枯草芽孢杆菌BS193对辣椒疫病的生防作用及其抗菌机制初探

发布时间：2018-04-30 08:43

本文选题：辣椒疫病 + 枯草芽孢杆菌　；参考：《福建农林大学》2017年硕士论文

【摘要】：由辣椒疫霉菌(Phytophthora capsici)侵染引起的辣椒疫病,是辣椒生产上的重要病害之一,广泛分布于世界各辣椒主产区。目前,在生产上控制辣椒疫病最易行且见效快的方法是施用化学药剂,但长期施用药剂使病原菌易产生一定抗药性,且农药残留问题突出,故寻找高效且不污染环境的方法对防治辣椒疫病具有重要意义,其中拥有广泛前景的方法是利用生防菌进行生物防治。本研究利用实验室早期分离获得一株具有生防作用的枯草芽孢杆菌BS193为材料,以辣椒疫病为对象,研究该菌株抗菌谱、生防作用及其抗菌机制,为辣椒疫病的生物防治和该生防菌的深入开发应用提供实践依据。得出如下结论:1、枯草芽孢杆菌BS193的抗菌作用具有广谱性。以平板对峙法对枯草芽孢杆菌BS193的抗菌谱进行测定,结果表明,枯草芽孢杆菌BS193对9个属10种常见的植物病原菌均有抑制作用。2、枯草芽孢杆菌BS193对辣椒疫病具有良好的生防效果。在平板对峙试验中,结果显示出菌株BS193对4种辣椒疫霉菌小种抑制作用稳定;在离体抑菌实验中,表明经BS193发酵液处理后抑制辣椒疫霉菌侵染叶片,其防效达到63.50%;在室内盆栽防病试验中,结果表明BS193菌体、无菌滤液、发酵液原液处理后24 h接种病原菌,9 d后各处理的防治效果分别为:30.75%、29.76%、52.56%;BS193发酵液、100倍稀释液、1000倍稀释液处理辣椒苗,接种病原菌9 d后的防效为:52.56%、34.56%、23.53%;BS193发酵液灌根后间隔0、24、48、72 h后接种病原菌,9 d后各处理的防治效果分别为:23.43%、41.25%、49.41%、51.44%。由此可知,枯草芽孢杆菌BS193对辣椒疫病的防病效果良好。同时,试验中发现BS193发酵液对辣椒幼苗还具有很好的促生作用。用BS193发酵液灌根30 d后测量发现,与对照组相比,辣椒苗株高、根长、鲜重和干重分别增加:50.04%、48.16%、73.27%和 48.83%。3、枯草芽孢杆菌BS193无菌滤液对辣椒疫霉菌菌丝生长、孢子囊形成、游动孢子萌发等具有显著的抑制活性。试验结果表明经菌株BS193无菌滤液处理后菌丝生长速率显著下降,在光学和电子显微镜下观察到菌丝分枝增多、畸形膨大、扭曲变形、皱缩等;辣椒疫霉孢子囊和游动孢子萌发的数量均明显减少,其抑制率都显著上升。4、初步鉴定BS193所合成的抗菌物质:采用浓盐酸沉淀法粗提抗菌物质,检测抑菌活性并进行ULPC-QTOF/MS分析。试验结果显示抗菌粗提物对辣椒疫霉菌菌丝生长抑制效果显著;经ULPC-QTOF/MS分析出现多处明显的质谱响应峰,核质比m/z值比较,初步鉴定抗菌粗提物中含有Iturin(伊枯草菌素)、Feycin(丰原菌素)、Surfactin(表面活性素)这3类脂肽类抗菌物质。
[Abstract]:Pepper blight caused by Phytophthora capsicii is one of the most important diseases in pepper production. At present, the most convenient and effective way to control pepper blight in production is to use chemical chemicals, but long-term application of insecticides makes pathogens easily develop resistance to certain drugs, and the pesticide residue problem is prominent. Therefore, it is of great significance to find efficient and non-polluting methods for the prevention and control of pepper blight, among which the biological control with biocontrol bacteria is one of the most promising methods. In this study, a biocontrol strain of Bacillus subtilis (BS193) was isolated in the early stage of laboratory. The antimicrobial spectrum, biocontrol effect and antibacterial mechanism of this strain were studied. It provides a practical basis for the biological control of pepper blight and the further development and application of this biocontrol bacterium. The results showed that the antimicrobial activity of Bacillus subtilis BS193 was broad spectrum. The antimicrobial spectrum of Bacillus subtilis BS193 was determined by plate confrontation method. Bacillus subtilis BS193 can inhibit 9 genera and 10 common plant pathogens. Bacillus subtilis BS193 has a good biocontrol effect on pepper blight. In the plate confrontation test, the results showed that the inhibition effect of the strain BS193 on the four strains of Phytophthora capsici was stable, and in vitro, the inhibition of Phytophthora capsici infection on leaves was shown by the treatment of BS193 fermentation broth. The control effect reached 63.50. The results showed that BS193 bacteria, aseptic filtrate, and sterility filtrate were used in pot experiment. The control effects of the fermentation broth after 24 h inoculation with pathogenic bacteria for 9 d were as follows: 1: 30.75 and 29.76% and 52.56% BS193 100 times diluent and 1000 times diluent to treat capsicum seedlings, respectively. 9 days after inoculation with pathogenic bacteria, the control effect was:% 52.56 and 34.560.The control effect of each treatment after inoculating the pathogen 9 days after inoculation with BS193 fermentation broth was 0.24 and 4872 hours after inoculation, and the control effects were respectively 51.441.49.41and 51.441.41.41.41.41.41.41.41.41.41.41.41.41.The control effect of each treatment was as follows: 59.41.41.41.44 after inoculating the pathogen for 9 days. It can be seen that Bacillus subtilis BS193 has a good effect on preventing pepper blight. At the same time, it was found that BS193 broth had a good effect on the growth of pepper seedlings. Compared with the control group, the plant height, root length, fresh weight and dry weight of capsicum seedlings were increased by 73.27% and 48.833.27% and 48.833.3.3respectively, respectively. The growth of Phytophthora capsici hyphae and the formation of sporangia were observed by BS193 aseptic filtrate of Bacillus subtilis. The germination of zoospore showed significant inhibitory activity. The results showed that the growth rate of mycelium decreased significantly after treatment with BS193 aseptic filtrate. The mycelium branching, deformity, distortion and shrinkage were observed under optical and electron microscopy. The number of sporangia and zoospore germination of Phytophthora capsici was significantly reduced, and the inhibition rate was increased significantly. The antimicrobial substances synthesized by BS193 were preliminarily identified. The antimicrobial substances were extracted by concentrated hydrochloric acid precipitation method, the antibacterial activity was detected and ULPC-QTOF/MS analysis was carried out. The results showed that the antimicrobial crude extract had significant inhibitory effect on the growth of Phytophthora capsici hyphae, and there were many obvious mass spectrum response peaks by ULPC-QTOF/MS analysis, and the ratio of nucleus to cytoplasm m / z was compared. Three kinds of lipopeptide antimicrobial substances Iturin (Iturin) Feycinin (surfacin) were identified in the crude antimicrobial extracts.
【学位授予单位】：福建农林大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S436.418

【参考文献】