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Polyphasic characterization of the microbial population of drinking water biofilms
Zitatschlüssel Kalmbach1998
Autor Kalmbach, Sibylle
Jahr 1998
Schule Technische Universität Berlin, FG Umweltmikrobiologie
Zusammenfassung The present study aimed at the comprehensive characterization of the microbial biofilm population in drinking water distribution systems. At first, the dynamics of biofilm formation were investigated with regard to cell density, metabolic potential and phylogenetic affiliation of the microbial population. Polyethylene and glass surfaces were rapidly colonized by mostly rod-shaped bacteria, and cell densities were controlled by eukaryotic and possibly by prokaryotic predators. In situ hybridizations with group-specific rRNA-targeted oligonucleotide probes revealed the predominance of bacteria affiliated to the beta-subclass of Proteobacteria within the biofilm community. Although the metabolic potential of the cells, determined by CTC-reduction and the presence of hybridization signals, decreased during biofilm formation, most cells retained the ability for ribosome synthesis and elongation measured by the newly developed probe active count (PAC) assay. In general, planktonic cells were found to be smaller and less active than their sessile counterparts. The information gained in the first part of this study was applied for the successful isolation of the prevalent bacteria of the young biofilm population. Phylogenetic analysis based on 16S rRNA sequences revealed all of the eight isolates B1 to B8 as hitherto unknown beta-Proteobacteria. Strains B4, B6 and B8 formed a cluster of closely related bacteria within the Rubrivivax-Leptothrix-branch of this taxon. Subsequently, specific oligonucleotide probes, termed beta1 to beta8b, were developed in order to determine the occurrence and abundance of strains B1 to B8 in their natural habitat. In situ probing of house installation system biofilms revealed that (i) 67 to 72% of total cell counts, corresponding to more than 80% of all beta-Proteobacteria, could be encompassed with this suite of probes, (ii) the phylogenetically closely related strains B4, B6 and B8 constituted the in situ dominant population and (iii) the dominating bacterial species were culturable on R2A medium. More than 65% of the microbial population could be shown to be in a viable but nonculturable (VBNC) state by using PAC assays. The comparison of isolation frequencies with the in situ abundance indicated variable ratios of culturable to VBNC cells among the investigated strains. In situ probing of biofilms from groundwater and a main pipe, as well as additional house installation system biofilms, revealed strain B8 as by far the most abundant species. Strain B8 could also be detected in high numbers in biofilms from drinking water systems of Stockholm, Mainz and Hamburg, demonstrating that this is a ubiquitous, in situ dominant drinking-water-biofilm organism. Substratum-induced population shifts could be shown for soft-PVC, which favored growth of strain B4 or other yet to be identified beta-Proteobacteria. Applying the newly developed probes in other freshwater ecosystems, including activated sludge, river and lake water, cells yielding clear hybridization signals could be detected with all probes except beta7. Phenotypic characterization of strains B4, B6 and B8 showed them to be motile, microaerophilic, gram-negative rods, which were oxidase positive and catalase negative and contained polyalkanoate and polyphosphate as storage polymers. The metabolized substrates included a broad range of organic acids, but no carbohydrates at all. Based on phenotypic and genotypic characterization, the new genus Aquabacterium is proposed, including the species Aquabacterium citratiphilum sp. nov. (strain B4), Aquabacterium parvum sp. nov. (strain B6) and Aquabacterium commune sp. nov. (strain B8).
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