防治黄瓜霜霉病的细菌制剂与叶围本土微生物的互作研究

Downy mildew caused by the Pseudoperonospora cubensis is an important disease of cucumber. Our group selected 3 bacterial strains of different genera from more than 2,100 isolates. These 3 kinds of bacterial preparation can be used to efficiently control cucumber downy mildew. According to DGGE analysis, biocontrol treatment increased microbial diversity in cucumber leaf surface, suggesting that the biocontrol bacteria change the local microbial community which plays a key role in the process of biocontrol of downy mildew. In this project, a greenhouse experiment will be carried out before the field trials in which qualitatively and quantitatively colonization pattern of the 3 biocontrol bacterial strains on cucumber foliage and their impacts on phyllospere microecology will be detected in different climates, cultivars, and growth stages in 3 continues years by using PCR-DGGE, the traditional culture and isolation, Q-PCR, and confocal laser microscopy. The different DNA stripes from DGGE gel pattern between treatment and control groups will be sequenced and the microbes will be identified, and then verified by traditional culture and isolation method, thus the microbial interactions between biocontrol agents with cucumber phyllosphere microflora can be analyzed respectively, and their influence on cucumber phyllospere microflora will be found. Meanwhile, we will also test the impacts of biocontrol agents on the plant heights, stem diameters, leaves numbers, amount of chlorophyll, fruits production and quality, and the natural conservation time, to evaluate the efficacy of plant growth promotion, disease control, and economic increase by our biocontrol agents; establish the interaction pattern of biocontrol agents and cucumber phyllospere microflora, and lay the theoretical foundation for the biocontrol of plant leaf diseases.

由古巴假霜霉菌引起的霜霉病是黄瓜重要病害。本课题组前期从2100个细菌菌株中筛选到3株能够高效防治黄瓜霜霉病的不同属细菌，并发现生防菌处理增加了黄瓜叶围微生物多样性，推测生防菌与本土微生物的互作在霜霉病生防过程中发挥重要作用。本项目拟在完成温室限菌条件实验后，利用DGGE、传统分离培养、Q-PCR与激光共聚焦检测相互验证的方法，从不同气候条件、不同适宜品种、不同生育期黄瓜上连续3年定性定量检测3株生防菌在黄瓜叶面的定殖规律及其对黄瓜叶表微生态的影响。将DGGE检测出的与对照组相比差异性条带回收测序，定性检测3株生防菌分别与黄瓜叶围本土微生物的互作结果；寻找其对黄瓜叶围微生态的影响规律。同时检测生防菌与叶围微生物互作对植物株高、茎粗、叶片数、叶绿素含量、黄瓜产量及品质、自然保鲜期的影响，评估其防病促生增值效果，建立生防菌与黄瓜叶表本土微生物互作模式，为植物叶部病害生防奠定理论基础。

由古巴假霜霉菌引起的霜霉病是黄瓜重要病害。本项目选择黄瓜生境的22个菌株和小麦、生姜、香蕉生境中的27个菌株总计49株颉颃细菌进行温室试验，接种病原菌15 d后，颉颃细菌对黄瓜霜霉病的防效为6.64–83.21%。选择防效在75%以上的3个颉颃细菌开展其田间设施大棚的小区试验，颉颃细菌处理组在其初次喷雾后第17、27、37 d的病害严重度均显著低于对照组，第一轮田间防效在2.64-73.44%，第二轮田间防效在48.34-75.68%。分别在温室和田间条件下，将6个颉颃细菌菌液（108 CFU/ml）喷雾于黄瓜叶部，分别统计其第0、3、7、15、21、30d在叶围的定殖情况，结果显示颉颃菌株在第30 d 时，大部分颉颃细菌在黄瓜叶片上定殖量保持在103 CFU/g。对黄瓜霜霉病防效较好的菌株进行叶围和根围DGGE微生态分析，DGGE电泳结果表明生防菌处理能显著增加了叶围和根围微生物的多样性。