鲁道夫对盲囊线虫C新种的ITS rDNA序列测定及种系发育关系分析

ITS rDNA sequencing and phylogenetic analysis of the new species Contracaecum rudolphii C(Nematoda:Anisakidae)

以鲁道夫对盲囊线虫C种为研究对象,对其rDNA的内转录间隔区(ITS)序列进行PCR扩增、测序和序列分析,以ClustalⅩ1.81程序对序列进行比对,然后用MEGA4.0程序的MP法和NJ法及PUZ-ZLE4.1程序的ML法构建鲁道夫对盲囊线虫A、B、C种以及北方对盲囊线虫的种系发育关系。结果显示,鲁道夫对盲囊线虫C种的ITS-1,5.8 S,ITS-2序列大小分别为451 bp、127 bp和277 bp,鲁道夫对盲囊线虫3个种与北方对盲囊线虫ITS序列种内差异小,种间差异明显。种系发育关系分析发现,这3种方法(NJ法、MP法和ML法)构建的种系发育树的拓扑结构基本一致,其中C种和A种聚在一起,构成一支,再和B种聚在一起;北方对盲囊线虫单独成一支。研究结果证明鲁道夫对盲囊线虫C种确为一新种。

The internal transcribed spacers （ITS） of Contracaecum rudolphii C were amplified, sequenced and compared with those of C. rudolphii A,C. rudolphii B,and C. septentrionale available in Gen- Bank^TM, and followed by phylogenetic reconstruction. The sequences were aligned using the Clustal X 1.81 ,MP and NJ trees were reconstructed using the softwares MEGA version 4.0 ,and ML tree was reconstructed using PUZZLE version 4.1. The results revealed that the ITS sequences of C. rudolphii C was 855 bp in length,including 451 bp for ITS 1,127 bp for 5.8S and 277 bp for ITS 2,and that the average intraspecific variations were comparatively lower and the inter-specific differences were significantly higher. Topologies of 3 phylogenetic trees inferred by different methods（NJ, MP or ML） were identical or similar. These phylogenetic trees showed two distinct clades：C, rudolphii and C. septentrionale. In the C. rudolphii clade,C, rudolphii A was genetically closer to C. rudolphii C than to C. rudolphii B.