长期稻-稻-绿肥轮作对水稻根际微生物多样性及功能群落的影响

The application of green manures to rice field is a traditional agricultural practice in China, especially in southern-central part of China. It has been demonstrated that the green manure can increase sustainability of soil eco-system by reducing soil erosion, ameliorating soil physical properties and promote soil fertility; it plays an important role in improving rice yield and maintaining the stability of soil eco-system. However, the effects of green manure on microbial diversity and functional communities in the rhizosphere of succession crops are still not investigated extensively and systematically and our knowledge on the mechanism of effects is still very limited so far. This project will focus on the studies of the effects of long-term winter-growing green manure plants on microbial diversity and nitrogen-cycle related functional communities (e.g.: nitrogen-fixing bacteria and ammonia oxidation microbes) in the rhizosphere of rice based on a 30-year long-term experiments in the Key Field Monitoring Experimental Station for Red Soil Eco-environment of Ministry of Agriculture, Qiyang County of Hunan Province. Four treatments will be setup as rice-rice-winter fallow, rice-rice-Chinese milk vetch, rice-rice-rape and rice-rice-ryegrass and these three different kinds of plants will be used as the three types of green-manure applied commonly in the rice field: Leguminosae, Cruciferae and Gramineae. To look into the effects of long-term rice-rice-green manure rotation on microbial diversity and function communities in the rhizosphere of rice, Culture-independent methods based on the analysis of 16S rRNA, nifH, amoA and amoB genes and traditional Culture-dependent methods will be used cooperatively in this study. Phylogenetics of cultured bacteria by using the method of polyphasic taxonomy; characteristics of cultured strains by testing the production of IAA, phosphate-dissolving ability, nitrogen-fixing and ammonia-oxidizing will be investigated as well. The results of this project will be important for us to better understand the functions of green manure and its effects on the interactions between microorganisms and rice plant. It will be very helpful for us to achieve some rice growth-promotion related microbial resources.

大量研究证实，绿肥是保持和提高土壤肥力和质量的有效措施之一，虽然已有绿肥对土壤微生物影响的相关研究报道，但由于方法和技术所限，相关研究不够系统深入，特别是绿肥对水稻根际微生物影响的研究尚属空白。本课题拟以湖南祁阳农田系统国家野外科学观测研究站具有30年长期定位历史的稻-稻-绿肥（豆科植物紫云英、禾本科植物黑麦草和十字花科植物油菜）及稻-稻-冬闲轮作水稻系统为材料，采用基于16S rRNA、nifH、amoA等基因为基础的先进的分子生态学方法与传统的培养方法相结合，深入研究不同绿肥轮作水稻根际微生物的多样性、氮循环重要的微生物类群（固氮和氨氧化微生物）组成，及其对一些功能性微生物的影响；同时针对性地分离培养微生物类群。研究结果可为长期轮作绿肥对水稻产量的影响提供理论依据，并有利于揭示微生物与植物的相互作用关系，同时将获得提高水稻产量的有益微生物资源。

研究证实，绿肥是保持和提高土壤肥力和质量的有效措施之一，虽然已有绿肥对土壤微生物影响的相关研究报道，但由于方法和技术所限，相关研究不够系统深入，特别是绿肥对水稻根际微生物影响的研究尚属空白。本课题以湖南祁阳农田系统国家野外科学观测研究站具有30年长期定位历史的稻-稻-绿肥及稻-稻-冬闲轮作水稻系统为材料，采用基于16S rRNA、nifH、amoA等基因为基础的先进的分子生态学方法与传统的培养方法相结合，研究不同绿肥轮作水稻根际微生物的多样性、氮循环重要的微生物类群组成；针对性地分离培养微生物类群。通过本项目的研究获得主要结果如下：1、长期绿肥轮作显著增加了水稻根际细菌的数量，而不同绿肥处理间根际微生物数量没有差异。α-多样性分析表明，绿肥轮作降低了根际微生物的多样性及丰度，β-多样性分析表明长期绿肥轮作显著的改变了水稻根际的微生物区系,这种变化是土壤和植物特性共同作用的结果。在轮作水稻根际，植物促生类群Pseudomonas和 Acinetobacter 得以富集；2、比较长期紫云英绿肥与冬闲轮作的水稻根内生菌，发现长期与紫云英绿肥轮作水稻根的内生细菌数量显著增多，且种类更加丰富；3、氨氧化微生物的相关基因分析发现，长期绿肥轮作土壤及水稻根际AOB的数量得以增加，施用绿肥土壤中AOA的拷贝数低于冬闲，但在根际则没有明显的规律；4、本研究共获得微生物资源300余株，对179株优势种进行促生功能初筛，获得80株具有溶解无机磷活性的菌株，64株具有分泌铁载体能力，93株具有分泌IAA功能。对40株初筛结果较好的菌株进行复筛，28株拮抗串珠镰刀菌，5株固氮菌株； 5、获得了多个水稻内生及根际细菌的新类群，已发表8株：Rhizobium oryziradicis、Paenibacillus oryzae、Novosphingobium oryzae、Rhizobium oryzicola、Paenibacillus rhizoryzae、Paenibacillus oryzisoli、Bacillus oryzisoli、Rhizobium rhizoryzae。研究结果为长期轮作绿肥增加水稻水稻产量提供了理论依据，并揭示了该生态系统中，土壤、微生物与植物三者的相互作用关系，同时将获得提高水稻产量的有益微生物资源。