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烟草青枯病菌拮抗菌的筛选、鉴定及生防特性研究
引用本文:夏 艳,徐 茜,董 瑜,林 勇,孔凡玉,张成省,王 静,宋毓峰.烟草青枯病菌拮抗菌的筛选、鉴定及生防特性研究[J].中国生态农业学报,2014,22(2):201-207.
作者姓名:夏 艳  徐 茜  董 瑜  林 勇  孔凡玉  张成省  王 静  宋毓峰
作者单位:1.烟草行业烟草病虫害监测与综合治理重点实验室 中国农业科学院烟草研究所 青岛 266101 3.中国农业科学院研究生院 北京 100081;福建省烟草公司南平市分公司 南平 353000;1.烟草行业烟草病虫害监测与综合治理重点实验室 中国农业科学院烟草研究所 青岛 266101 3.中国农业科学院研究生院 北京 100081;福建省烟草公司南平市分公司 南平 353000;烟草行业烟草病虫害监测与综合治理重点实验室 中国农业科学院烟草研究所 青岛 266101;烟草行业烟草病虫害监测与综合治理重点实验室 中国农业科学院烟草研究所 青岛 266101;烟草行业烟草病虫害监测与综合治理重点实验室 中国农业科学院烟草研究所 青岛 266101;1.烟草行业烟草病虫害监测与综合治理重点实验室 中国农业科学院烟草研究所 青岛 266101 3.中国农业科学院研究生院 北京 100081
基金项目:国家自然科学基金项目(31000878)和福建省南平市烟草公司科技项目(201203)资助
摘    要:烟草青枯病危害严重,以拮抗菌进行防病的生物防治手段成为研究热点。从不同烟田分离纯化出238株细菌菌株,首先经牙签接种初筛,选取对青枯病菌抑制效果较好的菌株制备其抑菌物质的粗提物,以牛津杯法复筛,最终获得3株对烟草青枯病菌有明显抑制作用的拮抗细菌。全细胞脂肪酸、16S rDNA及gyrB基因测序等分析结果表明,菌株H19、Y6为解淀粉芽孢杆菌(Bacillus amyloliquefaciens),菌株H34为甲基营养型芽孢杆菌(B.methylotrophicus)。3株拮抗菌经CAS检测平板法和Salkowski比色法,发现均具有产铁载体和吲哚-3-乙酸(IAA)的能力,以菌株H19能力最强。温室促生试验结果表明,3株拮抗菌能显著促进烟草株高、鲜重及干重等指标,与对照相比,平均增长率分别达到70%~115%、40%~49%和32%~42%。温室控病试验结果表明,菌株H19、H34和Y6明显降低烟草青枯病的发病率,防效达76.57%、60.98%和69.83%,稍逊于农用链霉素处理的78.66%。

关 键 词:烟草青枯病  拮抗细菌  铁载体  植物生长素  促生作用  防治效果
收稿时间:2013/10/23 0:00:00
修稿时间:2013/12/26 0:00:00

Screening, identification and characterization of antagonistic bacteria against Ralstonia solanacearum
XIA Yan,XU Qian,DONG Yu,LIN Yong,KONG Fanyu,ZHANG Chengsheng,WANG Jing and SONG Yufeng.Screening, identification and characterization of antagonistic bacteria against Ralstonia solanacearum[J].Chinese Journal of Eco-Agriculture,2014,22(2):201-207.
Authors:XIA Yan  XU Qian  DONG Yu  LIN Yong  KONG Fanyu  ZHANG Chengsheng  WANG Jing and SONG Yufeng
Institution:1. Tobacco Integrated Pest Management of China Tobacco; Tobacco Research Institute of Chinese Academy of Agricultural Sciences, Qingdao 266101, China;3. Graduate School of Chinese Academy of Agricultural Sciences, Beijing 100081, China;Tobacco Research Institute of Chinese Academy of Agricultural Sciences,Qingdao 266101, China;1. Tobacco Integrated Pest Management of China Tobacco; Tobacco Research Institute of Chinese Academy of Agricultural Sciences, Qingdao 266101, China;3. Graduate School of Chinese Academy of Agricultural Sciences, Beijing 100081, China;Tobacco Research Institute of Chinese Academy of Agricultural Sciences,Qingdao 266101, China;Tobacco Integrated Pest Management of China Tobacco; Tobacco Research Institute of Chinese Academy of Agricultural Sciences, Qingdao 266101, China;;Tobacco Integrated Pest Management of China Tobacco; Tobacco Research Institute of Chinese Academy of Agricultural Sciences, Qingdao 266101, China;;Tobacco Integrated Pest Management of China Tobacco; Tobacco Research Institute of Chinese Academy of Agricultural Sciences, Qingdao 266101, China;;1. Tobacco Integrated Pest Management of China Tobacco; Tobacco Research Institute of Chinese Academy of Agricultural Sciences, Qingdao 266101, China;3. Graduate School of Chinese Academy of Agricultural Sciences, Beijing 100081, China
Abstract:Tobacco bacterial wilt caused by Ralstonia solanacearum is one of the most destructive bacterial diseases of tobacco. As a recurring soil-borne disease, it is controlled mainly by chemical methods that are not only costly and less efficient but also have food safety and environment safety problems. Consequently, biological control methods that use antagonistic strains have gained strength in tobacco bacterial wilt disease research. To find new antagonist bacteria against R. solanacearum, 238 strains were identified and screened for in vitro antibiosis. By placing each strain on cultured pathogen plates with toothpicks 24 strains showed inhibition zones. Among the 24 strains, the width of the inhibition zones of 7 antagonist strains exceeded 10 mm. Then pathogen inhibitive ingredients of the 7 strains were extracted to determine bacteriostatic efficacy. Finally, 3 promising strains (H19, Y6, H34) were obtained. H19 and Y6 were identified as Bacillus amyloliquefaciens and the other strain (coded H34) identified as B. methylotrophicus. The strain classification was based on phylogenetic analysis of 16S rDNA sequence, gyrB sequence and fatty acid composition. Siderophores secreted by microorganisms to take up iron from the environment had an action mode of disease suppression that was solely based on competition with the pathogens for iron. Besides, indole-3-acetic acid (IAA) is a phytohormone known to be involved in root initiation, cell division and cell enlargement. In follow-up experiments, all the three strains showed the ability to produce siderophores and plant growth promotor IAA. However, productions of siderophores and IAA were different, with H19 and Y6 as the highest and lowest producers under improved CAS and Salkowski colorimetric methods. The three strains were further characterized for plant growth promoting traits and disease control effects. Compared with control treatment, the strains significantly improved the growth of tobacco with increases of 19% 24% in the maximum leaf length, 7% 12% in the maximum leaf width, 70% 115% in plant height, 2% 14% in stem diameter, 40% 49% in fresh weight and 32% 42% in dry weight. H19, H34 and Y6 reduced bacterial wilt incidence by 76.57%, 60.98% and 69.83%, respectively in greenhouse experiments. This was slightly lower than using 40% agricultural streptomycin which had disease incidence control efficacy of 78.66%. In conclusion, the research isolated new efficient antagonistic strains against tobacco bacterial wilt. It also proved the mechanism of disease control and plant growth promotion by H19, H34 and Y6. The study further showed that the three strains had great potential for plant growth promotion and biological con-trol under greenhouse conditions, which developed the needed reference for follow-up studies.
Keywords:Tobacco bacterial wilt  Antagonistic bacteria  Siderophore  Phytohormone  Growth promotion  Control effect
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