Colin Yannick, Goñi-Urriza Marisol, Caumette Pierre, Guyoneaud Rémy
Equipe Environnement et Microbiologie, IPREM UMR CNRS 5254, Université de Pau et des Pays de l'Adour, IBEAS, BP 1155, 64013 Pau Cedex, France.
Equipe Environnement et Microbiologie, IPREM UMR CNRS 5254, Université de Pau et des Pays de l'Adour, IBEAS, BP 1155, 64013 Pau Cedex, France.
J Microbiol Methods. 2017 Jul;138:100-105. doi: 10.1016/j.mimet.2017.05.015. Epub 2017 May 30.
The development of new high-throughput cultivation methods aims to increase the isolation efficiency as compared to standard techniques that often require enrichment procedures to compensate the low microbial recovery. In the current study, estuarine sulfate-reducing bacteria were isolated using an anaerobic isolation procedure in 384-well microplates. Ninety-nine strains were recovered from initial sediments. Isolates were identified according to their partial 16S rRNA sequences and clustered into 13 phylotypes. Besides, the increase in species richness obtained through enrichments or resampling was investigated. Forty-four enrichment procedures were conducted and shifts in sulfate-reducing bacterial communities were investigated through dsrAB gene fingerprinting. Despite efforts in conducting numerous enrichment conditions only few of them were statistically different from initial sample. The cultural diversity obtained from 3 of the most divergent enrichments, as well as from resampled sediments equally contributed to raise the sulfate-reducing diversity up to 22 phylotypes. Enrichments (selection of metabolism) or resampling (transient populations and micro-heterogeneity) may still be helpful to assess new microbial phylotypes. Nevertheless, all the newly cultivated strains were all representatives of minor Operational Taxonomic Units and could eventually be recovered by maintaining high-throughput isolation effort from the initial sediments.
新的高通量培养方法的发展旨在提高分离效率，与通常需要富集程序来弥补低微生物回收率的标准技术相比。在当前研究中，使用384孔微孔板中的厌氧分离程序分离河口硫酸盐还原菌。从初始沉积物中回收了99株菌株。根据其部分16S rRNA序列对分离株进行鉴定，并聚类为13个系统发育型。此外，还研究了通过富集或重新采样获得的物种丰富度的增加。进行了44次富集程序，并通过dsrAB基因指纹图谱研究了硫酸盐还原细菌群落的变化。尽管在进行大量富集条件方面做出了努力，但只有少数条件与初始样品在统计学上有差异。从3种差异最大的富集培养物以及重新采样的沉积物中获得的培养多样性同样有助于将硫酸盐还原多样性提高到22个系统发育型。富集（代谢选择）或重新采样（瞬时种群和微异质性）可能仍然有助于评估新的微生物系统发育型。然而，所有新培养的菌株都是次要操作分类单元的代表，最终可以通过持续从初始沉积物中进行高通量分离来回收。
