基于组学探究生防真菌Esteya vermicola吸引松材线虫的分子机理

The pine wood nematode (PWN) disease caused by PWN causes devastating ecological disasters and great economic losses to forestry in China and even in Asia, and it is of great significance to study the sustainable control technology of PWN. Esteya vermicola (Ev) attracts PWN by releasing volatile organic substances. The high infectivity and lethality of Ev against PWN make it a high potential biocontrol fungus for controlling PWN. The process of Ev attracting PWN is the key to effective prevention and control of pine wood nematode. However, the mechanism by which the fungus release chemicals to attract PWN is still unclear. Previous studies have found that there are intracellular symbiotic bacteria in Ev fungal spores and hyphae, so this project uses Ev symbiont as the research object. The key regulatory genes involved in the induction of PWN will be screened and identified based on genomics, transcriptomics, metabolomics, combined with qPCR technology, and these gene regulatory networks will be constructed to elucidate the mechanism of the Ev symbiont attracting PWN. The research results are not only important for the further study and use of the biocontrol fungi to prevent and control PWN in the future, but also help reveal the complex symbiotic-pathogenic relationship among the ternary "fungus-bacteria-nematode", and also show positive significance for coming up with better technology to control PWN.

由松材线虫引起的松材线虫病给我国乃至亚洲地区的林业造成毁灭性的生态灾害和重大经济损失，研究松材线虫的可持续控制技术意义重大。Esteya vermicola (Ev)通过释放挥发性物质吸引松材线虫，该菌对松材线虫表现出的高感染性和致死性使其成为防控松材线虫的高潜力生防菌。Ev吸引松材线虫的过程是该菌有效防控松材线虫的关键，但至今对该菌释放化学物质吸引松材线虫的机理尚不清楚。前期研究发现Ev孢子和菌丝内存在共生细菌，本项目以Ev共生体为研究对象，基于基因组学、转录组学和代谢组等多组学的研究与分析方法，结合qPCR技术，筛选和鉴定出产生吸引松材线虫化学物质的关键调控基因，构建其基因调控网络，阐明Ev共生体吸引松材线虫的分子机理。研究结果不仅对今后深入研究和利用该生防菌防控松材线虫具有重要意义，而且对揭示复杂的“真菌－细菌－线虫”三元的共生－致病关系、对松材线虫新防治技术的研发亦具有积极意义。

松材线虫（PWN）为我国第一大林业入侵有害生物，它引起的松材线虫病给我国的林业造成毁灭性的生态灾害和重大经济损失。伊氏线虫真菌Esteya vermicola对PWN具有高侵染能力和致死能力，该真菌具有应用于PWN生物防治的潜力。但真菌E. vermicola如何吸引PWN、及E. vermicola胞内共生细菌在吸引或致死PWN的过程中起到什么样的生物学作用尚不明确。本项目通过松材线虫诱导E. vermicola，研究E. vermicola的表型及转录变化，并对其胞内共生细菌进行基因组测序及比较基因组分析。研究发现，E. vermicola可能通过昆虫信息素合成途径释放挥发性物质吸引PWN。E. vermicola对线虫的响应不仅表现为信息素受体基因和信号转导途径相关基因表达量增加，而且为后续的粘附阶段和穿透线虫体壁阶段上调表达相关基因，为迅速杀死PWN做准备。共生细菌的基因组大小为3.96 Mb，GC含量为58.93%，注释到的基因4542个，初步鉴定为Phyllobacterium属。基于全基因组879个单拷贝基因构建的系统发育树表明，该共生细菌与P. myrsinacearum亲缘关系最近。基因家族分析表明，该内共生细菌中的多拷贝基因家族（42个）和多拷贝基因（91个）数量多，这可能与真菌细胞内共生细菌的生态适应性有关；此外，该内共生细菌基因组具有高比例的重复序列、多个电子传递链基因、及多个侵染和致死线虫的关键酶-枯草杆菌蛋白酶，这可能是特定环境条件下共进化的结果，适应其生态位并与真菌宿主协同杀死线虫。