松材线虫内寄生真菌Esteya vermicola 防治松材线虫病的机理研究

Pine wood nematode is the most serious forest pest. It is classified as the dangerous forest pest by many countries in the world because of its serious damage to pine trees and prevention difficulties. For the past few years control of pine wilt disease using endoparasitic fungus, Esteya vermicola has got good results. However, the control mechanism of pine wilt disease using E. vermicola is not clear, and it has not been interpreted systematically. This project studies on the host plant living pine tree, dead pine tree, media sawyer beetle which were sprayed by conidia of E. vermicola, and they will be investigated using cultivation of pine tree segments, checking-tube of additional nutrition and DNA detection method. Firstly, the control mechanism of E. vermicola against pinewood nematode in dead pine trees after spraying conidia of E. vermicola, and the change of pinewood nematode in pupa of sawyer beetle after spraying conidia will be revealed. Secondly, the conidia of E. vermicola and pinewood nematode carried by sawyer beetles caught in the spraying area will be investigated. Thirdly, the prevention mechanism of E. vermicola against pinewood nematode in living pine trees will be investigated, and the evidence of conidia in living pine tree and its killing pinewood nematode will be checked. Therefore, the control mechanism of pine wilt disease using E. vermicola will be revealed in the project, and the key technique of biocontrol of pine wilt disease will be established.

松材线虫是目前最为严重的森林有害生物，由于危害严重、防治困难而受到世界各国的高度重视。近年来利用松材线虫内寄生真菌Esteya vermicola防治松材线虫病取得了较好的效果，但是其防治机理不清晰。本项目拟对松材线虫的寄主生存松树、枯死松树、其传播媒介松褐天牛喷洒真菌, 利用样本切片培养法、外源营养管检测法和DNA检测法对树体内该真菌进行检测，揭示喷洒一定浓度的孢子防治枯死松树体内松材线虫，并进一步控制枯死木中化蛹期松褐天牛所携带松材线虫的机理；喷洒孢子防治林间松褐天牛体内所携带的松材线虫，并调查真菌在其体表生长的状况；揭示预先喷洒真菌孢子到生存松树，当松树受到松材线虫感染时体内真菌预防松材线虫的机理，并收集生存松树体内真菌生存并寄生松材线虫的证据，从而系统的阐明该真菌林间防治松材线虫病的机理，确立关键防治技术，为松材线虫病的林间真菌喷洒防治打下理论基础。

松材线虫是目前最为严重的森林有害生物，由于危害严重、防治困难而受到世界各国的高度重视。近年来利用松材线虫内寄生真菌Esteya vermicola防治松材线虫病取得了较好的效果，但是其防治机理不清晰。通过样本切片培养法、外源营养管检测法和DNA分子检测法成功找到了E. vermicola真菌在寄主生存松树、枯死松树及其传播媒介松褐天牛的证据。得到了E. vermicola真菌在树体内捕食松材线虫并萌发孢子的照片，喷洒E. vermicola真菌孢子悬浮液后孢子在松褐天牛体表生长产生大量菌丝和孢子的照片，以及喷洒E. vermicola真菌孢子悬浮液后用外源营养管检测法清晰地检测到健康松树体内的新月型孢子，用实时荧光定量PCR检测到喷洒E. vermicola真菌孢子悬浮液后在健康松树的证据。通过喷洒一定浓度的孢子防治枯死树体内松材线虫，降低松褐天牛携带松材线虫的数量，进而降低松褐天牛啃食松枝而造成的松树死亡率，实验结果表明喷洒松材线虫致死木段所羽化出的松褐天牛对松树幼苗致死率为0%，而对照组致死率为100%；通过温室松苗实验、松材线虫病疫区树干注射实验、林间喷洒实验等揭示了当松树收到松材线虫感染时体内真菌预防松材线虫病的机理；通过六年时间的观察，树干注射固体孢子培养基、线虫携带孢子的松树生存率为50%，注射液体孢子的松树生存率为60%，然而对照组在第一年全部死亡；通过松材线虫和内寄生真菌E. vermicola在初始浓度、培养时间的实验，揭示了初始浓度、培养时间影响松材线虫和内寄生真菌E. vermicola捕食和被捕食的关系。本实验通过对松材线虫的寄主生存松树、枯死松树、其传播媒介松褐天牛喷洒E. vermicola真菌，以及调查初始孢子浓度、喷洒时间等实验，成功揭示了松材线虫内寄生真菌E. vermicola防治松材线虫病的机理，为松材线虫病的林间真菌喷洒打下了理论基础。