嗜线虫致病杆菌抑制灰葡萄孢的效应

The inhibitory effect of Xenorhabdus nematophilus on Botrytis cinerea

摘要:

[背景] 灰葡萄孢（Botrytis cinerea）是引起葡萄采后病害的主要病原菌之一，严重影响葡萄的贮期和品质，给葡萄产业带来极大损失。利用拮抗微生物抑制采后病原菌生长已逐渐成为防治葡萄采后灰霉病的重要手段。[目的] 利用昆虫病原线虫共生细菌广谱高效的抑菌特性，从现有共生细菌资源中筛选对灰葡萄孢具有高拮抗作用的菌株，为葡萄采后灰霉病的抑制提供新的材料和研究方向。[方法] 通过平板对峙培养法和菌丝生长速率法分离筛选拮抗共生细菌，并对优选的高效拮抗共生细菌进行16S rRNA基因序列进化分析，采用扫描电镜观察其对灰葡萄孢菌丝生长的影响，利用损伤接种法对红地球葡萄防治效果进行验证。[结果] 初步分离筛选共获得9株拮抗菌，复筛与复测得到一株抑菌效果显著的共生细菌（命名为ALL），经进化分析其为嗜线虫致病杆菌（Xenorhabdus nematophila），其16S rRNA基因序列的Genbank登录号为MW488402，与菌株Xenorhabdus nematophi la NC116聚于同一分支，相似性达99.79%。扫描电镜观察该菌株导致灰葡萄孢菌丝扭曲变形、表面皱缩、失水塌陷，该菌株发酵（36 h）上清液浓度为1%时对灰葡萄孢菌丝抑制率达44.5%。在葡萄常温防效实验中，与对照组比较，ALL菌株发酵上清液对灰霉菌防治效果较好，3 d后防效为63.50%。[结论] 本研究应用昆虫病原线虫共生细菌生物防治葡萄贮期灰霉病，筛选出一株高效拮抗灰葡萄孢的昆虫病原线虫共生细菌，而且其上清液对灰葡萄孢具有良好的抑制效果，为生物防治贮期葡萄灰霉病提供了新的生物材料和相关研究基础。

Abstract:

[Background] Botrytis cinerea is one of the main pathogenic of grapes during postharvest, which seriously affects the storage period and quality of grapes and brings great losses to its industry. The use of antagonistic microorganisms to inhibit the growth of postharvest pathogens has gradually become an important method for the prevention and control of postharvest gray mold in grapes. [Objective] In order to provide new materials and research direction for the inhibition of Botrytis cinerea postharvest. Using the broad-spectrum and efficient antibacterial characteristics of entomopathogenic nematode symbiotic bacteria and screening high antagonistic effect on Botrytis cinerea from the symbiotic bacteria resources. [Methods] Antagonistic symbiotic bacteria were isolated and screened by plate confrontation culture method and mycelial growth rate method, and antagonistic symbiotic bacteria with highly effective, its 16S rRNA gene phylogeny analysis was carried out. The effect of antagonistic symbiotic bacteria on mycelial growth of Botrytis cinerea was studied by scanning electron microscope, and the control effect on Red Globe grape was verified by damage inoculation method. [Results] Nine strains had antagonistion were isolated from 14 bacteria, that one of symbiotic bacteria (ALL) was obtained through rescreening and retesting with significant antibacterial effect. According to phylogeny analysis, it clustered in the same branch with strain Xenorhabdus nematophila NC116 and 99.79% homology, which was identified to Xenorhabdus nematophila (Genbank accession number:MW488402). The strain caused the Botrytis cinerea hyphae to distort, shrink and collapse by scanning electron microscopy observation. The inhibition rate of the growth of Botrytis cinerea hyphae reached 44.5%, When the supernatant concentration was 1% in medium after fermentation (36 h). Compared with the control group, the fermentation supernatant of strain ALL had a better control effect on Botrytis cinerea, and the control effect was 63.50% after 3 days at room temperature. [Conclusion] In this study, we used entomopathogenic nematode symbiotic bacteria to inhibit Botrytis cinerea for the first time during grapes storage, and one of entomopathogenic nematode symbiotic bacteria was screened with high efficiency against Botrytis cinerea, and its supernatant had good inhibitory effect on Botrytis cinerea. The research provided a new biomaterial and related basis for bio-control of Botrytis cinerea during grapes storage.