基于形态和分子特征的三孔线虫科系统分类研究

Nematodes of the family Tripylidae de Man, 1876 are primitive in the phylum Nematoda. Some species could be applied as bioindicators in monitoring environmental changes of soil and some could be used as biological control agents of plant parasitic nematodes. The in-depth systematic study of this family has theoretical values and application potentials. However, problems related to the taxonomy of this family remain unresolved, such as the ambiguous definition of some genera and species, the presence of numerous synonyms and homonyms. In terms of research on this group, there is still a gap between China and worldwide. In the past several years, the applicants have conducted research on four genera of the family morphologically and molecularly, which have built up the basis for further studies. In this project, we propose to perform a systematic survey on soil samples to study the biodiversity of Tripylidae in China. The proposed approaches will include collection of morphological data by means of optical and scanning electronic microscopes, and molecular data by obtaining sequences of cytochrome oxidase c subunit 1 (COI), small subunit (SSU) and D2D3 segment fragments of large subunit (LSU) rRNA genes. The obtained data will be used to identify the species of Tripylidae in China and construct cladistic and molecular phylogenetic trees to resolve the relationships among the Tripylidae in China and worldwide. The aims of the project are to describe new taxa and its distribution of Tripylidae in China, clarify the confusion of species’ names and revise the controversial taxonomic genera and species, reveal the phylogenetic relationships among genera and species. Therefore the expected results will fill the gap of Tripylidae research in China, resolve the ambiguous genera and species, and importantly also provide a basic knowledge for the application of Tripylidae as potential biological monitoring of soil environment and biological control of plant parasitic nematodes.

三孔线虫科是线虫门中较古老的一个类群，有些种类可作为土壤环境的指示生物，有些则可作为植物寄生线虫的生防资源；开展该科的系统分类研究，具有理论价值和应用前景。国内外有关研究目前存在着分类单元界定模糊，遗留大量同物异名和异物同名现象，而我国的相关研究基础至今仍十分薄弱。申请人近年来结合形态学和分子生物学，对该科四个属的种类进行了分类研究。本项目拟在前期研究基础上，通过在全国范围内系统采集样品，采用形态学和分子鉴定相结合的方法，开展中国三孔线虫科的系统分类研究；并应用支序分类学和分子系统发育学相结合的方法，对其进行系统演化关系研究。从而明确中国三孔线虫科的种类及分布；澄清历史遗留的疑难分类单元，厘清同物异名及异物同名；探明属间和种间系统演化关系，推断其系统演化过程，完善该科的分类系统。本项研究将为线虫分类、土壤环境生物监测及生防资源挖掘提供理论基础和科学依据。

三孔线虫科是线虫门中较古老的一个类群，有些种类可作为土壤环境的指示生物，有些则可作为植物寄生线虫的生防资源；国内外有关研究目前存在着分类单元界定模糊，遗留大量同物异名和异物同名现象，而我国的相关研究基础薄弱。开展该科的系统分类研究，具有理论价值和应用前景。本项目的主要成果如下：. （1）对全国29省市的植物根围2834份土壤样品进行分离，并采用形态学和分子鉴定相结合的方法将其鉴定到4属12种，包括2个新种和5个中国新记录种，分别为三孔属Tripyla paraquatica sp. n.，T. setifera和T. aquatica；Tripylella sinensis sp. n.；小三孔线虫属Tripylina ixayocensis，T. puxianensis和T. zhejiangensis；以及Trischistoma属中Trischistoma equatoriale，T. minor，T. monohystera，T. pellucidium和T. taiguensis。其中Tripylina zhejiangensis是优势种群，占检测总群体的19%。并编制了三孔线虫科中国种类的鉴定检索表。. （2）获得了以上12个种的SSU、D2-D3 LSU和COI基因序列，并首次报道了Tripyla aquatica和Trischistoma minor的SSU序列，Trischistoma equatoriale和Trischistoma monohystera的D2-D3 LSU序列，以及Tripyla setifera，Tripylina ixayocensis，Tripylina zhejiangensis，Trischistoma minor和Trischistoma taiguensis的COI基因序列，丰富了三孔线虫科鉴定的分子数据。. （3）基于SSU和D2-D3 LSU构建了分子系统发育树，并基于支序分析方法构建了分支系统发育树，对三孔线虫高阶分类单元及其属级分类阶元进行了探讨。. 本项目的完成极大地丰富了我国三孔线虫科的本底资料，深化我国该类群物种多样性的认识，弥补我国在土壤自由线虫系统分类研究上的不足。