本研究结果为羊毛尾线虫的分子分类、群体遗传结构和分子流行病学调查等的进一步研究奠定基础。

The partial sequences of mitochondrial cox1 gene and Cyt b gene of 12 Trichuris ovis samples isolated from goats in Liangshan,Sichuan province,were amplified and sequenced.The results showed that the partial sequences of cox1 gene and Cyt b gene of T.ovis were 843 bp and909 bp,respectively.Sequence variations between the pcox1 of T.ovis isolated from Liangshan were 0.0%-0.5%(with 7 variable sites),while variations in pCyt b were 0.0%-0.1%(with 1 variable site).7 haplotypes and 2 haplotypes were detected in pcox1 and pCyt b with the average haplotype diversity being 0.773 and 0.409 respectively,the nucleotide diversities were 0.001 74 for pcox1 and 0.000 45 for pCyt b.The phylogenetic trees showed that all 12 T.ovis isolated from Liangshan were clustered in the same clade,which can be identified with other Trichuris.The populations of T.ovis from China isolated from different hosts clustered together,but separated from the isolates of Spain.The results suggested the existence of two cryptic species or genotypes from different geographical regions.The results indicated that cox1 gene and Cyt b gene can be used as genetic markers to identify T.ovis,and certain genetic variations and genetic diversities were presented among T.ovis isolated from Liangshan,the diversity of cox1 gene was higher than Cyt b gene.The results of this study provide the foundation for further research on molecular classification,population genetics and epidemiological survey of T.ovis.