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Molecular Ecological Approach to Sulfur-Metabolizing Bacteria in Aquatic Ecosystems

水生生态系统中硫代谢细菌的分子生态学方法

基本信息

批准号：
12440219
负责人：
FUKUI Manabu
金额：
$ 9.28万
依托单位：
Tokyo Metropolitan University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12440219/
关键词：
16S rRNA DGGE Sulfur-oxidizing bacteria freshwater sediment Thioploca Lake Biwa Sulfur cycle Sulfate reduction 16Sr RNA Desulfonema 16S rDNA 16SrRNA

项目摘要

Molecular ecological studies were done for sulfur-metabolizing microorganisms in various aquatic environments. Among these, freshwater filamentous gliding sulfide-oxidizing bacteria of the genus Thioploca were found on sediments in profundal areas of Lake Biwa, a Japanese freshwater mesotrophic lake, and characterized morphologically, ecologically and phylogenetically. The Lake Biwa Thioploca resembled morphologically Thioploca ingrica, a brackish water species from a Danish fjord. The diameter of individual trichomes was 3-5.6 μm ; the diameter of complete Thioploca filaments was in the range of 18-75 μm. Cell lengths ranged from 1.2 to 3.8 μm. In TEM specimens stained with uranyl acetate, dense intracellular particles were found, which did not show any positive signal of phosphorus and sulfur in the X-ray analysis. The 16S rRNA gene of Thioploca from Lake Biwa was amplified using newly designed Thioploca-specific primers (706-Thioploca, Biwa160F and Biwa829R) in combination with general bacterial primers, to avoid unspecific amplification of contaminating bacterial DNA. DGGE analysis of all three overlapping PCR products resulted in single DGGE bands, indicating that a single 16S rRNA gene has been amplified. With the same method, Thioploca from Lake Constance was examined. The 16S rRNA sequence was verified with FISH hybridization targeted at specific motifs of Lake Biwa Thioploca. Positive signals were obtained for the bacterial probe EUB-338, the γ-proteobacterial probe GAM42a, and the probe Biwa829 targeting Lake Biwa Thioploca. Based on the near-complete 16S rRNA sequence and on morphological similarities, Thioploca from Lake Biwa and Lake Constance are closely related to Thioploca ingrica and to each other.

对不同水环境中的硫代谢微生物进行了分子生态学研究。其中，淡水丝状滑翔硫化物氧化细菌属Thioploca被发现在沉积物中的深海区的琵琶湖，日本淡水中营养湖，其特征形态，生态和水生。琵琶湖的Thioploca在形态上类似于Thioploca ingrica，一种来自丹麦峡湾的半咸水物种。单个毛状体的直径为3-5.6 μm ;完整的Thioploca细丝的直径在18-75 μm范围内。细胞长度范围为1.2 - 3.8 μm。电镜下醋酸双氧铀染色可见细胞内致密颗粒，X线分析未见磷、硫阳性信号。为了避免污染细菌DNA的非特异性扩增，采用新设计的Thioploca特异性引物（706-Thioploca，Biwa 160 F和Biwa 829 R）结合细菌通用引物，对琵琶湖Thioploca的16 S rRNA基因进行了扩增。所有三个重叠的PCR产物的DGGE分析导致单个DGGE条带，表明单个16 S rRNA基因已被扩增。用同样的方法，对博登湖的Thioploca进行了检测。16 S rRNA序列进行了验证与FISH杂交针对特定图案的琵琶湖Thioploca。细菌探针EUB-338、γ-变形菌探针GAM 42 a和靶向琵琶湖Thioploca的探针Biwa 829获得阳性信号。基于接近完整的16 S rRNA序列和形态相似性，来自琵琶湖和博登湖的Thioploca与Thioploca ingrica密切相关。