松材线虫致病相关基因的协同致病机理研究

Pine wilt disease (Bursaphelenchus xylophilus) is the most serious forest disease in our country at present, but the lagging pathogenesis research and disputed results of previous studies severely retarded the propulsion of pine wilt disease basic research and the scientific prevention and control measures’ out puts. In this study, we paid more attention on the terpene metabolic disorders and cavitation development which also were the typical pathological phenomenon of the pine wilt disease. We focused on the cytochrome p450 genes and cloned its full length cDNA and then analyzed its bioinformatics characteristic. After inoculated pine tree with RNAi nematode and expression proteins of these cytochrome p450 genes we researched the temporal specificity of these genes, the pathogenicity of pine wood nematode and the physiological response, histopathological changes of inoculated host plant. Based on these results, we illuminated the function of cytochrome p450 genes from pine wood nematode during the pathogenic process. Our study laid the foundation for the further study on pathological synergism mechanisms interaction with other pathogenic genes of pine wood nematode.

松材线虫病（Bursaphelenchus xylophilus）是我国目前最严重的森林灾害，其致病机理研究滞后和已有研究结果的争议，严重制约了松材线虫病的基础研究进展和有效防控技术产出。本研究在以松材线虫病病理学过程中的特征性事件-萜烯类物质代谢紊乱和空洞化为重点，聚焦松材线虫细胞色素氧化还原蛋白CYP450基因，在对其进行cDNA全长克隆和生物信息学分析的基础上，通过制备表达产物和RNA干扰研究松材线虫RNAi前后该基因表达的时间特征和致病力变化及寄主的生理响应、组织病理变化，阐明CYP450基因在松材线虫致病过程中的功能，为进一步揭示该基因与其它致病相关基因在松材线虫致病过程中的协同致病机理奠定基础。

松材线虫病的发生主要是寄主植物与病原线虫互作的过程。松材线虫入侵后，取食寄主薄壁细胞，诱导寄主防御反应，导致寄主萜烯类特别是单萜类次级代谢产物大量积累。萜烯类物质作为松树主要防御产物，对松材线虫具有毒害作用，为了成功在树体内定殖，松材线虫通过氧化还原解毒途径消除该类物质的毒害作用。CYP450基因是松材线虫氧化还原解毒途径中的关键基因。研究发现，松材线虫CYP450基因响应寄主萜烯类物质积累而大量表达。同时，松材线虫CYP450基因沉默后，松材线虫在体外萜烯胁迫条件下的活性和繁殖率降低。CYP450基因沉默松材线虫接种寄主松树后，寄主萜烯合成酶基因在第6天时表达量降低，萜烯类物质含量降低，寄主发病症状减轻，松材线虫致病性降低。因此，CYP450基因参与了松材线虫应对寄主体内萜烯胁迫的解毒作用，从而影响其致病性，是松材线虫重要致病基因。本研究成果为松材线虫致病机理深入研究提供了重要研究基础。