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Rudolf Amann - One of the best experts on this subject based on the ideXlab platform.

  • morphological and compositional changes in a planktonic bacterial community in response to enhanced protozoan grazing
    Applied and Environmental Microbiology, 1999
    Co-Authors: Klaus Jurgens, Jakob Pernthaler, Sven Schalla, Rudolf Amann
    Abstract:

    We analyzed changes in bacterioplankton morphology and composition during enhanced protozoan grazing by image analysis and fluorescent in situ hybridization with group-specific rRNA-targeted oligonucleotide probes. Enclosure experiments were conducted in a small, fishless freshwater pond which was dominated by the cladoceran Daphnia magna. The removal of metazooplankton enhanced protozoan grazing pressure and triggered a microbial succession from fast-growing small bacteria to larger grazing-resistant morphotypes. These were mainly different types of filamentous bacteria which correlated in biomass with the population development of heterotrophic nanoflagellates (HNF). Small bacterial rods and cocci, which showed increased proportion after removal of Daphnia and doubling times of 6 to 11 h, belonged nearly exclusively to the beta subdivision of the class Proteobacteria and the Cytophaga-Flavobacterium cluster. The majority of this newly produced bacterial biomass was rapidly consumed by HNF. In contrast, the proportion of bacteria belonging to the gamma and alpha subdivisions of the Proteobacteria increased throughout the experiment. The alpha subdivision consisted mainly of rods that were 3 to 6 μm in length, which probably exceeded the size range of bacteria edible by protozoa. Initially, these organisms accounted for less than 1% of total bacteria, but after 72 h they became the predominant group of the bacterial assemblage. Other types of grazing-resistant, filamentous bacteria were also found within the beta subdivision of Proteobacteria and the Cytophaga-Flavobacterium cluster. We conclude that the predation regimen is a major structuring force for the bacterial community composition in this system. Protozoan grazing resulted in shifts of the morphological as well as the taxonomic composition of the bacterial assemblage. Grazing-resistant filamentous bacteria can develop within different phylogenetic groups of bacteria, and formerly underepresented taxa might become a dominant group when protozoan predation is the major selective pressure.

  • seasonal community and population dynamics of pelagic bacteria and archaea in a high mountain lake
    Applied and Environmental Microbiology, 1998
    Co-Authors: Jakob Pernthaler, Frank Oliver Glöckner, Albin Alfreider, Roland Psenner, Stefanie Unterholzner, Rudolf Amann
    Abstract:

    The seasonal variations in community structure and cell morphology of pelagic procaryotes from a high mountain lake (Gossenkollesee, Austria) were studied by in situ hybridization with rRNA-targeted fluorescent- ly labeled oligonucleotide probes (FISH) and image-analyzed microscopy. Compositional changes and biomass fluctuations within the assemblage were observed both in summer and beneath the winter ice cover and are discussed in the context of physicochemical and biotic parameters. Proteobacteria of the beta subclass (beta- proteobacteria) formed a dominant fraction of the bacterioplankton (annual mean, 24% of the total counts), whereas alpha-proteobacteria were of similar relative importance only during spring (mean, 11%). Bacteria of the Cytophaga-Flavobacterium cluster, although less abundant, constituted the largest fraction of the filamen- tous morphotypes during most of the year, thus contributing significantly to the total microbial biomass. Successive peaks of threadlike and rod-shaped archaea were observed during autumn thermal mixing and the period of ice cover formation, respectively. A set of oligonucleotide probes targeted to single phylotypes was constructed from 16S rRNA-encoding gene clone sequences. Three distinct populations of uncultivated mi- crobes, affiliated with the alpha- and Beta-Proteobacteria, were subsequently monitored by FISH. About one- quarter of all of the Beta-Proteobacteria (range, 6 to 53%) could be assigned to only two phylotypes. The bacterial populations studied were annually recurrent, seasonally variable, and vertically stratified, except dur- ing the periods of lake overturn. Their variability clearly exceeded the fluctuations of the total microbial as- semblage, suggesting that the apparent stability of total bacterioplankton abundances may mask highly dy- namic community fluctuations.

  • Contrasting bacterial strategies to coexist with a flagellate predator in an experimental microbial assemblage.
    Applied and Environmental Microbiology, 1997
    Co-Authors: Jakob Pernthaler, Thomas Posch, Karel Šimek, Jaroslav Vrba, Rudolf Amann, Roland Psenner
    Abstract:

    We studied predator-induced changes within a slowly growing mixed microbial assemblage that was sustained by algal exudates in a continuous cultivation system. In situ hybridization with fluorescent monolabeled oligonucleotide probes was used for a tentative community analysis. This method also allowed us to quantify the proportions of predators with ingested bacteria of different taxonomic groups. In addition, we determined grazing rates on bacteria with fluorescently labelled prey. Bacteria belonging to the alpha and beta subdivisions of the phylum Proteobacteria ((alpha)- and (beta)-Proteobacteria, respectively) showed very different responses to the addition of a bacterivorous flagellate, Bodo saltans. Within one day, filamentous protist-inedible bacteria developed; these belonged to the (beta)-Proteobacteria and constituted between 8.7 and 34% of bacteria from this subgroup. Total abundance of (beta)-Proteobacteria decreased from 3.05 x 10(sup6) to 0.23 x 10(sup6) cells ml(sup-1), and estimated cell division rates were low. Other morphologically inconspicuous protist-edible bacteria belonging to the (alpha)-Proteobacteria were found to respond to predation by an increase in growth rate. Although these bacteria were heavily grazed upon, as on average >85% of flagellate cells had ingested (alpha)-Proteobacteria, they numerically dominated after the addition of B. saltans (mean, 1.35 x 10(sup6) cells ml(sup-1)). It was thus mainly those fast-dividing strains of (alpha)-Proteobacteria that supported the growth of the flagellate population. We conclude that bacteria in mixed assemblages can adopt at least two distinct strategies as a reaction to intense flagellate predation: to outgrow predation pressure or to develop inedible, inactive filaments. Since these strategies occurred within 24 h after the addition of the flagellate, we hypothesize that chemical stimuli released by the predator may have triggered bacterial responses.

  • identification in situ and dynamics of bacteria on limnetic organic aggregates lake snow
    Applied and Environmental Microbiology, 1996
    Co-Authors: P Weiss, Rudolf Amann, Bernd Schweitzer, Meinhard Simon
    Abstract:

    Microbial assemblages on large organic aggregates (lake snow) of Lake Constance, Germany, were analyzed with rRNA-directed fluorescent oligonucleotide probes specific for the domain Bacteria and the alpha-, beta-, and gamma-subclasses of the class Proteobacteria. Lake snow aggregates were either collected in situ by SCUBA diving or in a sediment trap at 50 m or formed of natural lake water incubated in rolling cylinders under simulated in situ conditions. For the latter aggregates, the time course of the microbial colonization was also examined. The natural aggregates and those made in rolling cylinders were composed of the particulate organic material present in the lake and thus reflected the composition of the ambient plankton community. All types of lake snow aggregates examined were heavily colonized by microbial cells and harbored between 0.5 x 10(6) and > 2 x 10(6) cells aggregate -1. Between 55 and 100% of the microbial cells stained with 4', 6-diamidino-2-phenylindole (DAPI) could be visualized with the domain Bacteria-specific probe. In most samples, beta-subclass proteobacteria dominated the microbial community, constituting 27 to 42% of total cells as counted by DAPI staining, irrespective of the composition of the aggregates. During the time course experiments with the laboratory-made aggregates, the fraction of beta-subclass proteobacteria usually increased over time. Except for a few samples, alpha- and gamma-subclass proteobacteria were far less abundant than beta-subclass proteobacteria, constituting 11 to 25 and 9 to 33% of total cells, respectively. Therefore, we assume that a specific aggregate-adapted microbial community was established on the aggregates. Because the compositions of the microbial assemblages on natural and laboratory-made aggregates were similar, we conclude that aggregates made in rolling cylinders are good model system with which to examine the formation and microbial colonization of macroscopic organic aggregates.

  • Probing Activated Sludge withOligonucleotides Specific for Proteobacteria: Inadequacy ofCulture-Dependent
    1993
    Co-Authors: Michael Wagner, H Lemmer, Rudolf Amann, Karl-heinz Schleifer
    Abstract:

    Bacterial community structures inactivated sludge samples fromaeration tanksofatwo-stage system with ahigh-load first stage andalow-load second stage were analyzed witholigonucleotide probes. Theprobes were complementary toconserved regions oftherRNAofthealpha, beta, andgamma subclasses ofproteobacteria andofallbacteria. Group-specific cell counts were determined byinsituhybridization withfluorescent probe derivatives. Contributions oftheproteobacterial subclasses tototal bacterial rRNAwerequantified bydotblot hybridization withdigoxigenin-labeled oligonucleotides. Theactivated sludge samples were dominatedby proteobacteria fromthealpha, beta, or gamma subclass. Theseproteobacteria account forabout80% ofall active bacteria foundintheactivated sludge. Forbothsamples thecommunity structures determined with molecular techniques were compared withthecomposition oftheheterotrophic saprophyte flora isolated on nutrient-rich medium. Probes wereusedtorapidly classify theisolates andtodirectly monitor population shifts innutrient-amended, activated sludge samples. Therichmediumfavoredgrowthofgamma-subclass proteobacteria (e.g., enterobacteria) andselected against beta-subclass proteobacteria. Theculture-dependent community structure analysis ofactivated sludge produced partial andheavily biased results. A more realistic viewwill beobtained byusing insitu techniques. Thetreatment ofwastewater byactivated sludge systems is,intermsofmetabolized matter,probably today's most important biotechnological process.A lotofeffort hasbeen funneled intoprocess engineering, whereasour current knowledge ofmicrobial community structure-function correlations andconsequently a microbiological understanding

Michael Wagner - One of the best experts on this subject based on the ideXlab platform.

  • Obligate bacterial endosymbionts of Acanthamoeba spp. related to the Beta-Proteobacteria: proposal of 'Candidatus Procabacter acanthamoebae' gen. nov., sp. nov.
    International Journal of Systematic and Evolutionary Microbiology, 2002
    Co-Authors: Matthias Horn, Thomas R. Fritsche, Tanja Linner, Romesh K. Gautom, Marit D. Harzenetter, Michael Wagner
    Abstract:

    All obligate bacterial endosymbionts of free-living amoebae currently described are affiliated with the alpha-Proteobacteria, the Chlamydiales or the phylum Cytophaga-Flavobacterium-Bacteroides. Here, six rod-shaped gram-negative obligate bacterial endosymbionts of clinical and environmental isolates of Acanthamoeba spp. from the USA and Malaysia are reported. Comparative 16S rDNA sequence analysis demonstrated that these endosymbionts form a novel, monophyletic lineage within the Beta-Proteobacteria, showing less than 90% sequence similarity to all other recognized members of this subclass. 23S rDNA sequence analysis of two symbionts confirmed this affiliation and revealed the presence of uncommon putative intervening sequences of 146 bp within helix-25 that shared no sequence homology to any other bacterial rDNA. In addition, the 23S rRNA of these endosymbionts displayed one polymorphism at the target site of oligonucleotide probe BET42a that is conserved in all other sequenced Beta-Proteobacteria. Intra-cytoplasmatic localization of the endosymbionts within the amoebal host cells was confirmed by electron microscopy and fluorescence in situ hybridization with a specific 16S rRNA-targeted oligonucleotide probe. Based on these findings, the provisional name 'Candidatus Procabacter acanthamoebae' is proposed for classification of a representative of the six endosymbionts of Acanthamoeba spp. studied in this report. Comparative 18S rDNA sequence analysis of the Acanthamoeba host cells revealed their membership with either Acanthamoeba 18S rDNA sequence type T5 (Acanthamoeba lenticulata) or sequence type T4, which comprises the majority of all Acanthamoeba isolates.

  • characterization of bacterial communities from activated sludge culture dependent numerical identification versus in situ identification using group and genus specific rrna targeted oligonucleotide probes
    Microbial Ecology, 1996
    Co-Authors: R. Erhart, Judith Böhringer, Claudia Beimfohr, Peter Kampfer, Michael Wagner, Rudolf I. Amann
    Abstract:

    The structures of bacterial communities were studied in activated sludge samples obtained from the aerobic and anaerobic zones of a wastewater treatment plant showing enhanced phosphorous removal. Samples were analyzed by in situ hybridization with oligonucleotide probes complementary to selected regions of the 16S and 23S ribosomal RNA (rRNA) characteristic for defined phylogenetic entities (genera and larger groups). The microbial community structures revealed by molecular techniques were compared with the compositions of culturable bacterial communities, obtained from the characterization of 255 isolates from tryptone-soy (TS) agar and R2A agar. These isolates were characterized by 89 physiological tests and their cellular fatty acid patterns, and identified. Culture-dependent techniques indicated the following distribution: different Aeromonas spp. (2.7–8.3% on R2A agar; 45.0–63.7% on TS agar), Acinetobacter spp. (5.4–9.0% on R2A agar; 5.0–9.1% on TS agar), Pseudomonas spp. (up to 10% on R2A agar) and Shewanella putrefaciens (up to 3.0% on R2A agar), all members of the gamma subclass of Proteobacteria, were isolated most frequently. The relatively rare isolates of the beta subclass were identified as Acidovorax spp., Alcaligenes spp., and Comamonas spp., The Gram-positive bacteria (high DNA G+C) were assigned mainly to Arthrobacter spp., Microbacterium spp., and Mycobacterium phlei. In order to assess the in situ abundance of the most frequently isolated genus, Aeromonas, two rRNA-targeted oligonucleotide probes were developed. The two gamma proteobacterial genera Aeromonas and Acinetobacter constituted less than 5% of all bacteria. In situ, Proteobacteria belonging to the beta subclass and high G+C Gram-positive bacteria were dominant. From filamentous bacteria, Sphaerotilus spp. and Leptothrix spp. could be detected occasionally. In addition, one sample contained a high proportion of the morphologically distinct filaments of Microthrix parvicella.

  • Probing Activated Sludge withOligonucleotides Specific for Proteobacteria: Inadequacy ofCulture-Dependent
    1993
    Co-Authors: Michael Wagner, H Lemmer, Rudolf Amann, Karl-heinz Schleifer
    Abstract:

    Bacterial community structures inactivated sludge samples fromaeration tanksofatwo-stage system with ahigh-load first stage andalow-load second stage were analyzed witholigonucleotide probes. Theprobes were complementary toconserved regions oftherRNAofthealpha, beta, andgamma subclasses ofproteobacteria andofallbacteria. Group-specific cell counts were determined byinsituhybridization withfluorescent probe derivatives. Contributions oftheproteobacterial subclasses tototal bacterial rRNAwerequantified bydotblot hybridization withdigoxigenin-labeled oligonucleotides. Theactivated sludge samples were dominatedby proteobacteria fromthealpha, beta, or gamma subclass. Theseproteobacteria account forabout80% ofall active bacteria foundintheactivated sludge. Forbothsamples thecommunity structures determined with molecular techniques were compared withthecomposition oftheheterotrophic saprophyte flora isolated on nutrient-rich medium. Probes wereusedtorapidly classify theisolates andtodirectly monitor population shifts innutrient-amended, activated sludge samples. Therichmediumfavoredgrowthofgamma-subclass proteobacteria (e.g., enterobacteria) andselected against beta-subclass proteobacteria. Theculture-dependent community structure analysis ofactivated sludge produced partial andheavily biased results. A more realistic viewwill beobtained byusing insitu techniques. Thetreatment ofwastewater byactivated sludge systems is,intermsofmetabolized matter,probably today's most important biotechnological process.A lotofeffort hasbeen funneled intoprocess engineering, whereasour current knowledge ofmicrobial community structure-function correlations andconsequently a microbiological understanding

  • probing activated sludge with oligonucleotides specific for proteobacteria inadequacy of culture dependent methods for describing microbial community structure
    Applied and Environmental Microbiology, 1993
    Co-Authors: Michael Wagner, Rudolf I. Amann, H Lemmer, Karl-heinz Schleifer
    Abstract:

    Bacterial community structures in activated sludge samples from aeration tanks of a two-stage system with a high-load first stage and a low-load second stage were analyzed with oligonucleotide probes. The probes were complementary to conserved regions of the rRNA of the alpha, beta, and gamma subclasses of proteobacteria and of all bacteria. Group-specific cell counts were determined by in situ hybridization with fluorescent probe derivatives. Contributions of the proteobacterial subclasses to total bacterial rRNA were quantified by dot blot hybridization with digoxigenin-labeled oligonucleotides. The activated sludge samples were dominated by proteobacteria from the alpha, beta, or gamma subclass. These proteobacteria account for about 80% of all active bacteria found in the activated sludge. For both samples the community structures determined with molecular techniques were compared with the composition of the heterotrophic saprophyte flora isolated on nutrient-rich medium. Probes were used to rapidly classify the isolates and to directly monitor population shifts in nutrient-amended, activated sludge samples. The rich medium favored growth of gamma-subclass proteobacteria (e.g., enterobacteria) and selected against beta-subclass proteobacteria. The culture-dependent community structure analysis of activated sludge produced partial and heavily biased results. A more realistic view will be obtained by using in situ techniques. Images

  • phylogenetic oligodeoxynucleotide probes for the major subclasses of proteobacteria problems and solutions
    Systematic and Applied Microbiology, 1992
    Co-Authors: Werner Manz, Rudolf Amann, Michael Wagner, Wolfgang Ludwig, Karl-heinz Schleifer
    Abstract:

    Summary Based on comparative analyses of 16S and 23S ribosomal RNA sequences we have located sites specific for the alpha-, beta-, and gamma-subclasses of Proteobacteria. Short oligodeoxynucleotides complementary to these signature regions were evaluated as potential nucleic acid probes for the differentiation of the major subclasses of Proteobacteria. Hybridization conditions were optimized by the addition of form-amide to the hybridization buffer and high stringency post-hybridization washing. Single-mismatch discrimination of probes was further improved by blocking nontarget probe binding sites with competitor oligonucleotides. Nonisotopic dot-blot hybridization to reference strains demonstrated the expected probe specificities, whole cell hybridization with fluorescent probe derivatives allowed the classification of individual microbial cells. The probes will be useful for determinative studies and for the in situ monitoring of population distribution and dynamics in microbial communities.

Rudolf I. Amann - One of the best experts on this subject based on the ideXlab platform.

  • Predominance of β‐proteobacteria in summer melt pools on Arctic pack ice
    Limnology and Oceanography, 2004
    Co-Authors: Robin Brinkmeyer, Elisabeth Helmke, Frank Oliver Glöckner, Rudolf I. Amann
    Abstract:

    The diversity and community structure of bacteria in melt pools on Arctic pack ice floes were dominated by bproteobacteria. Thirty-five percent of the pure cultures isolated in 1997 from pack ice floes north of Svalbard and in the Fram Strait were from the b-proteobacteria group. Within this group, there were only two phylotypes clustering within the widespread Beta I cluster, also known as the Comamonadaceae clade. One phylotype, most closely related to Aquaspirillum arcticum(96.0‐97.3% identical), was frequent among cultures isolated from 10 melt pools. A 16S ribosomal RNA (rRNA) gene clone library, constructed from a melt pool that was sampled 2 yr later in the Fram Strait, was also dominated by b-proteobacteria, in particular the same recurrent isolate phylotype designated ‘‘MP-BetaI’’. Fluorescence in situ hybridization of 20 melt pools corroborated the cultivation and cloning data. bProteobacteria were the most abundant bacterial group, constituting ;49% of the bacteria that were stained by 496diamidino-2-phenylindole (DAPI). a- and g-proteobacteria accounted for only 2% each, the Cytophaga‐Flavobacterium group accounted for 9%, and the Actinobacteria spp. accounted for 9%. Approximately 63% of the b-proteobacterial fraction that was found in the melt pools was determined with a newly developed probe to be the recurrent b-proteobacterial MP-BetaI phylotypes, indicating that it is particularly adapted for success in this extreme environment.

  • Characterization of bacterial communities from activated sludge: Culture-dependent numerical identification versus in situ identification using group- and genus-specific rRNA-targeted oligonucleotide probes
    Microbial Ecology, 1996
    Co-Authors: Peter Kampfer, R. Erhart, Judith Böhringer, Claudia Beimfohr, M. Wagner, Rudolf I. Amann
    Abstract:

    The structures of bacterial communities were studied in activated sludge samples obtained from the aerobic and anaerobic zones of a wastewater treatment plant showing enhanced phosphorous removal. Samples were analyzed by in situ hybridization with oligonucleotide probes complementary to selected regions of the 16S and 23S ribosomal RNA (rRNA) characteristic for defined phylogenetic entities (genera and larger groups). The microbial community structures revealed by molecular techniques were compared with the compositions of culturable bacterial communities, obtained from the characterization of 255 isolates from tryptone-soy (TS) agar and R2A agar. These isolates were characterized by 89 physiological tests and their cellular fatty acid patterns, and identified. Culture-dependent techniques indicated the following distribution: different Aeromonas spp. (2.7–8.3% on R2A agar; 45.0–63.7% on TS agar), Acinetobacter spp. (5.4–9.0% on R2A agar; 5.0–9.1% on TS agar), Pseudomonas spp. (up to 10% on R2A agar) and Shewanella putrefaciens (up to 3.0% on R2A agar), all members of the gamma subclass of Proteobacteria, were isolated most frequently. The relatively rare isolates of the beta subclass were identified as Acidovorax spp., Alcaligenes spp., and Comamonas spp., The Gram-positive bacteria (high DNA G+C) were assigned mainly to Arthrobacter spp., Microbacterium spp., and Mycobacterium phlei . In order to assess the in situ abundance of the most frequently isolated genus, Aeromonas , two rRNA-targeted oligonucleotide probes were developed. The two gamma proteobacterial genera Aeromonas and Acinetobacter constituted less than 5% of all bacteria. In situ, Proteobacteria belonging to the beta subclass and high G+C Gram-positive bacteria were dominant. From filamentous bacteria, Sphaerotilus spp. and Leptothrix spp. could be detected occasionally. In addition, one sample contained a high proportion of the morphologically distinct filaments of Microthrix parvicella . As for the genus Acinetobacter , the relative abundance of the most frequently gamma-proteobacterial genus Aeromonas was overestimated by the intrinsic selectivity of cultivation. Cultivation on nutrient-rich medium (TS-agar) especially supported an enhanced isolation of bacteria belonging to these two genera.

  • characterization of bacterial communities from activated sludge culture dependent numerical identification versus in situ identification using group and genus specific rrna targeted oligonucleotide probes
    Microbial Ecology, 1996
    Co-Authors: R. Erhart, Judith Böhringer, Claudia Beimfohr, Peter Kampfer, Michael Wagner, Rudolf I. Amann
    Abstract:

    The structures of bacterial communities were studied in activated sludge samples obtained from the aerobic and anaerobic zones of a wastewater treatment plant showing enhanced phosphorous removal. Samples were analyzed by in situ hybridization with oligonucleotide probes complementary to selected regions of the 16S and 23S ribosomal RNA (rRNA) characteristic for defined phylogenetic entities (genera and larger groups). The microbial community structures revealed by molecular techniques were compared with the compositions of culturable bacterial communities, obtained from the characterization of 255 isolates from tryptone-soy (TS) agar and R2A agar. These isolates were characterized by 89 physiological tests and their cellular fatty acid patterns, and identified. Culture-dependent techniques indicated the following distribution: different Aeromonas spp. (2.7–8.3% on R2A agar; 45.0–63.7% on TS agar), Acinetobacter spp. (5.4–9.0% on R2A agar; 5.0–9.1% on TS agar), Pseudomonas spp. (up to 10% on R2A agar) and Shewanella putrefaciens (up to 3.0% on R2A agar), all members of the gamma subclass of Proteobacteria, were isolated most frequently. The relatively rare isolates of the beta subclass were identified as Acidovorax spp., Alcaligenes spp., and Comamonas spp., The Gram-positive bacteria (high DNA G+C) were assigned mainly to Arthrobacter spp., Microbacterium spp., and Mycobacterium phlei. In order to assess the in situ abundance of the most frequently isolated genus, Aeromonas, two rRNA-targeted oligonucleotide probes were developed. The two gamma proteobacterial genera Aeromonas and Acinetobacter constituted less than 5% of all bacteria. In situ, Proteobacteria belonging to the beta subclass and high G+C Gram-positive bacteria were dominant. From filamentous bacteria, Sphaerotilus spp. and Leptothrix spp. could be detected occasionally. In addition, one sample contained a high proportion of the morphologically distinct filaments of Microthrix parvicella.

  • probing activated sludge with oligonucleotides specific for proteobacteria inadequacy of culture dependent methods for describing microbial community structure
    Applied and Environmental Microbiology, 1993
    Co-Authors: Michael Wagner, Rudolf I. Amann, H Lemmer, Karl-heinz Schleifer
    Abstract:

    Bacterial community structures in activated sludge samples from aeration tanks of a two-stage system with a high-load first stage and a low-load second stage were analyzed with oligonucleotide probes. The probes were complementary to conserved regions of the rRNA of the alpha, beta, and gamma subclasses of proteobacteria and of all bacteria. Group-specific cell counts were determined by in situ hybridization with fluorescent probe derivatives. Contributions of the proteobacterial subclasses to total bacterial rRNA were quantified by dot blot hybridization with digoxigenin-labeled oligonucleotides. The activated sludge samples were dominated by proteobacteria from the alpha, beta, or gamma subclass. These proteobacteria account for about 80% of all active bacteria found in the activated sludge. For both samples the community structures determined with molecular techniques were compared with the composition of the heterotrophic saprophyte flora isolated on nutrient-rich medium. Probes were used to rapidly classify the isolates and to directly monitor population shifts in nutrient-amended, activated sludge samples. The rich medium favored growth of gamma-subclass proteobacteria (e.g., enterobacteria) and selected against beta-subclass proteobacteria. The culture-dependent community structure analysis of activated sludge produced partial and heavily biased results. A more realistic view will be obtained by using in situ techniques. Images

Peter Kampfer - One of the best experts on this subject based on the ideXlab platform.

  • Characterization of bacterial communities from activated sludge: Culture-dependent numerical identification versus in situ identification using group- and genus-specific rRNA-targeted oligonucleotide probes
    Microbial Ecology, 1996
    Co-Authors: Peter Kampfer, R. Erhart, Judith Böhringer, Claudia Beimfohr, M. Wagner, Rudolf I. Amann
    Abstract:

    The structures of bacterial communities were studied in activated sludge samples obtained from the aerobic and anaerobic zones of a wastewater treatment plant showing enhanced phosphorous removal. Samples were analyzed by in situ hybridization with oligonucleotide probes complementary to selected regions of the 16S and 23S ribosomal RNA (rRNA) characteristic for defined phylogenetic entities (genera and larger groups). The microbial community structures revealed by molecular techniques were compared with the compositions of culturable bacterial communities, obtained from the characterization of 255 isolates from tryptone-soy (TS) agar and R2A agar. These isolates were characterized by 89 physiological tests and their cellular fatty acid patterns, and identified. Culture-dependent techniques indicated the following distribution: different Aeromonas spp. (2.7–8.3% on R2A agar; 45.0–63.7% on TS agar), Acinetobacter spp. (5.4–9.0% on R2A agar; 5.0–9.1% on TS agar), Pseudomonas spp. (up to 10% on R2A agar) and Shewanella putrefaciens (up to 3.0% on R2A agar), all members of the gamma subclass of Proteobacteria, were isolated most frequently. The relatively rare isolates of the beta subclass were identified as Acidovorax spp., Alcaligenes spp., and Comamonas spp., The Gram-positive bacteria (high DNA G+C) were assigned mainly to Arthrobacter spp., Microbacterium spp., and Mycobacterium phlei . In order to assess the in situ abundance of the most frequently isolated genus, Aeromonas , two rRNA-targeted oligonucleotide probes were developed. The two gamma proteobacterial genera Aeromonas and Acinetobacter constituted less than 5% of all bacteria. In situ, Proteobacteria belonging to the beta subclass and high G+C Gram-positive bacteria were dominant. From filamentous bacteria, Sphaerotilus spp. and Leptothrix spp. could be detected occasionally. In addition, one sample contained a high proportion of the morphologically distinct filaments of Microthrix parvicella . As for the genus Acinetobacter , the relative abundance of the most frequently gamma-proteobacterial genus Aeromonas was overestimated by the intrinsic selectivity of cultivation. Cultivation on nutrient-rich medium (TS-agar) especially supported an enhanced isolation of bacteria belonging to these two genera.

  • characterization of bacterial communities from activated sludge culture dependent numerical identification versus in situ identification using group and genus specific rrna targeted oligonucleotide probes
    Microbial Ecology, 1996
    Co-Authors: R. Erhart, Judith Böhringer, Claudia Beimfohr, Peter Kampfer, Michael Wagner, Rudolf I. Amann
    Abstract:

    The structures of bacterial communities were studied in activated sludge samples obtained from the aerobic and anaerobic zones of a wastewater treatment plant showing enhanced phosphorous removal. Samples were analyzed by in situ hybridization with oligonucleotide probes complementary to selected regions of the 16S and 23S ribosomal RNA (rRNA) characteristic for defined phylogenetic entities (genera and larger groups). The microbial community structures revealed by molecular techniques were compared with the compositions of culturable bacterial communities, obtained from the characterization of 255 isolates from tryptone-soy (TS) agar and R2A agar. These isolates were characterized by 89 physiological tests and their cellular fatty acid patterns, and identified. Culture-dependent techniques indicated the following distribution: different Aeromonas spp. (2.7–8.3% on R2A agar; 45.0–63.7% on TS agar), Acinetobacter spp. (5.4–9.0% on R2A agar; 5.0–9.1% on TS agar), Pseudomonas spp. (up to 10% on R2A agar) and Shewanella putrefaciens (up to 3.0% on R2A agar), all members of the gamma subclass of Proteobacteria, were isolated most frequently. The relatively rare isolates of the beta subclass were identified as Acidovorax spp., Alcaligenes spp., and Comamonas spp., The Gram-positive bacteria (high DNA G+C) were assigned mainly to Arthrobacter spp., Microbacterium spp., and Mycobacterium phlei. In order to assess the in situ abundance of the most frequently isolated genus, Aeromonas, two rRNA-targeted oligonucleotide probes were developed. The two gamma proteobacterial genera Aeromonas and Acinetobacter constituted less than 5% of all bacteria. In situ, Proteobacteria belonging to the beta subclass and high G+C Gram-positive bacteria were dominant. From filamentous bacteria, Sphaerotilus spp. and Leptothrix spp. could be detected occasionally. In addition, one sample contained a high proportion of the morphologically distinct filaments of Microthrix parvicella.

  • taxonomic position of aromatic degrading denitrifying pseudomonad strains k 172 and kb 740 and their description as new members of the genera thauera as thauera aromatica sp nov and azoarcus as azoarcus evansii sp nov respectively members of the beta
    International Journal of Systematic and Evolutionary Microbiology, 1995
    Co-Authors: Hansjoachim Anders, Peter Kampfer, Wolfgang Ludwig, Anette Kaetzke, Georg Fuchs
    Abstract:

    In the past workers have isolated several pseudomonad strains which have been used for studies of anaerobic aromatic metabolism. The best studied of these strains are strains KB 740T(T = type strain) and K172T. The taxonomic positions of these two organisms were determined by classical methods, including experiments to determine substrate spectrum, quinone type, and total fatty acid composition. Our results clearly excluded these strains from the authentic genus Pseudomonas, which belongs to the gamma subclass of the Proteobacteria. Instead, the properties of these organisms indicated that they belong to the beta subclass of the Proteobacteria. The sequences of the 16S ribosomal DNA genes confirmed this conclusion and indicated that strain K 172Trepresents a new species of the genus Thauera, Thauera aromatica, and that strain KB 740Trepresents a new species of the genus Azoarcus, Azoarcus evansii.

Karl-heinz Schleifer - One of the best experts on this subject based on the ideXlab platform.

  • Probing Activated Sludge withOligonucleotides Specific for Proteobacteria: Inadequacy ofCulture-Dependent
    1993
    Co-Authors: Michael Wagner, H Lemmer, Rudolf Amann, Karl-heinz Schleifer
    Abstract:

    Bacterial community structures inactivated sludge samples fromaeration tanksofatwo-stage system with ahigh-load first stage andalow-load second stage were analyzed witholigonucleotide probes. Theprobes were complementary toconserved regions oftherRNAofthealpha, beta, andgamma subclasses ofproteobacteria andofallbacteria. Group-specific cell counts were determined byinsituhybridization withfluorescent probe derivatives. Contributions oftheproteobacterial subclasses tototal bacterial rRNAwerequantified bydotblot hybridization withdigoxigenin-labeled oligonucleotides. Theactivated sludge samples were dominatedby proteobacteria fromthealpha, beta, or gamma subclass. Theseproteobacteria account forabout80% ofall active bacteria foundintheactivated sludge. Forbothsamples thecommunity structures determined with molecular techniques were compared withthecomposition oftheheterotrophic saprophyte flora isolated on nutrient-rich medium. Probes wereusedtorapidly classify theisolates andtodirectly monitor population shifts innutrient-amended, activated sludge samples. Therichmediumfavoredgrowthofgamma-subclass proteobacteria (e.g., enterobacteria) andselected against beta-subclass proteobacteria. Theculture-dependent community structure analysis ofactivated sludge produced partial andheavily biased results. A more realistic viewwill beobtained byusing insitu techniques. Thetreatment ofwastewater byactivated sludge systems is,intermsofmetabolized matter,probably today's most important biotechnological process.A lotofeffort hasbeen funneled intoprocess engineering, whereasour current knowledge ofmicrobial community structure-function correlations andconsequently a microbiological understanding

  • probing activated sludge with oligonucleotides specific for proteobacteria inadequacy of culture dependent methods for describing microbial community structure
    Applied and Environmental Microbiology, 1993
    Co-Authors: Michael Wagner, Rudolf I. Amann, H Lemmer, Karl-heinz Schleifer
    Abstract:

    Bacterial community structures in activated sludge samples from aeration tanks of a two-stage system with a high-load first stage and a low-load second stage were analyzed with oligonucleotide probes. The probes were complementary to conserved regions of the rRNA of the alpha, beta, and gamma subclasses of proteobacteria and of all bacteria. Group-specific cell counts were determined by in situ hybridization with fluorescent probe derivatives. Contributions of the proteobacterial subclasses to total bacterial rRNA were quantified by dot blot hybridization with digoxigenin-labeled oligonucleotides. The activated sludge samples were dominated by proteobacteria from the alpha, beta, or gamma subclass. These proteobacteria account for about 80% of all active bacteria found in the activated sludge. For both samples the community structures determined with molecular techniques were compared with the composition of the heterotrophic saprophyte flora isolated on nutrient-rich medium. Probes were used to rapidly classify the isolates and to directly monitor population shifts in nutrient-amended, activated sludge samples. The rich medium favored growth of gamma-subclass proteobacteria (e.g., enterobacteria) and selected against beta-subclass proteobacteria. The culture-dependent community structure analysis of activated sludge produced partial and heavily biased results. A more realistic view will be obtained by using in situ techniques. Images

  • phylogenetic oligodeoxynucleotide probes for the major subclasses of proteobacteria problems and solutions
    Systematic and Applied Microbiology, 1992
    Co-Authors: Werner Manz, Rudolf Amann, Michael Wagner, Wolfgang Ludwig, Karl-heinz Schleifer
    Abstract:

    Summary Based on comparative analyses of 16S and 23S ribosomal RNA sequences we have located sites specific for the alpha-, beta-, and gamma-subclasses of Proteobacteria. Short oligodeoxynucleotides complementary to these signature regions were evaluated as potential nucleic acid probes for the differentiation of the major subclasses of Proteobacteria. Hybridization conditions were optimized by the addition of form-amide to the hybridization buffer and high stringency post-hybridization washing. Single-mismatch discrimination of probes was further improved by blocking nontarget probe binding sites with competitor oligonucleotides. Nonisotopic dot-blot hybridization to reference strains demonstrated the expected probe specificities, whole cell hybridization with fluorescent probe derivatives allowed the classification of individual microbial cells. The probes will be useful for determinative studies and for the in situ monitoring of population distribution and dynamics in microbial communities.