松材线虫VAP基因和CYP450基因协同致病机理研究

Pine Wilt Disease (Bursaphelenchus xylophilus) is one of the most serious forestry disease in China. The lagging pathogenesis research severely retarded the out puts of scientific prevention and control measures. The invasion of pine wood nematode (PWN) stimulated the defense reaction and induced the accumulation of terpenes within host tree. And then, PWN alleviated the stress of terpenes through the detoxification pathway to ensure that the nematode successfully propagated within host tree. The PWN continuously induced the accumulation of terpenes within host tree which caused pine trees extensive cavitation and wilt due to lack of water. In our study we 1) to determine the key pathogenic genes of PWN by studying correlation between stimulating genes (VAP) and detoxification genes (CYP450) of PWN and terpene metabolism of host trees; 2) studied the function of key pathogenic genes of PWN by RNAi and prokaryotic expression technology; 3) compared the dynamic changes of stimulating genes (VAP), detoxification genes (CYP450) and terpene metabolism in host trees under different treatments and their role during the cavitation process. The function of the key pathogenic gene of pine wood nematode; 3) By comparing the dynamic changes of the pine wood nematode challenge gene (VAP), detoxification gene (CYP450) and host terpene under different treatment conditions and its role in the cavity process. These results revealed the synergistic pathogenesis of stimulating genes (VAP) and detoxification genes (CYP450) in PWN during the pathogenic process. Our study provided a new idea and method for the study of the pathogenesis of PWN and provided a scientific basis for the prevention and treatment of the disease.

松材线虫病（Bursaphelenchus xylophilus）是我国目前危害最为严重的森林灾害之一，其致病机理研究的滞后严重制约了该病有效防控技术产出。松材线虫入侵后，激发寄主防御反应，诱导寄主产生萜烯类物质，同时通过解毒作用消除萜烯类物质的抑制作用，保证线虫在树体内成功繁殖，大量繁殖的线虫持续诱导寄主萜烯类物质大量积累，最终引起松树产生大面积空洞而缺水死亡。本研究1）通过研究松材线虫激发基因（Vap）和解毒基因（CYP450）与寄主萜烯类物质代谢之间的相关性确定松材线虫关键致病基因；2）通过RNAi和原核表达技术研究松材线虫关键致病基因的功能；3）通过比较不同处理条件下松材线虫激发基因（Vap）、解毒基因（CYP450）和寄主萜烯类物质动态变化及其在空洞化过程中的作用。阐明松材线虫激发基因（Vap）和解毒基因（CYP450）协同作用机制，为松材线虫病致病机理的研究提供新思路和新方法。

松材线虫病是我国最具毁灭性的森林灾害之一，该病致病机理仍存在争议。本研究通过气象色谱、实时定量、MicroCT、冷冻扫描技术等方法深入研究了松材线虫致病机理。研究发现，松材线虫入侵后，寄主α-蒎烯、β-蒎烯、柠檬烯、长叶烯、月桂烯和石芹烯等萜烯类物质大量积累。其中α-蒎烯含量最高，其含量为总的萜烯类物质的75%。松树发病程度越严重，萜烯类物质含量越高。另外，寄主松树病程发生情况与侵染早期α-蒎烯、β-蒎烯、柠檬烯和长叶烯比例有关，健康松树α-蒎烯：β-蒎烯：长叶烯：柠檬烯为1:0.17:0.12:0.1，松材线虫入侵后，发病早期α-蒎烯：β-蒎烯：长叶烯：柠檬烯为1:0.05:0.15:0.15，发病后期为1:0.1:0.05:0.05。发病早期寄主发病情况越轻，β-蒎烯含量越高。松材线虫致病过程中， Bx-cyp33c9和Bx-vap-1基因具有协同致病作用，且与寄主萜烯类物质存在显著相关性，vap-1基因可激发寄主萜烯类物质大量积累，而Bx-cyp33c9基因响应寄主萜烯类物质大量积累而上调表达，从而保证松材线虫在树体内大量繁殖，持续诱导萜烯类物质积累，诱导寄主空洞化现象持续发生，最终导致寄主水分运输紊乱而死亡。本研究深入揭示了松材线虫致病机理，为该病防控策略产出提供了理论基础，寄主萜烯类物质含量和比例的发现也为该病早期检测提供了新靶标。