AM真菌对甘薯连作障碍的生防作用及其机制研究

Arbuscular mycorrhiza (AM), a ubiquitous symbiotic association established between AM fungi and roots of higher plant in most terrestrial ecosystems, is essentially important for plant adaptation to various environmental stresses, such as nutrient deficiency, environmental pollution and soil-borne disease, etc. Many studies have proved the positive influence of AM on resistance of soil-borne disease and made efforts to uncover the underlying mechanisms. However, experimental evidences are still lacking to support the ecological significance of AM associations in natural ecosystems suffering continuous cropping obstacle, and available information is so far very limited as for the molecular mechanisms underlying the enhanced plant disease tolerance by AM symbiosis. In the proposed project, the planting base of sweet potato in Qingdao that covers different continuous cropping years is selected as target research area to reveal the variations of microbial biomass and community along the increase of continuous cropping years, by using phospholipid fatty acid technology (PLFA) and MiSeq Illumina sequencing technologies (Illumina Miseq). Meanwhile, the dominant AM fungi and pathogens populations will also be revealved by these methods. Furthermore, the dominant AM fungi and pathogens obtained by the above field investigation is chose as the model microorganisms to the biocontrol mechanism of AM fungi on sweet potato through pot experiment, which is characterized by plant physiological function of the AM symbiosis. Finally, the biocontrol effect and mechanism of AM fungi on sweet potato cropping obstacles are proved by the field cultivation of sweet potato with AM fungal inoculation. The study is expected to demonstrate the ecological significance of AM symbiosis in plant disease tolerance, and more importantly to reveal the underlying molecular mechanisms. It will also lay foundations for further research into the synergetic stress resistance of AM symbiosis.

针对甘薯栽培中克服连作障碍的实际生产需求，立足于土壤微生物生态学研究的前沿，本项目拟采用野外分子生态调查、室内盆栽机制研究及大田效应验证的三位一体的研究体系，系统地揭示AM真菌对甘薯连作障碍的生防作用及机制。以山东省青岛市胶州甘薯研究体系的种植基地为平台，采用磷脂脂肪酸技术和高通量测序技术相结合的方法，定量和定性比较甘薯不同连作年限的土壤微生物（AM真菌、细菌、真菌）生物量及群落组成的差异，揭示AM真菌抑制甘薯连作障碍的微生态学机制，明确甘薯的优势AM真菌及病原菌的种群信息；选用上述研究获得的主导病原菌与优势AM真菌为模式微生物，通过室内盆栽模拟实验，探讨AM真菌对甘薯连作障碍的植物生防机制；最后采用纸袋法田间接种优势AM真菌种群，在田间条件下验证AM真菌对甘薯连作障碍的生防效应及其机制。通过上述研究系统地揭示AM真菌在防御甘薯连作障碍中的生理生态机制，为防治甘薯连作障提供技术和理论支撑。

甘薯连作障碍成为制约其植株生长及产量形成的关键因素，然而其连作障碍的土壤微生态机制尚未见报道，对其连作病害的生物防治措施的研究也尚显薄弱。为了揭示甘薯连作障碍的土壤微生态机制，以及为甘薯连作病害的生物防治提供理论及技术支持，本研究主要从三个方面展开。.（1）首先采用现代分子生态学方法对甘薯连作的微生态机制进行了系统的研究。结果表明，连作后土壤酸化、有机质减少、土壤主要酶活性降低、微生物区系改变、有害病菌数量增加及有益菌数量减少，是导致甘薯连作病害发生的主要原因。该研究揭示了与连作相关的细菌、真菌、AM真菌的指示物种，为甘薯连作病害的防治及生防菌的筛选奠定了理论基础。.（2）通过第一部分的研究发现，除了AM真菌，细菌和真菌这两类微生物也在“甘薯连作土壤的自然微生态平衡机制”中起到了非常关键的作用。本研究以我国及研究区域甘薯主要病害“黑斑病”为主要目标，从甘薯典型发病区的“抑病土”土壤中，分离筛选出12株对甘薯黑斑病有拮抗效果的优良生防细菌及真菌菌株，并对其进行形态特征、生长特性研究和16S、18S rRNA 基因序列鉴定，为甘薯连作病害的防治提供了高效生防菌株资源及理论基础。.（3）以优势AM真菌，及筛选出来的优势拮抗细菌（荧光假单胞菌）为材料，通过室内盆载模拟实验，研究这两种生防菌对于甘薯黑斑病的的生防效果及其机制。结果表明AM真菌和荧光假单胞菌均能通过促进甘薯生长，增加其地上部和根系的生物量，提高甘薯地上部和根系的CAT、PPO、POD、降低抗病相关的防御性酶活性，以及增加土壤中微生物的数量，来增强甘薯对于黑斑病的抗性，有效减轻病害的发生。.综上，本研究系统的揭示了甘薯连作障碍的土壤微生态机制，获得了12株高效生防菌菌株资源，也初步探讨了AM真菌和生防细菌对甘薯连作病害的防治效应及其机制，为甘薯连作病害的生物防治奠定了基础。