基于荧光定量PCR和高通量测序技术的葡萄园土壤细菌群落结构多样性分析

Analysis of Bacterial Community Structure in Vineyard Soil by RT-PCR and High Throughput Sequencing

土壤细菌在葡萄园生态系统中起着重要作用，具有十分重要的生态功能。以贵州镇远葡萄园基地根际、非根际土壤为研究对象，在对土壤的基本理化指标及酶活检测的基础上，采用荧光定量PCR（Q-PCR）和Illumina Miseq高通量测序技术对葡萄园根际和非根际土壤的细菌菌群结构进行分析。结果表明，葡萄园根际土壤细菌16S rRNA基因的拷贝数在1.38×108～1.17×109 copies/g之间，高于非根际土壤。Illumina Miseq高通量测序显示，葡萄园根际土壤细菌呈多样性分布且优势菌群突出，主要涵盖了变形菌门（Protecbacteria），放线菌门（Actinbacteria），酸杆菌门（Acidobacteri），厚壁菌门（Firmicute），绿弯菌门（Chloroflexi），蓝藻门（Cyanobacteria），泉古菌门（Crenar-choeo），拟杆菌门（Bacteroidetes），芽单胞菌门（Gemmatimonadete），硝化螺旋菌门（Nitrospirae）。其中变形菌门和放线菌门为优势菌群，两者相对丰度之和占到了53.33%～67.28%。葡萄园根际土壤的主要优势细菌属包括乳球菌属（Lactococcus），节细菌属（Arthrobacter），Candidatus nitrososphaera，亚硝化螺菌属（Nitrososphaera），热单胞菌属（Thermomonas），Phycicoccus，泛菌属（Pantoea），芽孢杆菌属（Bacillus），埃希氏杆菌属（Escherichia），鞘脂单胞菌属（Sphingomonas），和溶杆菌属（Lysobacter）。α多样性分析表明，不同位点的根际土壤细菌多样性比较接近，而根际土壤细菌多样性低于非根际土壤。冗余分析（RDA）进一步分析表明，葡萄园土壤细菌群落多样性与土壤养分及酶活存在不同程度的相关关系，群落结构多样性受多种因素的影响。

Soil bacteria have important ecological functions and play a very important role in vineyard ecological system. In this study, we per-formed quantitative PCR （Q-PCR） and Illumina Miseq high-throughput sequencing to compare the basic physiochemical parameters, enzymat-ic activities and bacterial groups between the rhizosphere and non-rhizosphere soil from a vineyard from Zhenyuan, Guizhou. Q-PCR results in-dicated that, the bacterial 16S rRNA gene copy numbers in the rhizosphere soils ranged from 1.38×108 to 1.17×109 copies/g, higher than that in non-rhizosphere soil. Illumina Miseq high-throughput sequencing results indicated that, the bacterial communities in rhizosphere vineyard soil mainly included Protecbacteria, Actinbacteria, Acidobacteria, Firmicute, Chloroflexi, Cyanobacteria, Crenarchoeo,Bacteroidetes,Gemmatimon-adete, and Nitrospirae, among them, the dominant bacterial taxa members were Proteobacteria and Actinobacteria which accounted for 53.33%~67.28%of total abundance. At genus level, Lactococcus, Arthrobacter, Candidatus Nitrososphaera, Nitrososphaera, Thermomonas, Phycicoccus, Pantoea, Bacillus, Escherichia, Sphingomonas, Lysobacter were the dominant bacteria genera in rhizosphere vineyard soil.α-di-versity analysis showed that, the bacterial community diversity of rhizosphere soil was similar in different site but were lower than that of the non-rhizosphere soil. Redundancy analysis showed that, there were certain correlations between bacterial communities and nutrients and en-zyme activity in vineyard soil..