番茄灰霉病生防链霉菌筛选及鉴定

Screening and identification of the biocontrol Streptomyces against tomato Botrytis cinerea

【背景】由灰葡萄孢侵染所致的番茄灰霉病是一类重要的真菌病害,生物防治具有环境友好、病原菌不易产生抗药性等特点,是果蔬灰霉病绿色防控的有效措施。【目的】筛选对番茄灰霉病具有防病作用且能促进番茄种子发芽的广谱拮抗性链霉菌,并明确该菌株种级分类地位。【方法】采用琼脂块法筛选拮抗番茄灰霉病菌的链霉菌菌株,采用对峙培养法和生长速率法检测菌株T22抑菌谱,通过产胞外酶活性检测、离体叶片防效和种子发芽试验明确该菌株的防病促生相关特性,根据形态学特征、生理生化特性和分子生物学方法对该菌株进行种类鉴定。【结果】从分离的56株放线菌中筛选到14株对番茄灰霉病菌具有拮抗效果的放线菌菌株,其中链霉菌T22对番茄灰霉病菌抑制作用最强,且具有较广抑菌谱,同时菌株T22具有产生纤维素酶和几丁质酶的能力。菌株T22无菌发酵滤液对番茄灰霉病菌、桃褐腐病菌、黄瓜枯萎病菌抑菌率分别为84.6%、81.5%和79.1%;其无菌发酵滤液原液对番茄灰霉病离体防效为55.1%;100倍稀释液处理番茄种子,胚轴、胚根和种子活力指数分别增加15.1%、29.7%和43.9%。根据形态学特征、生理生化特性和多基因聚类分析将链霉菌T22鉴定为白黑链霉菌(Streptomycesalboniger)。【结论】白黑链霉菌T22具有较强的抗真菌、产胞外酶、防病和促生活性,在番茄灰霉病生物防治中具有较好的开发应用潜力。

[Background]Tomato grey mould caused by Botrytis cinerea is an important fungal disease.Biological control can be an effective measure in prevention and control of this disease,due to its environmentally-friendly and not easy to develop pathogen resistance characters.[Objective]To screen antagonistic Streptomyce sp.with broad-spectrum antifungal activity,confirm the biocontrol effects and define the classification status of the antagonistic strain.[Methods]Antagonistic actinomycetes were obtained by agar block method,the inhibition spectrum of strain T22 was detected using confrontation culture and mycelia growth assays.Biocontrol effects of strain T22 was detected using enzymes production,detached leaves and seed germination assays.Strain T22 was identified to species level based on morphological and cultural features,physiological-biochemical characters,as well as molecular methods.[Results]A total of 14 antagonistic isolates were screened from 56 soil actinomycetes,among which strain T22 showed obvious inhibition effect on Botrytis cinerea and other fungi.The inhibition rate of strain T22 on Botrytis cinerea,Monilinia fructicola and Fusarium oxysporum f.sp.cucumerinum were 84.6%,81.5%and 79.1%,respectively.In addition,the culture filtrate of strain T22 displayed obvious disease reduction effect(55.1%)on detached leaves of tomato plants.The hypocotyl length,radical length and seed vigor index were increased by 15.1%,29.7%and 43.9%,respectively,with 100-fold dilution treatment.Strain T22 was identified as Streptomyces alboniger based on morphological and cultural features,physiological-biochemical characters,as well as multiple sequences analyses.[Conclusion]Streptomyces alboniger T22 had obvious antifungal activity,extracellular hydrolase production ability,disease reduction and seed germination promotion effects on tomato,with application potential as biocontrol agent against tomato grey mould.