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ABSTRACT: The substrate partitioning of sympatric populations of freshwater bacterioplankton was studied via microautoradiography and fluorescence in situ hybridization. Fourteen radiolabeled tracers were used to assess microbial acquisition spectra of low-molecular-weight (LMW) organic compounds. The most abundant group, ac1 Actinobacteria, were highly active in leucine, thymidine and glucose assimilation, whereas Alphaproteobacteria from the LD12 lineage (the freshwater sister clade of SAR11) only weakly incorporated these tracers, but exhibited a distinct preference for glutamine and glutamate. Different Bacteroidetes showed contrasting uptake patterns: Flavobacteriales did not incorporate significant amounts of any LMW compound, and Cyclobacteriaceae were clearly specialized on leucine, glucose and arginine. Betaproteobacteria represented the most active and versatile bacterioplankton fraction and >90% of them could be assigned to eight species- to genus-like populations with contrasting substrate specialization. Limnohabitans sp. were the most abundant and active Betaproteobacteria, incorporating almost all tracers. While three closely related betaproteobacterial populations substantially differed in their uptake spectra, two more distantly related lineages had very similar preferences, and one population did not incorporate any tracer. The dominant phototrophic microorganism, the filamentous cyanobacterium Planktothrix rubescens, assimilated several substrates, whereas other (pico)cyanobacteria had no heterotrophic activity. The variable extent of specialization by the studied bacterial taxa on subsets of LMW compounds contrasts theoretical considerations about non-selective microbial substrate assimilation at oligotrophic conditions. This physiological niche separation might be one explanation for the coexistence of freshwater bacterioplankton species in a seemingly uniform environment.
The ISME Journal 05/2013; 7:896-907. · 7.38 Impact Factor
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ABSTRACT: The impact of different feed waters in terms of eukaryotic populations and organic carbon content on the biofilm structure formation and permeate flux during Gravity-Driven Membrane (GDM) filtration was investigated in this study. GDM filtration was performed at ultra-low pressure (65 mbar) in dead-end mode without control of the biofilm formation. Different feed waters were tested (River water, pre-treated river water, lake water, and tap water) and varied with regard to their organic substrate content and their predator community. River water was manipulated either by chemically inhibiting all eukaryotes or by filtering out macrozoobenthos (metazoan organisms). The structure of the biofilm was characterized at the meso- and micro-scale using Optical Coherence Tomography (OCT) and Confocal Laser Scanning Microscopy (CLSM), respectively. Based on Total Organic Carbon (TOC) measurements, the river waters provided the highest potential for bacterial growth whereas tap water had the lowest. An increasing content in soluble and particulate organic substrate resulted in increasing biofilm accumulation on membrane surface. However, enhanced biofilm accumulation did not result in lower flux values and permeate flux was mainly influenced by the structure of the biofilm. Metazoan organisms (in particular nematodes and oligochaetes) built-up protective habitats, which resulted in the formation of open and spatially heterogeneous biofilms composed of biomass patches. In the absence of predation by metazoan organisms, a flat and compact biofilm developed. It is concluded that the activity of metazoan organisms in natural river water and its impact on biofilm structure balances the detrimental effect of a high biofilm accumulation, thus allowing for a broader application of GDM filtration. Finally, our results suggest that for surface waters with high particulate organic carbon (POC) content, the use of worms is suitable to enhance POC removal before ultrafiltration units.
Water Research 04/2013; 47(6):2085-95. · 4.86 Impact Factor
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ABSTRACT: The vernal successions of phytoplankton, heterotrophic nanoflagellates (HNF) and viruses in temperate lakes result in alternating dominance of top-down and bottom-up factors on the bacterial community. This may lead to asynchronous blooms of bacteria with different life strategies and affect the channelling of particular components of the dissolved organic matter (DOM) through microbial food webs. We followed the dynamics of several bacterial populations and of other components of the microbial food web throughout the spring phytoplankton bloom period in a pre-alpine lake, and we assessed bacterial uptake patterns of two constituents of the labile DOM pool (N-acetyl-glucosamine [NAG] and leucine). There was a clear genotypic shift within the bacterial assemblage, from fast growing Cytophaga-Flavobacteria (CF) affiliated with Fluviicola and from Betaproteobacteria (BET) of the Limnohabitans cluster to more grazing resistant AcI Actinobacteria (ACT) and to filamentous morphotypes. This was paralleled by successive blooms of viruses and HNF. We also noted the transient rise of other CF (related to Cyclobacteriaceae and Sphingobacteriaceae) that are not detected by fluorescence in situ hybridization with the general CF probe. Both, the average uptake rates of leucine and the fractions of leucine incorporating bacteria were approximately five to sixfold higher than of NAG. However, the composition of the NAG-active community was much more prone to genotypic successions, in particular of bacteria with different life strategies: While 'opportunistically' growing BET and CF dominated NAG uptake in the initial period ruled by bottom-up factors, ACT constituted the major fraction of NAG active cells during the subsequent phase of high predation pressure. This indicates that some ACT could profit from a substrate that might in parts have originated from the grazing of protists on their bacterial competitors.
Environmental Microbiology 11/2011; 14(3):794-806. · 5.84 Impact Factor
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ABSTRACT: We examined the effect of light on the heterotrophic activity of the filamentous cyanobacterium Planktothrix rubescens and on its relationship with the accompanying bacteria. In situ leucine uptake by bacteria and cyanobacteria was determined in a subalpine mesotrophic lake, and natural assemblages from the zone of maximal P. rubescens abundances were incubated for 2 days at contrasting light regimes (ambient, 100× increased, dark). Planktothrix rubescens from the photic zone of the lake incorporated substantially more leucine, but some heterotrophic activity was maintained in filaments from the hypolimnion. Exposure of cyanobacteria to increased irradiance or darkness resulted in significantly lower leucine incorporation than at ambient light conditions. Highest abundances and leucine uptake of Betaproteobacteria from the genus Limnohabitans were found in the accompanying microflora at suboptimal irradiance levels for P. rubescens or in dark incubations. Therefore, two Limnohabitans strains (representing different species) were co-cultured with axenic P. rubescens at different light conditions. The abundances and leucine incorporation rates of both strains most strongly increased at elevated irradiance levels, in parallel to a decrease of photosynthetic pigment fluorescence and the fragmentation of cyanobacterial filaments. Our results suggest that Limnohabitans spp. in lakes might profit from the presence of physiologically stressed P. rubescens.
Environmental Microbiology 11/2011; 14(3):765-78. · 5.84 Impact Factor
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ABSTRACT: The pelagic zone of large, deep freshwater lakes features pronounced horizontal and vertical gradients of physicochemical parameters, which in turn might allow for a nonuniform occurrence of specifically adapted bacterial taxa. We, therefore, studied the spatial distribution patterns of different heterotrophic bacteria, picocyanobacteria, and the dominant primary producer, the filamentous cyanobacterium Planktothrix rubescens, in a large, canyon-shaped, prealpine lake (Lake Zurich, Switzerland), in six vertical profiles along a 21.7-km longitudinal transect. Highest total densities and proportions of cells with high nucleic acid content were in the warm epilimnion and the hypoxic zone of the hypolimnion. P. rubescens formed a dense layer in the metalimnion throughout the lake, whereas picocyanobacteria populated the water layers above. The epilimnion was mainly inhabited by ultramicrobacteria related to the LD12-lineage of Alphaproteobacteria and to Actinobacteria; the latter group preferred the shallow regions. Cytophaga–Flavobacteria, in particular a population related to Fluviicola sp. were more frequent in and below the layer of maximal P. rubescens abundances. Betaproteobacteria, on the other hand, were highly abundant in the epi- and hypolimnion, but not in the P. rubescens layer. Four betaproteobacterial subpopulations with contrasting longitudinal and/or vertical habitat preferences were distinguished: putatively methylotrophic bacteria of the LD28 lineage (beta IV) preferentially inhabited the hypolimnion, Polynucleobacter acidiphobus was found throughout the epilimnion, Limnohabitans (R-BT065) more in the shallow regions of the lake, and Polynucleobacter necessarius ssp. asymbioticus only in hypoxic waters. Our results stress the potential importance of spatial niche differentiation in freshwater bacterioplankton.
Limnology and oceanography 04/2011; 56(6):2027-2039. · 3.42 Impact Factor
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ABSTRACT: Alphaproteobacteria are common members of marine bacterioplankton assemblages, but are believed to be rare in lacustrine systems. However, uncultured Alphaproteobacteria of the freshwater LD12 lineage form a tight monophyletic sister group with the numerically dominant bacteria in marine epipelagic waters, the SAR11 clade or genus Pelagibacter. Comparative rRNA sequence analysis reveals a global occurrence of LD12 bacteria in freshwater systems. The association of genotypic subclades with single-study systems moreover suggests a regional diversification. LD12 bacteria exhibit distinct and annually recurring spatio-temporal distribution patterns in prealpine lakes, as assessed by seasonally resolved vertical profiling and high-throughput cell counting. During the summer months, these ultramicrobacteria can form cell densities in the surface (epilimnetic) water layers that are comparable to those of their marine counterparts (>5 × 10(8) cells per l). LD12 bacteria had a pronounced preference for glutamine and glutamate over 7 other amino acids in situ, and they exhibited substantially higher uptake of these two substrates (and glycine) than the microbial assemblage in general. In addition, members of LD12 were also able to exploit other monomeric sources of organic carbon such as glucose, fructose or acetate. LD12 seemed to follow an oligotrophic lifestyle with slow but efficient uptake already at low substrate concentrations. Thus, LD12 bacteria do not only share phenotypic and metabolic traits with Pelagibacter, but also seem to thrive in the analogous spatiotemporal niche in freshwaters. The two groups together form one of the rare monophyletic lineages of ultramicrobacteria that have successfully traversed the barrier between marine and freshwater habitats.
The ISME Journal 03/2011; 5(8):1242-52. · 7.38 Impact Factor
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Aquatic Microbial Ecology 01/2009; 54(54):113. · 2.39 Impact Factor
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ABSTRACT: We investigated the diversity of planktonic Betaproteobacteria and the seasonal population changes of betaproteobacterial taxa in an oligo-mesotrophic lake (Piburger See, Austria). Focus was put on the vertical distribution of the investigated populations and on differences between their respective cell fractions with apparent amino acid incorporation. On average, 66% of betaproteobacterial cells and 73% of their diversity could be attributed to four clades within three lineages that were further analysed by fluorescence in situ hybridization. The numbers of bacteria from the R-BT subclade of the beta I lineage and from the PnecB subgroup of the beta II lineage were rather constant throughout the water column. In contrast, members of another subgroup of beta II (PnecC) and bacteria related to Methylophilus (beta IV) were particularly numerous in the oxygen-depleted zone. In general, only moderate seasonal changes in abundance were observed in the upper water layers, whereas there was a clear relationship between decreasing oxygen levels and the rise of bacteria from the PnecC and beta IV clades in deeper strata. On average, almost 80% of beta I bacteria, but < 15% of cells from the beta IV clade, showed amino acid incorporation. Our results suggest that the studied populations occupy distinct vertical and ecophysiological niches in Piburger See.
Environmental Microbiology 04/2008; 10(8):2074-86. · 5.84 Impact Factor
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ABSTRACT: We simultaneously studied the impact of top-down (protistan grazing) and bottom-up (phosphorus availability) factors on the numbers and biomasses of bacteria from various phylogenetic lineages, and on their growth and activity parameters in the oligo-mesotrophic Piburger See, Austria. Enhanced grazing resulted in decreased proportions of bacteria with high nucleic acid content (high-NA bacteria) and lower detection rates by FISH. There was a change in the composition of the bacterial assemblage, whereby Betaproteobacteria were heavily grazed while Alphaproteobacteria and Cytophaga-Flavobacterium-Bacteroides were less affected by predators. Changes in bacterial assemblage composition were also apparent in the treatments enriched with phosphorus, and even more pronounced in the incubations in dialysis tubes (allowing relatively free nutrient exchange). Here, Betaproteobacteria became dominant and appeared to act as successful opportunistic competitors for nutrients. In contrast, Actinobacteria did not respond to surplus phosphorus by population growth, and, moreover, maintained their small size, which resulted in a very low biomass contribution. In addition, significant relationships between high-NA bacteria and several bacterial phylogenetic clades were found, indicating an enhanced activity status. By combining several single-cell methods, new insight is gained into the competitive abilities of freshwater bacteria from a variety of phylogenetic lineages under contrasting sets of bottom-up and top-down constraints.
FEMS Microbiology Ecology 05/2007; 60(1):40-50. · 3.41 Impact Factor
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Aquatic Microbial Ecology 01/2007; 49(223-49):232. · 2.39 Impact Factor
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ABSTRACT: We tested a previously described protocol for fluorescence in situ hybridization of marine bacterioplankton with horseradish peroxidase-labeled rRNA-targeted oligonucleotide probes and catalyzed reporter deposition (CARD-FISH) in plankton samples from different lakes. The fraction of Bacteria detected by CARD-FISH was significantly lower than after FISH with fluorescently monolabeled probes. In particular, the abundances of aquatic Actinobacteria were significantly underestimated. We thus developed a combined fixation and permeabilization protocol for CARD-FISH of freshwater samples. Enzymatic pretreatment of fixed cells was optimized for the controlled digestion of gram-positive cell walls without causing overall cell loss. Incubations with high concentrations of lysozyme (10 mg ml(-1)) followed by achromopeptidase (60 U ml(-1)) successfully permeabilized cell walls of Actinobacteria for subsequent CARD-FISH both in enrichment cultures and environmental samples. Between 72 and >99% (mean, 86%) of all Bacteria could be visualized with the improved assay in surface waters of four lakes. For freshwater samples, our method is thus superior to the CARD-FISH protocol for marine Bacteria (mean, 55%) and to FISH with directly fluorochrome labeled probes (mean, 67%). Actinobacterial abundances in the studied systems, as detected by the optimized protocol, ranged from 32 to >55% (mean, 45%). Our findings confirm that members of this lineage are among the numerically most important Bacteria of freshwater picoplankton.
Applied and Environmental Microbiology 05/2003; 69(5):2928-35. · 3.83 Impact Factor
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ABSTRACT: Bacterivorous protists are known to induce changes in bacterial community composition (BCC). We hypothesized that changes in BCC could be related quantitatively to a measure of grazing: the ratio of bacterial mortality to growth rate. To test this hypothesis, we analyzed time-course changes in BCC, protistan grazing rate, and bacterial production from 3 in situ studies conducted in a freshwater reservoir and three laboratory studies. In the field experiments, samples were manipulated to yield different levels of protistan bacterivory and incubated in dialysis bags. Laboratory investigations were continuous cultivation studies in which different bacterivorous protists were added to bacterial communities. BCC was assessed using 4-6 different rRNA-targeted oligonucleotide probes for community analysis. Change in BCC (delta BCC) was estimated as the sum of changes in the proportions of the two phylogenetic groups that showed the largest shifts. Analysis of a set of 22 estimates of shifts in the ratio of grazing to production rate over periods of 48-72 h and A BCC showed that delta BCC was positively and tightly correlated (r2 = 0.784) with shifts in the ratio of grazing mortality to cell production. While the nature of a shift in BCC is unpredictable, the magnitude of the change can be related to changes in the balance between bacterial production and protistan grazing. bacterial community composition
Antonie van Leeuwenhoek 09/2002; 81(1-4):453-63. · 2.09 Impact Factor
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ABSTRACT: Since nearly one hundred years Traunsee experiences the import of tons of liquid and solid waste originating from salt and soda production. Today, the lake exhibits chloride concentrations of up to 170 mg L-1 and 19% of the lake floor are directly or indirectly influenced by industrial deposits (ID). Based on the comparison of several microbial parameters in unaffected, directly affected and intermediate lake bottom sediments, the ecological integrity of the lake was evaluated. The highly alkaline ID, which were exclusively colonized by microorganisms, harbored a bacterial community reduced by a factor of 10 in abundance and biomass compared to undisturbed sediment areas within the lake. The bacterial community of ID was furthermore characterized by a reduced content of actively respiring cells (INT-formazan reduction), a lower frequency of dividing cells (FDC) and a significantly reduced cell and biomass production. A 80 to 90% reduction in carbon recycling is estimated for the area exclusively covered by ID. Protists, although occasionally absent from the industrial sediments, were in general found to be less sensitive to the contaminant stress. Differences in alkalinity and dissolved organic carbon (DOC) concentrations of sediment porewaters as well as the total organic content and C/N ratios of sediments partly explain the microbial pattern observed at the various sampling sites. Possible consequences of the continuous industrial tailings for the whole lake ecosystem and the validation of the ecological integrity are discussed.
Water Air and Soil Pollution Focus 06/2002; 2(4):33-62.
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ABSTRACT: The holomictic Traunsee is the deepest and second largest lakein Austria. The special characteristic of this ecosystem isthe fact that local salt and soda industries presumably alterthe lake by the discharge of waste materials. Since thebeginning of the 20th century salt and soda works areannually releasing up to 50,000 tons of solid wastes and up to150,000 tons of chloride into Traunsee. To assess potentialeffects of these anthropogenic impacts on the bacterioplanktonthree sampling sites, influenced as well as not influenced bythe industrial discharge, were chosen for comparison andsampled monthly from November 1997 to October 1998. Bacterialabundance ranged between 0.4 to 3.0 106 cells ml-1 with decreasing numbers along the depth profile. Theproportion of actively respiring bacteria, i.e. INT [2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium chloride]reducing cells, never exceeded 10% of DAPI (4'',6''-diamidino-2-phenylindole) stained cells. Fluorescence in situ hybridization (FISH) was used to examine the seasonal and spatial distribution of dominant phylogenetic groups of thebacterioplankton. Up to 84% of bacteria detected with DAPIcould be detected via FISH applying the universal bacterialprobe EUB338. Percentages of alpha- and beta-Proteobacteriaand members of the Cytophaga-Flavobacterium cluster did notexceed 60% of DAPI-stained cells.Beta-Proteobacteriaappeared to be the most abundant group, not only in Traunsee butalso in two reference lakes, Attersee and Hallstttersee. No significant differences in any of the bacterial parameters couldbe detected between the three sampling sites and all measurementswere found in the range reported for oligotrophic lakes. The highdischarge of the Traun River, resulting in a lake water renewaltime of only one year, may diminish possible effects of industrial waste discharge in the pelagic zone.
Water Air and Soil Pollution Focus 06/2002; 2(4):137-163.
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ABSTRACT: Traunsee is a deep oligotrophic lake in Austria characterised by an artificial enrichment of chloride in the hypolimnion (up to 170 mg L-1) caused by waste disposal of soda and salt industries. Protists were collected monthly over one year, observed alive and after Quantitative Protargol Staining (ciliates) or via epifluorescence microscopy (heterotrophic flagellates). Three sites within the lake (0–40 m depths) were compared to deeper water layers from 60–160 m depths where chloride concentrations and conductivity were increased. In addition, we observed the protozooplankton of two neighbouring lakes, i.e. reference systems, during one sampling occasion. In Traunsee the abundance of ciliates was low (200–36 600 cells L-1) in contrast to high species diversity (at least 60 different species; HS = 2.6) throughout the year. The main pelagic species in terms of abundance were small oligotrichs and prostomatids like Rimostrombidium brachykinetum/hyalinum, Balanion planctonicum and Urotricha spp. throughout the investigation period. Among free-living heterotrophic flagellates, which occurred at densities of 40–2800 cells mL-1, small morphotypes dominated in the pelagial. No differences at the community level between the three lakes could be observed and pelagic ciliates and flagellates seemed not to be affected by increased chloride concentrations or by enhanced conductivity.
Water Air and Soil Pollution Focus 06/2002; 2(4):211-226.
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ABSTRACT: Background
Flow cytometry is an invaluable tool for the analysis of large series of samples in aquatic microbial ecology. However, analysis of the resulting data is often inefficient or does not reflect the complexity of natural communities. Because bacterioplankton assemblages frequently fall into several clusters with respect to their cellular properties, these subgroups seem to be a promising level of abstraction. Image analysis was used to detect clusters from flow cytometry data. The method was tested on a bacterial community under heavy protozoan grazing pressure.MethodsA bivariate histogram of flow cytometry data was transformed into a gray-scale image for image analysis. After low-pass filtration, regional maxima were delimited by a watershed algorithm. The resulting areas were then used as gates on the original measurements.ResultsThree clusters could be detected from the bacterial assemblage. Protozoan grazing had a strong impact on the bacterial community, which could be analyzed in detail at the level of individual subgroups.Conclusions
Investigation at the level of bacterial subgroups allowed a more detailed analysis than whole-community statistics and delivered essential and ecologically meaningful information. Image analysis proved to be an adequate tool to detect the subgroups without a priori knowledge. Cytometry 44:218–225, 2001. © 2001 Wiley-Liss, Inc.
Cytometry 06/2001; 44(3):218 - 225.
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ABSTRACT: Current models on protistan size-selective feeding assume that contact probability is the factor that largely explains observed food preferences. Contact probability is generally expected to be positively correlated with prey size and therefore to explain observed food selection for larger prey items. We critically tested these basic assumptions on size-selective feeding using the interception-feeding chrysomonad nanoflagellates Ochromonas sp. and Spumella sp. Mechanisms of differential feeding were studied during distinct stages of the selection process (i.e. contact probability, capture efficiency, ingestion efficiency, and differential digestion) by means of high-resolution video microscopy. Food selection was investigated using a mixture of microspheres ranging from 0.3-2.2 microm in diam., as well as a mixed bacterial community. In contrast to current model assumptions, the contact probability was highest for microspheres of intermediate size (0.9-1.2 microm), but was not generally positively correlated with prey size over the whole prey size range. Capture and ingestion also proved to be involved in size selection: these patterns were also independent of the food concentration (p = 0.968 for Ochromonas, p = 0.971 for Spumella). Even though the capture rate was significantly higher for attached flagellates than for swimming flagellates (p < 0.001), size selectivity was not affected (p > 0.05). Our results indicate that: (i) size selection is not actively regulated by these flagellates, but is a passive process; (ii) contact probability is not generally positively correlated with prey size, but shows a maximum for intermediate-sized prey in the prey size spectrum of 0.3-2.2 microm; and (iii) selection steps other than contact probability are crucial for size selection and should be integrated in models on size selection.
Journal of Eukaryotic Microbiology 51(6):626-33. · 2.66 Impact Factor
