In situ characterization of Nitrospira-like nitrite oxidizing bacteria active in wastewater treatment plants
In situ characterization of Nitrospira-like nitrite oxidizing bacteria active in wastewater treatment plants
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DOI：
10.1128/aem.67.11.5273-5284.2001
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发表时间：
2001-11-01
影响因子：
4.4
通讯作者：
Wagner, M
中科院分区：
文献类型：
--
作者：
Daims, H;Nielsen, JL;Wagner, M
用不依赖培养的分子方法对不同生物膜和活性污泥样品中未培养的类硝化细菌进行了研究。首先，通过16S rRNA基因序列分析，确定了硝化生物膜中类硝化螺旋菌的系统发育关系。随后，构建了包含所有公开序列的Nitrospira门的系统发育共识树。这一分析揭示了Nitrospira属至少由四个不同的亚系组成。基于这些数据，我们开发了两个分别针对Nitrospira门和属的16S rRNA寡核苷酸探针，并对其用于荧光原位杂交(FISH)的适用性进行了评估。利用FISH、激光共聚焦扫描显微镜和计算机辅助三维可视化技术，对污水处理厂类硝酸菌细胞聚集体的结构进行了原位研究。荧光素染色显示，硝酸螺旋菌微菌落内的空腔和无细胞通道网络是透水性的。采用FISH和显微放射自显影相结合的方法，研究了类硝化细菌在不同培养条件下对不同碳源的吸收。在好氧条件下，生物反应器样品中的类硝酸螺旋菌可以吸收无机碳(以HCO3-或CO2形式)和丙酮酸，但不能吸收醋酸盐、丁酸盐和丙酸，这表明这些细菌可以在丙酮酸存在的情况下进行混合营养生长。相比之下，在缺氧或厌氧条件下，类硝化螺旋菌对任何碳源的吸收都没有观察到。
Uncultivated Nitrospira-like bacteria in different biofilm and activated-sludge samples were investigated by cultivation-independent molecular approaches. Initially, the phylogenetic affiliation of Nitrospira-like bacteria in a nitrifying biofilm was determined by 16S rRNA gene sequence analysis. Subsequently, a phylogenetic consensus tree of the Nitrospira phylum including all publicly available sequences was constructed. This analysis revealed that the genus Nitrospira consists of at least four distinct sublineages. Based on these data, two 16S rRNA-directed oligonucleotide probes specific for the phylum and genus Nitrospira, respectively, were developed and evaluated for suitability for fluorescence in situ hybridization (FISH). The probes were used to investigate the in situ architecture of cell aggregates of Nitrospira-like nitrite oxidizers in wastewater treatment plants by FISH, confocal laser scanning microscopy, and computer-aided three-dimensional visualization. Cavities and a network of cell-free channels inside the Nitrospira microcolonies were detected that were water permeable, as demonstrated by fluorescein staining. The uptake of different carbon sources by Nitrospira-like bacteria within their natural habitat under different incubation conditions was studied by combined FISH and microautoradiography. Under aerobic conditions, the Nitrospira-like bacteria in bioreactor samples took up inorganic carbon (as HCO3- or as CO2) and pyruvate but not acetate, butyrate, and propionate,, suggesting that these bacteria can grow mixotrophically in the presence of pyruvate. In contrast, no uptake by the Nitrospira-like bacteria of any of the carbon sources tested was observed under anoxic or anaerobic conditions.