九龙江河口区nirS型反硝化细菌多样性及系统发育学分析

Phylogenetic diversity of nirS-type denitrifying bacteria in Jiulong River estuary

摘要:

【目的】结合16S rRNA基因克隆文库和nirS基因克隆文库的分析，揭示九龙江河口区nirS型反硝化细菌多样性。【方法】选取九龙江河口区一富营养化采样点，分别采集水样及沉积物样品，进行理化因子的测定并提取细菌总DNA。以水样DNA构建16S rRNA基因克隆文库，以沉积物DNA构建nirS基因克隆文库，分析微生物群落结构的多样性并构建系统发育树。【结果】从16S rRNA基因克隆文库中获得86条有效序列，按97%的序列相似性划分为53个OTU，分别属于Proteobacteria门、Planctomycetes门、Bacteroidetes门、Actinobacteria门、Firmicutes门和Chloroflexi门。其中属于Proteobacteria门OTU的克隆子占克隆数的62.9%，是最优势的类群，分属于Alphaproteobacteria、Betaproteobacteria、Gammaproteobacteria和Deltaproteobacteria纲等。从nirS基因克隆文库中获得190条有效序列，翻译为氨基酸序列后，按82%的序列相似性划分为60个OTU，并定位到属的水平。其中Proteobacteria门是最优势的类群，占文库克隆子总数的71.6%，包括Alphaproteobacteria纲(5.8%)、Betaproteobacteria纲(49.0%)和Gammaproteobacteria纲(16.9%)。nirS基因克隆文库中丰度最高的OTU与GenBank中的一株可培养反硝化菌Thauera sp. R-26906具有100%的序列相似性。【结论】九龙江河口区的微生物以及亚硝酸盐还原酶基因(nirS)具有丰富的多样性。大部分NirS序列在GenBank中的最相似序列来源于河口、海湾等相似的环境。

Abstract:

[Objective] To understand the microbial community of denitrifying bacteria in Jiulong River estuary. [Methods] The sampling site was in Jiulong River estuary that was characterized by eutrophication. The water sample and sediment sample were collected and the environmental physicochemical factors were analyzed. The 16S rRNA gene and nirS gene clone libraries were constructed based on water sample and sediment sample respectively. [Results] In the 16S rRNA gene clone library, 86 valid sequences were grouped into 53 OTUs based on 97% sequence similarity and clustered into 6 different bacterial groups on the phylum level: Proteobacteria (Alpha, Beta, Gamma and Delta), Planctomycetes, Bacteroidetes, Actinobacteria, Firmicutes and Chloroflexi. Among these groups, Proteobacteria was the predominant member (62.9%). Of the nirS gene clone library 190 valid sequences were translated to conceptual amino acid sequences and group into 60 OTUs based on 82% similarity for genus identification. Among the 190 sequences, 71.6% were clustered into Proteobacteria phylum, including Alphaproteobacteria (5.8%), Betaproteobacteria (49.0%) and Gammaproteobacteria (16.9%). The most abundant OTU shared 100% sequence similarity to sequences from the nitrite reductase of Thauera sp. R-26906, a species of cultured denitrifier. [Conclusion] High diversity of the microbial community as well as nirS genes were found in the Jiulong River estuary. Most of the NirS sequences of this study shared quite high similarity to known sequences from various similar environments such as estuary and gulf.