Search

A complex microbial system consisting of six different interconnected localities was thoroughly investigated at full scale for over a year. The metacommunity concept originating from macro-ecology was used to uncover mechanisms of community assembly by observing microbial interrelationships in and between the different localities via correlation and network analysis. The individual-based observation approach was applied using high-throughput microbial community cytometry in addition to next generation sequencing. We found robust α-diversity values for each of the six localities and high β-diversity values despite directed connectivity between localities, classifying for endpoint assembly of organisms in each locality. Endpoint characteristics were based on subcommunities with high cell numbers whereas those with lower cell numbers were involved in dispersal. Perturbation caused abiotic parameters to alter local community assembly with especially the rare cells announcing community restructuration processes. The mass-effect paradigm as part of the metacommunity concept was identified by an increase in interlocality biotic correlations under perturbation which, however, did not unbalance the predominant species-sorting paradigm in the studied full scale metacommunity. Data as generated in this study might contribute to the development of individual-based models for controlling managed multispecies natural systems in future.

Micro-organisms are known to degrade a wide range of toxic substances. How the environment shapes microbial communities in polluted ecosystems and thus influences degradation capabilities is not yet fully understood. In this study, we investigated microbial communities in a highly complex environment: the capillary fringe and subjacent sediments in a hydrocarbon-contaminated aquifer. Sixty sediment sections were analysed using terminal restriction fragment length polymorphism (T-RFLP) fingerprinting, cloning and sequencing of bacterial and archaeal 16S rRNA genes, complemented by chemical analyses of petroleum hydrocarbons, methane, oxygen and alternative terminal electron acceptors. Multivariate statistics revealed concentrations of contaminants and the position of the water table as significant factors shaping the microbial community composition. Micro-organisms with highest T-RFLP abundances were related to sulphate reducers belonging to the genus Desulfosporosinus, fermenting bacteria of the genera Sedimentibacter and Smithella, and aerobic hydrocarbon degraders of the genus Acidovorax. Furthermore, the acetoclastic methanogens Methanosaeta, and hydrogenotrophic methanogens Methanocella and Methanoregula were detected. Whereas sulphate and sulphate reducers prevail at the contamination source, the detection of methane, fermenting bacteria and methanogenic archaea further downstream points towards syntrophic hydrocarbon degradation.

The influences of poorly water-soluble anthracene on ester-linked phospholipid fatty acid (PLFA) and glycolipid fatty acid (GLFA) profiles of Mycobacterium sp. LB501T were studied. Bacteria were cultivated on either anthracene or glucose (one culture with successively amended small doses of this substrate and one with excess concentrations) to distinguish between influences of the chemical structure and the bioavailability of the growth substrate. Results revealed that GLFA and PLFA profiles of M . sp. LB501T depended on the availability and the structure of the carbon source. Fatty acid profiles obtained with anthracene differed from those obtained with excess glucose. They were interpreted as a specific adaptation to this poorly bioavailable polycyclic aromatic hydrocarbon (PAH). In contrast, profiles obtained with low glucose concentrations showed clear signs of starvation stress. Stable carbon isotopic ratios (delta(13) C) of GLFA and PLFA of M. sp. LB501T were analysed to characterize the (13) C-fractionation during the biosynthesis of individual fatty acids and to evaluate their value as markers for substrate usage. Although the delta(13) C values of PLFA and GLFA showed differential isotope fractionation during anthracene- and glucose-degradation, they were sufficiently distinct to be used as signatures of bacterial substrate usage.

A method for the quantitative analysis of the hydrophobicity of the mycelial mat of filamentous fungi based on contact angle measurements is presented. It was tested for a range of fungi belonging to the classes of basidiomycetes, ascomycetes and deuteromycetes. The measured contact angles of the mycelial mats ranged between hydrophilic (30 degrees) for the deuteromycetes Fusarium oxysporum Fo47 GUS1 and Trichoderma harzianum P1[pZEGA1] and hydrophobic (60 degrees) for the ascomycete Cladosporium sp. DSE48.1b and the basidiomycetes Paxillus involutus WSL 37.7, Hebeloma crustiliniforme WSL 6.2, Suillus bovinus WSL 48.1 and Laccaria bicolor WSL 73.1. For some fungi, variations in the hydrophobicity of the mycelium depending on the growth medium, the physiological state and the exposure to water were distinguished.

The influence of growth substrates on mycolic acid profiles of PAH-degrading Mycobacterium spp. LB501T, LB307T and VM552 was examined by high-performance liquid chromatography (HPLC) using glucose, alkanes, polycyclic aromatic hydrocarbons (PAH) or Luria-Bertani medium (LB) as the sole carbon source. The substrates gave rise to varying mycolic acid profiles, as bacteria growing on poorly water-soluble substrates exhibited more hydrophobic mycolic acids than cells grown on glucose. Our results indicate that mycobacteria respond to the growth substrate by changing the mycolic acid composition of their cell wall, pointing at the importance of the growth substrate for mycolic acid profiling as an identification method of actinomycetes.

Diffusion of octane from oily droplets in different microscale settings was measured using Escherichia coli expressing the stable green fluorescent protein (GFP) from the alkB promoter of Pseudomonas oleovorans. GFP fluorescence intensities were determined quantitatively at the single-cell level after 1.0 or 2.5 h incubation and compared with different calibration series using known concentrations of octane. By immobilizing the E. coli sensor cells on the bottom glass plate of a microscope flow chamber, it was possible to monitor the diffusion process for octane in aqueous solution as a function of time and distance from non-aqueous phase droplets of octane alone or oily octane mixtures. When a gas phase was included in the flow chambers, octane transport could be demonstrated from the oily mixtures to the cells through both gas and liquid phase. Assays of non-immobilized sensor cells in microdroplets in the presence or absence of soil particles incubated with octane through the vapour phase revealed a slight reduction in the total amount of induced E. coli cells in the presence of soil. Our results indicate the power of using GFP-marked single-cell biosensors in determining microscale bioavailability of organic pollutants.

Micro-organisms are known to degrade a wide range of toxic substances. How the environment shapes microbial communities in polluted ecosystems and thus influences degradation capabilities is not yet fully understood. In this study, we investigated microbial communities in a highly complex environment: the capillary fringe and subjacent sediments in a hydrocarbon-contaminated aquifer. Sixty sediment sections were analysed using terminal restriction fragment length polymorphism (T-RFLP) fingerprinting, cloning and sequencing of bacterial and archaeal 16S rRNA genes, complemented by chemical analyses of petroleum hydrocarbons, methane, oxygen and alternative terminal electron acceptors. Multivariate statistics revealed concentrations of contaminants and the position of the water table as significant factors shaping the microbial community composition. Micro-organisms with highest T-RFLP abundances were related to sulphate reducers belonging to the genus Desulfosporosinus, fermenting bacteria of the genera Sedimentibacter and Smithella, and aerobic hydrocarbon degraders of the genus Acidovorax. Furthermore, the acetoclastic methanogens Methanosaeta, and hydrogenotrophic methanogens Methanocella and Methanoregula were detected. Whereas sulphate and sulphate reducers prevail at the contamination source, the detection of methane, fermenting bacteria and methanogenic archaea further downstream points towards syntrophic hydrocarbon degradation.

Contaminant biodegradation in soil is frequently limited by hindered physical access of bacteria to the contaminants. In the frame of the development of novel bioremediation approaches based on ecological principles, we tested the hypothesis that fungal networks facilitate the movement of bacteria by providing continuous liquid films in which gradients of chemoattractants can form and chemotactic swimming can take place. Unlike bacteria, filamentous fungi spread with ease in water-unsaturated soil. In a simple laboratory model of a water-unsaturated environment, we studied the movement of polycyclic aromatic hydrocarbon-degrading Pseudomonas putida PpG7 (NAH7) along a mycelium of Pythium ultimum. Some undirected dispersal was observed in the absence of a chemoattractant or when the non-chemotactic derivative strain P. putida G7.C1 (pHG100) was used. The bacterial movement became fourfold more effective and clearly directed when the chemotactic wild type was used and salicylate was present as a chemoattractant. No dispersal of bacteria was found in the absence of the fungus. These findings point at a role of mycelia for the translocation of chemicals and microorganisms. The results suggest that fungi improve the accessibility of contaminants in water-unsaturated environments.

Bacterial diversity in 16S ribosomal DNA and reverse-transcribed 16S rRNA clone libraries originating from the heavy metal-contaminated rhizosphere of the metal-hyperaccumulating plant Thlaspi caerulescens was analysed and compared with that of contaminated bulk soil. Partial sequence analysis of 282 clones revealed that most of the environmental sequences in both soils affiliated with five major phylogenetic groups, the Actinobacteria, alpha-Proteobacteria, beta-Proteobacteria, Acidobacteria and the Planctomycetales. Only 14.7% of all phylotypes (sequences with similarities >97%), but 45% of all clones, were common in the rhizosphere and the bulk soil clone libraries. The combined use of rDNA and rRNA libraries indicated which taxa might be metabolically active in this soil. All dominant taxa, with the exception of the Actinobacteria, were relatively less represented in the rRNA libraries compared with the rDNA libraries. Clones belonging to the Verrucomicrobiales, Firmicutes, Cytophaga-Flavobacterium-Bacteroides and OP10 were found only in rDNA clone libraries, indicating that they might not represent active constituents in our samples. The most remarkable result was that sequences belonging to the Actinobacteria dominated both bulk and rhizosphere soil libraries derived from rRNA (50% and 60% of all phylotypes respectively). Seventy per cent of these clone sequences were related to the Rubrobacteria. subgroups 2 and 3, thus providing for the first time evidence that this group of bacteria is probably metabolically active in heavy metal-contaminated soil.

The influences of poorly water-soluble anthracene on ester-linked phospholipid fatty acid (PLFA) and glycolipid fatty acid (GLFA) profiles of Mycobacterium sp. LB501T were studied. Bacteria were cultivated on either anthracene or glucose (one culture with successively amended small doses of this substrate and one with excess concentrations) to distinguish between influences of the chemical structure and the bioavailability of the growth substrate. Results revealed that GLFA and PLFA profiles of M. sp. LB501T depended on the availability and the structure of the carbon source. Fatty acid profiles obtained with anthracene differed from those obtained with excess glucose. They were interpreted as a specific adaptation to this poorly bioavailable polycyclic aromatic hydrocarbon (PAH). In contrast, profiles obtained with low glucose concentrations showed clear signs of starvation stress. Stable carbon isotopic ratios (delta13C) of GLFA and PLFA of M. sp. LB501T were analysed to characterize the 13C-fractionation during the biosynthesis of individual fatty acids and to evaluate their value as markers for substrate usage. Although the delta13C values of PLFA and GLFA showed differential isotope fractionation during anthracene- and glucose-degradation, they were sufficiently distinct to be used as signatures of bacterial substrate usage.