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Description and characterization of bacteria attached to lotic organic aggregates (river snow) in the Elbe River of Germany and the South Saskatchewan River of Canada
Citation key Boeckelmann2001
Author Böckelmann, Uta
Year 2001
School Technische Universität Berlin, FG Umweltmikrobiologie
Abstract Aerobic and anaerobic cultivation techniques, 16S rDNA based phylogeny, and fluorescent in situ hybridization (FISH) were used to describe the phylogenetic diversity and physiological versatility of lotic microbial aggregates (river snow) obtained from the river Elbe. In the course of the year, the river snow community was characterized by a great bacterial diversity in spring with total bacterial cell counts of 2.5 x 108 cells per ml, the predominant occurrence of algae in summer (total bacterial cell counts 2.0 x 108), and the reduction of total bacterial cell counts in autumn 1.2 x 108 and winter 1.4 x 108 (all mean values). In all river snow samples, more than 70% of the bacteria, counted with the general DNA stain DAPI, also hybridized with the Bacteria specific probe EUB338. In situ analysis of the bacterial river snow community with a comprehensive suite of specific rRNA targeted probes revealed population dynamics to be governed by seasonal factors. During all seasons, beta-Proteobacteria constituted the numerically most important bacterial group forming up to 54% of the total cell counts. In contrast to this, the relative abundance of other major bacterial lineages ranged from 2% for the order Planctomycetales to 36% for Cytophaga-Flavobacteria. Batch cultures of river snow samples fed with sterile Elbe river water (0.2-µm-pore-size filtered) and supplemented with minimal amounts (0.1% w/v) of different substrates resulted in remarkable changes of the microbial community composition, with N-acetylglucosamine favouring the growth of beta-Proteobacteria. Cultivation of river snow under aerobic and anaerobic conditions with a variety of different media resulted in the isolation of 40 bacterial strains. Phenotypical and phylogenetical analysis revealed them as mostly unknown organisms affiliated to different bacterial phyla. Application of newly developed specific oligonucleotide probes proved the cultivated bacteria, including Aeromonadaceae, affiliated to the gamma-Proteobacteria, clostridia and the numerically abundant beta-Proteobacteria, as in situ relevant members of the river snow community. For the simultaneous detection of cellular components and extracellular polymeric substances (EPS) in lotic microbial aggregates (river snow) a new technique combining fluorescent in situ hybridization and lectin-binding-analysis (FISH-LBA) was developed. River snow aggregates were directly collected from the bulk water phase into coverslip chambers, in which the complete procedure including fixation, fluorescent in situ hybridization, lectin-binding and optical analysis by confocal laser scanning microscopy was performed. Neither autofluorescence originating from photosynthetic organisms nor inorganic particles did negatively interfere with the FISH-LBA technique. In Elbe river snow samples distinct compartments of the river snow structure could be visualized with FITC-labelled lectins from Triticum vulgaris, Limulus polyphemus, Arachis hypogaea, Phaseolus vulgaris and Pseudomonas aeruginosa, each binding to frequently occurring different saccharide residues in the EPS matrix. The analysis could be performed on different levels of complexity. The new combined technique visualized bacteria of different phylogenetic groups in the entire river snow structure as well as EPS components linked with various microcolonies. Slime-layers and cell-envelopes of individual eucaryotic and procaryotic cells could also be observed. Cultivation and isolation of members of the bacterial river snow community of the Canadian South Saskatchewan River on the oligotrophic medium FBM led to the discovery of the bacterial strain F8. 16S rDNA sequencing and phylogenetic analysis revealed this isolate as a deep branching gamma-Proteobacterium. Strain F8 was generally noticed by a remarkable kind of a self-produced filamentous network. The process started with rod shaped bacterial cells, accumulating a self-produced material around them, followed by the formation of filaments of different length. Filament formation developed into a network in form of a sponge like pattern. Cells moved along this network assembling firstly small and subsequently larger clusters. The process ended with large aggregates of cells, leaving an empty network of filaments behind which could be observed in cultures growing on solid and/or liquid media. Although strain F8 was able to grow in a variety of different media, filament formation occurred only in low nutrient media. Filaments could be stained with different dyes. The presence of filaments could be revealed by light microscopy, confocal laser scanning microscopy (CLSM) as well as by transmission electron microscopy (TEM).
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