水稻种子内生核心细菌的界定及其系统进化和功能特性研究

The importance of plant-associated microorganisms for plant growth and health has recently received greater recognition. It has been demonstrated that bacteria inhabiting seeds form an important group among the plant-associated bacteria. However, the role of seed-associated microorganisms, and especially seed endophytic bacteria, still is under-studied. Until now, only limited research has been conducted to unravel the composition and the exact role of seed endophytes. This project will focus on the definition, classification and functional characterization of the core endophytic bacteria microbiome of rice seeds using both emerging culture-independent and traditional culture-dependent methods. Seven rice varieties were carefully chosen, including japonica rice, indica rice and wild rice. The selected seeds will be planted in three experimental stations at the Chinese Academy of Agricultural Sciences (Langfang, Hebei; Qiyang, Hunan; Wenchang, Hainan) and one experimental station at the Heilongjiang Academy of Agricultural Sciences (Harbin). To define the core microbime of the rice seeds at different levels of ecological hierarchy, the endophytic bacterial composition of rice seeds will be investigated by Illumina next-generation sequencing of the 16S rRNA gene. Based on these results, three kinds of rice seeds, representing japonica rice, indica rice and wild rice, will be used to isolate culturable bacteria. The isolated bacteria that belong to the predominant core microbime of the rice seeds will be investigated on their phylogenetics by using the method of polyphasic taxonomy. Plant beneficial characteristics of cultured strains will include testing the production of IAA, the phosphate-dissolving ability, the nitrogen-fixing ability, as well as the production of 1-aminocyclopropane-1-carboxylate (ACC) deaminase and siderophores. The results of this project will be critical for us to better understand the role of bacterial communities in seeds and their contribution to their host. It will be very helpful for us to achieve some rice growth-promotion related microbial resources.

植物微生物已成为近年来的研究热点之一。然而，作为植物微生物的重要组成部分，种子微生物的研究却十分有限，特别是种子内生细菌的研究较为滞后。虽然对于核心微生物组的界定是植物微生物研究的难题之一，但由于种子微生物具有选择性转移到下一代的可能性，使得种子成为研究植物核心微生物组的理想材料。本研究在申请人前期研究水稻种子内生细菌多样性工作的基础上，选取我国主要栽培稻—粳稻和籼稻亚种的6个品种以及东乡野生稻为研究材料，分别在河北、湖南、海南和黑龙江进行种植，通过16S rRNA基因高通量测序，探讨不同地理区域水稻种子内生细菌群落结构特征，以界定不同生态尺度下的核心微生物组；在非培养研究结果基础上，选择代表性品种进行种子内生核心细菌地定向分离，确定其分类地位及不同来源菌株间的系统进化关系；探索核心类群菌株的植物促生功能。不但为植物与微生物间的共进化理论提供依据，也可为新型微生物接种剂的研制提供新资源。

植物微生物已成为近年来的研究热点之一。然而，作为植物微生物的重要组成部分，种子微生物的研究却十分有限，特别是种子内生细菌的研究较为滞后。虽然对于核心微生物组的界定是植物微生物研究的难题之一，但由于种子微生物具有选择性地转移到下一代的可能性，使得种子成为研究植物核心微生物组的理想材料。本研究选取我国主要栽培稻—粳稻和籼稻亚种的6个水稻品种，分别在河北、云南、海南和江西进行种植，通过16S rRNA高通量测序，系统调查不同地理区域水稻种子内生细菌群落结构，结果发现；水稻基因型与环境对水稻种子内生细菌的组成具有显著的影响，相对于基因型，环境变量影响更大；水稻种子内生细菌主要来源与上一代种子；根据高通量测序结果，水稻种子内生100%共有类群的OTUs有13个，其中主要为泛菌（Pantoea），进一步网络分析也证实泛菌属为关键核心类群；可培养方法研究发现，从不同地区的水稻种子中，除泛菌外，大量分离到糖黄单胞菌（Xanthomonas Sacchari）；因此初步确定，水稻内生核心微生物为泛菌属和糖黄单胞菌。进一步通过基因组分析和植物促生功能测定，确定其系统发育分析及植物促生有益特性。相关研究为植物与微生物间的共进化理论提供依据，也可为新型微生物接种剂的研制提供新资源和新思路。