分离自印度洋深海沉积物的部分链霉菌分离株多位点序列分析

Multilocus sequence analysis (MLSA) of Streptomyces strains isolated from the Indian Ocean sediment samples

摘要:

【目的】采用多位点序列分析方法,研究印度洋3 000 m 以下深海沉积物中分离得到的16S rRNA 基因比对高度相似的链霉菌菌株的种间系统发育关系,同时探讨各管家基因及多基因聚类分析后的种间区分能力。【方法】以分离自印度洋深海沉积物的7 株Streptomyces albidoflavus,11 株Streptomyces cavourensis,16 株Streptomyces pratensis 为研究对象,以16SrRNA、atpD、recA 和rpoB 基因片段为标记,通过PCR 扩增、测序,获得序列。同时从NCBI上下载5 株S. pratensis 上述4 个基因的序列,将所有序列在MLST 网站进行比对,并构建系统进化树进行比较。【结果】S. pratensis 各菌株种内比较发现,16S rRNA 基因构建的系统进化树中相同基因型的菌株没有聚在一起,系统进化树不稳定,区分度不高。其余3 个构建的系统进化树稳定,菌株的聚类关系与MLST 数据库得到的基因型一致。同时,多基因聚类分析后将菌株分为6个类群。在3个种的种间多位点序列比较中,除区分度明显增加、进化树更加稳定以外,还发现recA 基因进化上比较特殊的菌株。【结论】多位点序列分析将实验菌株分为很多不同的类型,成功地将所分离的链霉菌进行了更细的分类,同时也找到部分菌株在个别基因上差异较大。此方法可以用于相近种的快速鉴定。

Abstract:

[Objective] The objective of this study was to apply multilocus sequence analysis (MLSA) to evaluate the phylogenetic relationship among highly similar strains of the genus Streptomyces isolated from the Indian Ocean sediment samples under the depth of 3 000 m, based on 16S rRNA, atpD, recA and rpoB genes, and to further explore the potential of isolated Streptomyces species. [Methods] We cultured 7 strains of Streptomyces albidoflavus, 11 strains of Streptomyces cavourensis, 16 strains of Streptomyces pratensis isolated from the Indian Ocean deep-sea sediments. We used PCR amplification and sequenced the genes 16S rRNA, atpD, recA and rpoB from these strains and also downloaded 5 strains sequences of S. pratensis from NCBI. We used all sequences to find the gene types in MLST web site and to build the phylogenetic trees for comparison. [Results] After comparison of S. pratensis, the same genotype strains were not clustered together in phylogenetic tree based on 16S rRNA gene sequence, the tree was not stable and degree of differentiation was also low, the phylogenetic trees based on atpD, recA, rpoB genes were relatively stable. Strains in the phylogenetic tree based on multiple gene cluster analysis divided into six groups. In Multilocus sequence analysis of the three Streptomyces, the degree of differentiation increased and the phylogenetic tree was more stable and also found some special strains of recA gene. [Conclusion] Multilocus sequence analysis can divide experimental strains into different types and some strains have large differences in individual gene (recA gene). Multilocus sequence analysis is suitable for classification and identification of some closely related species.