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烟草棒孢霉叶斑病生防菌GK-6的抗病机制

引用本文：于心如,李琳新,吴惠平,张婷婷,谈发洋,夏春成,黄纯杨,丁婷.烟草棒孢霉叶斑病生防菌GK-6的抗病机制.植物保护学报,2025,52(6):1404-1415

DOI：10.13802/j.cnki.zwbhxb.2025.2025076

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作者	单位	E-mail
于心如	安徽农业大学植物保护学院, 植物病虫害生物学与绿色防控安徽普通高校重点实验室, 作物有害生物综合治理安徽省重点实验室, 合肥 230036
李琳新	安徽农业大学植物保护学院, 植物病虫害生物学与绿色防控安徽普通高校重点实验室, 作物有害生物综合治理安徽省重点实验室, 合肥 230036
吴惠平	安徽农业大学植物保护学院, 植物病虫害生物学与绿色防控安徽普通高校重点实验室, 作物有害生物综合治理安徽省重点实验室, 合肥 230036
张婷婷	安徽农业大学植物保护学院, 植物病虫害生物学与绿色防控安徽普通高校重点实验室, 作物有害生物综合治理安徽省重点实验室, 合肥 230036
谈发洋	安徽农业大学植物保护学院, 植物病虫害生物学与绿色防控安徽普通高校重点实验室, 作物有害生物综合治理安徽省重点实验室, 合肥 230036
夏春成	安徽农业大学植物保护学院, 植物病虫害生物学与绿色防控安徽普通高校重点实验室, 作物有害生物综合治理安徽省重点实验室, 合肥 230036
黄纯杨	贵州省烟草公司遵义市公司, 遵义 563000	Hcy_0701@126.com
丁婷	安徽农业大学植物保护学院, 植物病虫害生物学与绿色防控安徽普通高校重点实验室, 作物有害生物综合治理安徽省重点实验室, 合肥 230036	18355433@qq.com

中文摘要:为筛选烟草棒孢霉叶斑病的生防菌源，从贵州省遵义市发病烟区的健康烟株中分离内生细菌，以多主棒孢霉Corynespora cassiicola为指示菌进行拮抗菌株的筛选，通过形态特征与多基因序列串联分析对拮抗菌株进行鉴定，利用结晶紫染色法和平板运动性试验解析其生物膜形成能力及群集运动特性，采用盆栽试验评价其对烟草棒孢霉叶斑病的防效，通过分生孢子萌发抑制试验、菌丝生长速率法和基质辅助激光解析电离-飞行时间质谱技术探究其抑菌机制，并采用绿色荧光蛋白标记法测定其在烟株中的定殖动态。结果表明：筛选获得1株对多主棒孢霉有较高抑制作用的菌株，抑制率达63.75%，命名为GK-6，并被鉴定为贝莱斯芽胞杆菌Bacillus velezensis。该菌株表现出较强的生物膜形成能力及群集运动特性；其对烟草棒孢霉叶斑病的防效为41.67%；经菌株GK-6发酵上清液处理96 h的多主棒孢霉分生孢子萌发率仅为64.67%，显著低于阴性对照组（100.00%）；其粗提物对多主棒孢霉的EC₅₀为0.032 mg/mL，且含有丰原素、表面活性素、杆菌霉素等抗菌物质。该菌株可在烟株叶部稳定定殖，接种后35 d仍可检测到2.87×10³ CFU/g的定殖量。表明贝莱斯芽胞杆菌GK-6具有开发为生物农药的潜力。

中文关键词:烟草棒孢霉叶斑病多主棒孢霉贝莱斯芽胞杆菌脂肽分生孢子萌发

The mechanism of resistance in biocontrol bacterial strain GK-6 against tobacco Corynespora leaf spot

Author Name	Affiliation	E-mail
Yu Xinru	Anhui Province Key Laboratory of Integrated Pest Management on Crops, Key Laboratory of Biology and Sustainable Management of Plant Diseases and Pests of Anhui Higher Education Institutes, School of Plant Protection, Anhui Agricultural University, Hefei 230036, Anhui Province, China
Li Linxin	Anhui Province Key Laboratory of Integrated Pest Management on Crops, Key Laboratory of Biology and Sustainable Management of Plant Diseases and Pests of Anhui Higher Education Institutes, School of Plant Protection, Anhui Agricultural University, Hefei 230036, Anhui Province, China
Wu Huiping	Anhui Province Key Laboratory of Integrated Pest Management on Crops, Key Laboratory of Biology and Sustainable Management of Plant Diseases and Pests of Anhui Higher Education Institutes, School of Plant Protection, Anhui Agricultural University, Hefei 230036, Anhui Province, China
Zhang Tingting	Anhui Province Key Laboratory of Integrated Pest Management on Crops, Key Laboratory of Biology and Sustainable Management of Plant Diseases and Pests of Anhui Higher Education Institutes, School of Plant Protection, Anhui Agricultural University, Hefei 230036, Anhui Province, China
Tan Fayang	Anhui Province Key Laboratory of Integrated Pest Management on Crops, Key Laboratory of Biology and Sustainable Management of Plant Diseases and Pests of Anhui Higher Education Institutes, School of Plant Protection, Anhui Agricultural University, Hefei 230036, Anhui Province, China
Xia Chuncheng	Anhui Province Key Laboratory of Integrated Pest Management on Crops, Key Laboratory of Biology and Sustainable Management of Plant Diseases and Pests of Anhui Higher Education Institutes, School of Plant Protection, Anhui Agricultural University, Hefei 230036, Anhui Province, China
Huang Chunyang	Zunyi Company of Guizhou Tobacco Company, Zunyi 563000, Guizhou Province, China	Hcy_0701@126.com
Ding Ting	Anhui Province Key Laboratory of Integrated Pest Management on Crops, Key Laboratory of Biology and Sustainable Management of Plant Diseases and Pests of Anhui Higher Education Institutes, School of Plant Protection, Anhui Agricultural University, Hefei 230036, Anhui Province, China	18355433@qq.com

Abstract:In order to screen for biocontrol bacterial sources against tobacco Corynespora leaf spot, endophytic bacteria have been isolated from healthy tobacco in the disease-affected tobacco-growing areas in Zunyi City, Guizhou Province. Then the antagonistic strain was screened using the fungal pathogen Corynespora cassiicola, and identified by combining colony morphology characteristics with multilocus sequence analysis. Its biofilm-forming ability and swarming motility were analyzed using crystal violet staining and plate motility assays. Subsequently, the pot experiment was conducted to evaluate the control efficacy of the antagonistic strain against the tobacco Corynespora leaf spot. Furthermore, conidial germination inhibition assays, mycelial growth rate determination, and Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry analysis were performed to elucidate the antifungal mechanisms of this antagonistic strain. Subsequently green fluorescent protein labeling was employed to investigate the colonization dynamics of this antagonistic strain in tobacco plants. The results demonstrated that a bacterial strain, which exhibited an inhibition rate of 63.75% to Corynespora cassiicola, has been obtained and named GK-6. The strain GK-6 was identified as Bacillus velezensis, exhibiting strong biofilm-forming capacity and swarming motility. Its control efficacy on Corynespora leaf spot in tobacco reached 41.67%. The conidial germination of C. cassiicola treated with the fermentation supernatant of the strain GK-6 for 96 h was only 64.67%, which was lower than that of the control group（100.00%）. The crude extract of the strain GK-6 exhibited potent antifungal activity against C. cassiicola with an EC₅₀ value of 0.032 mg/mL, and the crude extract contained antimicrobial compounds such as fengycin, surfactin, and bacillomycin. Additionally, the strain GK-6 could stably colonize on the leaves of tobacco plants, on the 35 th day after inoculation, its population was 2.87×10³ CFU/g. The results of this study provided a theoretical basis for the development of B. velezensis GK-6 as a biocontrol agent.

keywords:tobacco Corynespora leaf spot Corynespora cassiicola Bacillus velezensis lipopeptide conidial germination

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