宁夏荒漠植物柠条内生细菌多样性及其生物学功能研究

Endophytic bacteria can be defined as those bacteria that can colonize the internal tissue of the plant without showing negative signs on their host. Many endophytes have potential uses as agents for stimulating plant growth and make bioactive natural compounds that are a potential source of novel products for use in medicine, agriculture, and industry. So the study of plant bacterial endophytes is important for understanding ecological interactions and to develop biotechnological applications. Howeve，very little is known that the diversity , structure composition and functions of endophytes within sand binding plants that grown in extreme environment. Caragana microphylia，is a perennial leguminous shrub that is highly tolerant to drought, salt and extreme cold environment in relatively poor or sandy, well-drained soil. It is often used as windbreaks to protect soils from desertification in the northern regions of China. Considering the ecological importance of the C. microphylia and the lack of information about its endophytes, it is essential to systematically study the relationship of endophytic bacteria and host plants. To this end, the genetic diversity and community composition diversity of endophytic bacteria associated with plant Caragana microphylia , grown in desertification grassland of Ningxia Baijitan and Shapotou National Nature Reserve, will be analyzed using methods of PCR-RFLP , sequencing , library analyses and DGGE based on 16S rRNA gene in this study. In order to screening the stress resistance and high efficient strains, the biological functions such as phosphorus dissolving , nitrogen fixation, secretion of growth hormone , anti-plant pathogenic and stress toleranc of endophytes will be detected. This project will not only can rich the microorganism resources of China, but also can promote the protection, development and utilization of microorganism resources in arid area, as well as has an important instructive significance to the ecological interactions and coevolution between the endophytic bacteria and host plants.

近几年来，植物内生细菌因具有多方面的生物学和生态学功能，成为潜力巨大、尚待开发的微生物新资源而受到广泛重视。但对极端环境中分布的具有重要生态意义的植物内生细菌的多样性及其作用的研究却很少涉及。本项目拟以鄂尔多斯荒漠区为背景，以宁夏白芨滩、沙坡头国家自然保护区为基地，对宁夏中部干旱风沙区内主要的荒漠固沙植物柠条不同组织中内生细菌的遗传多样性、群落组成多样性、抗逆性能及溶磷、固氮、分泌生长素、抗植物病原菌等生物学功能进行研究，对内生细菌增强宿主植物抗逆性的作用途径进行分析，筛选抗逆性强的高效促生菌株并进行功能验证。该项目的完成将为丰富我国的微生物资源、推动干旱地区微生物资源的保护、开发和利用提供依据，同时为探索极端环境中宿主植物- - 内生细菌抗逆性能的相互作用机制及协同进化研究有着重要的指导意义。

植物内生细菌因具有多方面的生物学和生态学功能，成为潜力巨大、尚待开发的微生物新资源而受到广泛重视。但对极端环境中分布的具有重要生态意义的植物内生细菌的多样性及其作用的研究却很少涉及。宁夏位于我国西北的东部，是典型的荒漠化草原地带，是我国西部土地沙漠化最严重的地区之一，自然生态恶化。柠条作为宁夏荒漠区长期发展的重要植物资源，在维护干旱荒漠区的生态平衡，改善沙区环境，发展畜牧业生产，提高沙区的经济效能中发挥着十分重要的作用。对干旱荒漠区的柠条内生细菌多样性和生物学功能的研究将有助于了解极端环境中内生细菌的生物学特性及未知的生态学作用、发掘新的微生物种质资源。本项目主要研究内容和结果：（1）运用可培养和免培养方法，对柠条各部分组织中内生细菌的遗传多样性、群落结构和分布特征进行了分析，建立和优化了柠条内生细菌的分离培养体系。基于16S rDNA的 RFLP 和序列分析表明柠条内生细菌的数量和群落组成存在明显的组织特异性，根部菌群密度最大，茎部和种子最少。芽孢杆菌属是优势内生菌，分布于所有组织中。确定了柠条内生细菌的多样性组成信息及优势菌群的系统发育地位，通过向Genbank提交序列，丰富了基因数据库中相关信息，实现了资源共享。（2）测定了78株分离菌株的耐盐性、耐酸碱性和生长温度范围。结果表明柠条内生细菌的抗逆能力较强，多数菌株可耐受6%的NaCl，能在pH值 5.0—10.0及10-45 ℃的条件下生长。根内细菌的耐盐性普遍较强，叶部菌株的耐盐性差异最大，茎部的耐盐性较弱。筛选获得4株能耐受10% NaCl和55 ℃高温的抗逆菌株。（3）对菌株的解磷、分泌生长素和抑菌活性等生物学功能进行了检测。部分菌株具备溶解有机磷或无机磷的能力，7株菌同时具备溶解有机磷和无机磷的能力，但解磷能力较弱。6个具有强的产IAA能力的菌株隶属于根瘤菌，多数芽孢杆菌无产IAA活性。多数菌株具有抑制植物病原菌生长的能力，筛选获得3株具有广谱抗菌活性的抗逆菌株。（4）初步分析了内生细菌对宿主植物生存适应性的影响和作用途径，为进一步探索极端环境中宿主植物——内生细菌抗逆性能的相互作用机制及协同进化研究奠定了基础。该项目的完成为丰富我国的微生物资源、推动干旱地区微生物资源的保护、开发和利用提供了依据。