黄河三角洲滨海湿地非培养放线菌多样性

Actinobacterial diversity in a coastal wetland of the Yellow River Delta revealed by culture-independent approach

【目的】利用免培养技术对黄河三角洲滨海湿地土壤中放线菌多样性进行分析。【方法】提取样品总DNA,利用放线菌门特异引物扩增放线菌16S rRNA基因序列,构建放线菌16S rRNA基因克隆文库,文库经RFLP分析后挑选代表序列测序并进行多样性指数分析和系统发育分析。【结果】构建的放线菌16S rRNA基因克隆文库覆盖率(C)为96.3%,116个测序序列可化分为46个OTUs,58.7%的OTUs(27个)存在于放线菌亚纲(Actinobacteridae)放线菌目(Actinomycetales)的7个亚目中,分布于10个科中,其中弗兰克氏菌亚目(Frankineae)最多,共7个OTUs,有30.4%的OTUs(14个)存在于酸微菌亚纲(Acidimicrobidae)醋微菌亚目(Acidimicrobineae)中,没有发现与红色杆菌亚纲(Rubrobacteridae)和科里氏杆菌亚纲(Coriobacteridae)亲缘关系较近的序列。有10.9%的OTUs序列与有效发表的所有类群无亲缘关系,在进化树上成为一个独立的进化分支,有可能代表新亚目或更高级分类单元的类群。【结论】黄河三角洲滨海湿地蕴含着丰富的放线菌物种多样性及潜在的放线菌新类群,具有深一步研究的价值。

[Objective] Using culture-independent method to analyze the diversity of actinomyces from Yellow River Delta coastal wetland. [Methods] Actinobacterium-specific 16S rRNA gene clone library was constructed. Representative clones selected based on the RFLP analysis were sequenced and placed into operational taxonomic unit （OTU） groups according to the 16S rRNA sequence similarity. Diversity statistics were analyzed using SPADE analysis software. [Results] 324 sequences were placed into 46 OTUs. 58.7% of the OTUs belonged to Actinobacteridae and 7 suborders of Actinomycetales. The dominant actinobacteria was Frankineae, which accounted for 7 OTUs. 30.4% of the OTUs belonged to subclass Aci-dimicrobidae and suborder Acidimicrobineae. 10.9% of the OTUs shared no close relationship with published sequences. Some sequences which formed a distinct clade in phylogenetic tree may represent new taxonomical groups of actinomyces. [Conclusion] Yellow River Delta coastal wetland harbors abundant and diverse actinomyces. The result also implied that there may be large numbers of unknown actinobacterial groups.