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通过定量寡核苷酸探针膜杂交揭示日本盐度分层湖凯伊克湖中的优势微生物组成及其垂直分布。

Dominant microbial composition and its vertical distribution in saline meromictic Lake Kaiike (Japan) as revealed by quantitative oligonucleotide probe membrane hybridization.

作者信息

Koizumi Yoshikazu, Kojima Hisaya, Fukui Manabu

机构信息

Department of Biological Sciences, Graduate School of Science, Tokyo Metropolitan University, Minami-ohsawa 1-1, Hachioji, Tokyo 192-0397, Japan.

出版信息

Appl Environ Microbiol. 2004 Aug;70(8):4930-40. doi: 10.1128/AEM.70.8.4930-4940.2004.

Abstract

Vertical distributions of dominant bacterial populations in saline meromictic Lake Kaiike were investigated throughout the water column and sediment by quantitative oligonucleotide probe membrane hybridization. Three oligonucleotide probes specific for the small-subunit (SSU) rRNA of three groups of Chlorobiaceae were newly designed. In addition, three general domain (Bacteria, Archaea, and Eukarya)-specific probes, two delta-Proteobacteria-specific probes, a Chlorobiaceae-specific probe, and a Chloroflexi-specific probe were used after optimization of their washing conditions. The abundance of the sum of SSU rRNAs hybridizing with probes specific for three groups of Chlorobiaceae relative to total SSU rRNA peaked in the chemocline, accounting for up to 68%. The abundance of the delta-proteobacterial SSU rRNA relative to total SSU rRNA rapidly increased just below the chemocline up to 29% in anoxic water and peaked at the 2- to 3-cm sediment depth at ca. 34%. The abundance of SSU rRNAs hybridizing with the probe specific for the phylum Chloroflexi relative to total SSU rRNA was highest (31 to 54%) in the top of the sediment but then steeply declined with depth and became stable at 11 to 19%, indicating the robust coexistence of sulfate-reducing bacteria and Chloroflexi in the top of the sediment. Any SSU rRNA of Chloroflexi in the water column was under the detection limit. The summation of the signals of group-specific probes used in this study accounted for up to 89% of total SSU rRNA, suggesting that the DGGE-oligonucleotide probe hybridization approach, in contrast to conventional culture-dependent approaches, was very effective in covering dominant populations.

摘要

通过定量寡核苷酸探针膜杂交技术,对盐度分层的凯伊克湖整个水柱和沉积物中优势细菌种群的垂直分布进行了研究。新设计了三种针对绿菌科三组小亚基(SSU)rRNA的寡核苷酸探针。此外,在优化洗涤条件后,使用了三种通用域(细菌、古菌和真核生物)特异性探针、两种δ-变形菌特异性探针、一种绿菌科特异性探针和一种绿弯菌门特异性探针。与绿菌科三组特异性探针杂交的SSU rRNA总和相对于总SSU rRNA的丰度在化学跃层达到峰值,占比高达68%。δ-变形菌SSU rRNA相对于总SSU rRNA的丰度在化学跃层下方的缺氧水中迅速增加,最高可达29%,并在沉积物2至3厘米深度处达到约34%的峰值。与绿弯菌门特异性探针杂交的SSU rRNA相对于总SSU rRNA的丰度在沉积物顶部最高(31%至54%),但随后随深度急剧下降并稳定在11%至19%,这表明硫酸盐还原菌和绿弯菌在沉积物顶部强烈共存。水柱中绿弯菌的任何SSU rRNA均低于检测限。本研究中使用的组特异性探针信号总和占总SSU rRNA的比例高达89%,这表明与传统的依赖培养的方法相比,DGGE-寡核苷酸探针杂交方法在覆盖优势种群方面非常有效。

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