基于SAFLP的我国常见索线虫科昆虫病原线虫亲缘关系分析

本研究建立并优化了索线虫科(Mermithidae)SAFLP体系，构建了我国常见索线虫科昆虫病原线虫6属11种/亚种的指纹图谱。采用EcoRⅠ和MseⅠ两种限制性内切酶单酶切，结果显示EcoRⅠ酶更适合作为索线虫科线虫SAFLP的内切酶。3个带有3个选择性碱基的EcoRⅠ引物进行扩增共得条带225个，片段大小为250～1 650 bp。通过NTsys-PC2.1软件计算了索线虫科6属11种/亚种的Nei-Li遗传距离(0.1980～034554)和相似系数，利用NTsys-PC2.1软件中的UPGMA方法构建其聚类图。索线虫科这6属11种/亚种从属级阶元可以分为两大类群：罗索属Romanomermis和八腱索属Octomyomermis聚为一支构成第一大类群；六索属Hexamermis与卵索属Ovomermis先聚在一起然后与多索属Agamermis聚在一起，再与两索属Amphimermis聚在一起构成第二大类群。在八腱索属和两索属中属内不同种/亚种间线虫采集地相距较近的其遗传距离较近。优化的索线虫科线虫SAFLP体系能够反映索线虫科线虫种属间的亲缘关系，并与形态学的研究结果基本相符，可以用于属种的分类和亲缘关系的研究。

Phylogenetic relationships among common species of Mermithidae (Nematoda) in China based on SAFLP

The method of SAFLP (single-enzyme amplified fragment length polymorphism) was optimized for Mermithidae in this study. The genetic relationships among 11 species or subspecies belonging to six genera of Mermithidae and common in China were detected using this method. Two kinds of restriction endonucleases were used in this study, and the results showed that EcoRⅠ was more suitable than MseⅠ for SAFLP analysis of Mermithidae. In total 225 bands (form 250 to 1 650 bp) were amplified using three EcoRⅠ primers each with three selective bases. Nei's similarity coefficients and genetic distance (0.1980－0.4554） were calculated by NTsys-PC2.1 software and the phylogenetic tree was constructed with the method of UPGMA. The results indicated that the eleven species/subspecies of Mermithidae could be distinguished with SAFLP markers and were grouped into two distinct clusters. Two major groups of genera are observed in the phylogenetic tree: Romanomermis + Octomyomermis; ((Hexamermis+Ovomermis) + Agamermis)+Amphimermis. Comparatively, the genetic distance of different species or subspecies, which are from closer area in the same genera, is smaller in Octomyomermis and Amphimermis. The result of this experiment is similar to that of the morphologic taxonomy system and provides further evidence for the studies of phylogenetic relationship among species or genera of Mermithidae. The results suggest that this optimized SAFLP is suitable for analysis of phylogenetic relationship and taxonomy among species of Mermithidae.