Nübel Ulrich, Bateson Mary M, Vandieken Verona, Wieland Andrea, Kühl Michael, Ward David M
Department of Land Resources and Environmental Sciences, Montana State University, Bozeman, Montana, USA.
Appl Environ Microbiol. 2002 Sep;68(9):4593-603. doi: 10.1128/AEM.68.9.4593-4603.2002.
We investigated the diversity, distribution, and phenotypes of uncultivated Chloroflexaceae-related bacteria in photosynthetic microbial mats of an alkaline hot spring (Mushroom Spring, Yellowstone National Park). By applying a directed PCR approach, molecular cloning, and sequence analysis of 16S rRNA genes, an unexpectedly large phylogenetic diversity among these bacteria was detected. Oligonucleotide probes were designed to target 16S rRNAs from organisms affiliated with the genus Chloroflexus or with the type C cluster, a group of previously discovered Chloroflexaceae relatives of this mat community. The application of peroxidase-labeled probes in conjunction with tyramide signal amplification enabled the identification of these organisms within the microbial mats by fluorescence in situ hybridization (FISH) and the investigation of their morphology, abundance, and small-scale distribution. FISH was combined with oxygen microelectrode measurements, microscope spectrometry, and microautoradiography to examine their microenvironment, pigmentation, and carbon source usage. Abundant type C-related, filamentous bacteria were found to flourish within the cyanobacterium-dominated, highly oxygenated top layers and to predominate numerically in deeper orange-colored zones of the investigated microbial mats, correlating with the distribution of bacteriochlorophyll a. Chloroflexus sp. filaments were rare at 60 degrees C but were more abundant at 70 degrees C, where they were confined to the upper millimeter of the mat. Both type C organisms and Chloroflexus spp. were observed to assimilate radiolabeled acetate under in situ conditions.
我们研究了碱性温泉（黄石国家公园蘑菇泉）光合微生物垫中未培养的绿弯菌科相关细菌的多样性、分布和表型。通过应用定向PCR方法、分子克隆和16S rRNA基因序列分析，检测到这些细菌中存在意想不到的大量系统发育多样性。设计了寡核苷酸探针，以靶向与绿弯菌属或C型簇相关的生物体的16S rRNA，C型簇是该垫群落中一组先前发现的绿弯菌科亲属。将过氧化物酶标记的探针与酪胺信号放大相结合，通过荧光原位杂交（FISH）在微生物垫中鉴定这些生物体，并研究它们的形态、丰度和小尺度分布。FISH与氧微电极测量、显微镜光谱分析和微放射自显影相结合，以检查它们的微环境、色素沉着和碳源利用情况。发现大量与C型相关的丝状细菌在蓝藻主导的、高度氧化的顶层中大量繁殖，并在被研究的微生物垫较深的橙色区域中数量占主导，这与细菌叶绿素a的分布相关。绿弯菌属的丝状体在60摄氏度时很少见，但在70摄氏度时更为丰富，它们局限于垫子的上毫米处。在原位条件下，观察到C型生物体和绿弯菌属都能同化放射性标记的乙酸盐。
