16S rDNA-based analysis of dominant bacterial populations associated with early life stages of coho salmon (Oncorhynchus kisutch)
16S rDNA-based analysis of dominant bacterial populations associated with early life stages of coho salmon (Oncorhynchus kisutch)
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DOI：
10.1007/s00248-006-9037-9
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发表时间：
2006-05-01
影响因子：
3.6
通讯作者：
Navarrete, Paola
中科院分区：
文献类型：
--
作者：
Romero, Jaime;Navarrete, Paola
在这项研究中，我们使用了16 S rDNA为基础的方法来确定与银鲑（Oncorhynchus kisutch）在其早期生命阶段的细菌种群，突出在淡水中生长过程中的胃肠道中的优势细菌。本文是第一个银鲑细菌群落的分子分析。通过比较直接显微镜计数（使用吖啶橙子）与菌落计数（在胰蛋白胨大豆琼脂中）来估计鲑鱼胃肠道微生物群的可培养性。一般而言，低部分（约1%）的微生物群可以作为可培养细菌回收。采用聚合酶链反应-变性梯度凝胶电泳（PCR-DGGE）技术，从同一批次的个体中直接提取DNA，对初喂鱼苗和幼鱼的卵和胃肠道中的细菌群落进行了监测。DGGE图谱显示了所有阶段的简单群落，并暴露了生长过程中细菌群落的变化。切下的DGGE条带的测序和系统发育分析揭示了在每个阶段中发现的主要细菌的性质。在鸡蛋中，优势细菌属于β-变形菌（Janthinobacterium和Rhodoferax）。在第一次进食阶段，胃肠道中最丰富的细菌与γ-变形菌（希瓦氏菌和气单胞菌）聚集。在2至15 g的青少年中，主要细菌为假单胞菌和气单胞菌。为了确定在幼年胃肠道中发现的优势假单胞菌和气单胞菌的假定起源，基于从DGGE凝胶中检索的序列设计了这些组的引物。随后，通过PCR扩增分析水流入物、颗粒饲料和蛋的样品。只有从鸡蛋和进水样品中获得的扩增子与DGGE的优势条带序列相同。总的来说，我们的研究结果表明，在第一个喂养阶段后建立了稳定的微生物群，其主要成分可能来自水和卵表观生物群。
In this study, we used a 16S rDNA-based approach to determine bacterial populations associated with coho salmon (Oncorhynchus kisutch) in its early life stages, highlighting dominant bacteria in the gastrointestinal tract during growth in freshwater. The present article is the first molecular analysis of bacterial communities of coho salmon. Cultivability of the salmon gastrointestinal microbiota was estimated by comparison of direct microscopic counts (using acridine orange) with colony counts (in tryptone soy agar). In general, a low fraction (about 1%) of the microbiota could be recovered as cultivable bacteria. Using DNA extracted directly from individuals belonging to the same lot, bacterial communities present in eggs and gastrointestinal tract of first-feeding fries and juveniles were monitored by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). The DGGE profiles revealed simple communities in all stages and exposed changes in bacterial community during growth. Sequencing and phylogenetic analysis of excised DGGE bands revealed the nature of the main bacteria found in each stage. In eggs, the dominant bacteria belonged to beta-Proteobacteria (Janthinobacterium and Rhodoferax). During the first feeding stage, the most abundant bacteria in the gastrointestinal tract clustered with gamma-Proteobacteria (Shewanella and Aeromonas). In juveniles ranging from 2 to 15 g, prevailing bacteria were Pseudomonas and Aeromonas. To determine the putative origin of dominant Pseudomonas and Aeromonas found in juvenile gastrointestinal tracts, primers for these groups were designed based on sequences retrieved from DGGE gel. Subsequently, samples of the water influent, pelletized feed, and eggs were analyzed by PCR amplification. Only those amplicons obtained from samples of eggs and the water influent presented identical sequences to the dominant bands of DGGE. Overall, our results suggest that a stable microbiota is established after the first feeding stages and its major components could be derived from water and egg epibiota.