Population analysis in a denitrifying sand filter: Conventional and in situ identification of Paracoccus spp. in methanol-fed biofilms
Population analysis in a denitrifying sand filter: Conventional and in situ identification of Paracoccus spp. in methanol-fed biofilms
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DOI：
10.1128/aem.62.12.4329-4339.1996
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发表时间：
1996-12-01
影响因子：
4.4
通讯作者：
Schleifer, KH
中科院分区：
文献类型：
--
作者：
Neef, A;Zaglauer, A;Schleifer, KH
采用常规技术和分子生物学技术对某城市污水处理厂的砂滤池微生物群落进行了研究，以确定参与硝酸盐去除的活性菌群。在该系统中，反硝化作为水处理的最后一步，通过生长在石英颗粒上的生物膜，以甲醇作为补充碳源进行，生物膜是非常不规则的，具有13至20 μ m的中值厚度。通过对56株砂滤器分离株的脂肪酸分析，确定了砂滤器中存在副球菌属，并设计了该属和Paracoccus spp. -Paracoccus versutus种簇的16 SrRNA探针，通过全细胞杂交进行了特异性检测，通过使用甲酰胺浓度梯度来调节探针的严格性要求，以实现甚至高度相似的靶序列的完全区分，全细胞杂交证实，属副球菌的成员是丰富的分离物。56个分离株中有27个与属特异性探针杂交，原位杂交在分离的生物膜中鉴定出密集的副球菌聚集体，与P、P. versutus和P. ficans模式株互补的探针与生物膜中的细胞不杂交，提示砂滤器中存在一个新的副球菌种，使用共聚焦激光扫描显微镜的分析检测到形态相同的细胞的球形聚集体，其表现出均匀的荧光，在彻底破坏生物膜并过滤到聚碳酸酯过滤器上后进行细胞定量。平均3.5%的总细胞计数对应于副球菌属，而在没有补充甲醇且没有可测量的反硝化作用的平行砂滤器中，仅能检测到非常少的paracocci（0.07%的细胞被4 ′，6-二脒基-2-苯基吲哚染色）。Hyphomicrobium spp，构成约2%的所有细胞的净化单元，并不能检测到在常规砂滤器，这种明确的联系，在原位丰度和反硝化表明一个积极参与的过程中的paracocci和hyphomicrobia，可能的选择优势，有利于paracocci在这个栖息地进行了讨论。
The microbial community of a denitrifying sand filter in a municipal wastewater treatment plant was examined by conventional and molecular techniques to identify the bacteria actively involved in the removal of nitrate, In this system, denitrification is carried out as the last step of water treatment by biofilms growing on quartz grains with methanol as a supplemented carbon source, The biofilms are quite irregular, having a median thickness of 13 to 20 mu m. Fatty acid analysis of 56 denitrifying isolates indicated the occurrence of Paracoccus spp, in the sand filter, 16S rRNA-targeted probes were designed for this genus and the species cluster Paracoccus denitrificans-Paracoccus versutus and tested for specificity by whole-cell hybridization, Stringency requirements for the probes were adjusted by use of a formamide concentration gradient to achieve complete discrimination of even highly similar target sequences, Whole-cell hybridization confirmed that members of the genus Paracoccus were abundant among the isolates. Twenty-seven of the 56 isolates hybridized with the genus-specific probes, In situ hybridization identified dense aggregates of paracocci in detached biofilms, Probes complementary to the type strains of P, denitrificans and P. versutus did not hybridize to cells in the biofilms, suggesting the presence of a new Paracoccus species in the sand filter, Analysis using confocal laser scanning microscopy detected spherical aggregates of morphologically identical cells exhibiting a uniform fluorescence, Cell quantification was performed after thorough disruption of the biofilms and filtration onto polycarbonate filters, An average of 3.5% of total cell counts corresponded to a Paracoccus sp,, whereas in a parallel sand filter with no supplemented methanol, and no measurable denitrification, only very few paracocci (0.07% of cells stained with 4',6-diamidino-2-phenylindole) could be detected. Hyphomicrobium spp, constituted approximately 2% of all cells in the denitrifying unit and could not be detected in the regular sand filter, This clear link between in situ abundance and denitrification suggests an active participation of paracocci and hyphomicrobia in the process, Possible selective advantages favoring the paracocci in this habitat are discussed.