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M Schloter

Publications and source records attributed to M Schloter.

At least 19 recordsLinked to original sources
PubMed2006

Archaea predominate among ammonia-oxidizing prokaryotes in soils.

Ammonia oxidation is the first step in nitrification, a key process in the global nitrogen cycle that results in the formation of nitrate through microbial activity. The increase in nitrate availability in soils is important for plant nutrition, but it also has considerable impact on groundwater pollution owing to leaching. Here we show that archaeal ammonia oxidizers are more abundant in soils than their well-known bacterial counterparts. We investigated the abundance of the gene encoding a subunit of the key enzyme ammonia monooxygenase (amoA) in 12 pristine and agricultural soils of three climatic zones. amoA gene copies of Crenarchaeota (Archaea) were up to 3,000-fold more abundant than bacterial amoA genes. High amounts of crenarchaeota-specific lipids, including crenarchaeol, correlated with the abundance of archaeal amoA gene copies. Furthermore, reverse transcription quantitative PCR studies and complementary DNA analysis using novel cloning-independent pyrosequencing technology demonstrated the activity of the archaea in situ and supported the numerical dominance of archaeal over bacterial ammonia oxidizers. Our results indicate that crenarchaeota may be the most abundant ammonia-oxidizing organisms in soil ecosystems on Earth.

Ammonia↗
PubMed2005

Characterization of bacterial community structure in rhizosphere soil of grain legumes.

Molecular techniques were used to characterize bacterial community structure, diversity (16S rDNA), and activity (16S rRNA) in rhizospheres of three grain legumes: faba beans (Vicia faba L., cv. Scirocco), peas (Pisum sativum L., cv. Duel) and white lupin (Lupinus albus L., cv. Amiga). All plants were grown in the same soil under controlled conditions in a greenhouse and sampled after fruiting. Amplified 16S rDNA and rRNA products (using universal bacterial primers) were resolved by denaturing gradient gel electrophoresis (DGGE). Distinct profiles were observed for the three legumes with most of the bands derived from RNA being a subset of those derived from DNA. Comparing the total bacterial profiles with actinomycete-specific ones (using actinomycete-specific primers) highlighted the dominance of this group in the three rhizospheres. 16S PCR and RT-PCR products were cloned to construct libraries and 100 clones from each library were sequenced. Actinomycetes and proteobacteria dominated the clone libraries with differences in the groups of proteobacteria. Absence of beta-subdivision members in pea and gamma-subdivision members of proteobacteria in faba bean rhizosphere was observed. Plant-dependent rhizosphere effects were evident from significant differences in the bacterial community structure of the legume rhizospheres under study. The study gives a detailed picture of both residing and "active" bacterial community in the three rhizospheres. The high abundance of actinomycetes in the rhizospheres of mature legumes indicates their possible role in soil enrichment after the legumes are plowed into the soil as biofertilizers.

DNA, Bacterial↗
PubMed2005

Mycorrhizosphere responsiveness to atmospheric ozone and inoculation with Phytophthora citricola in a phytotron experiment with spruce/beech mixed cultures.

The aim was to analyze functional changes in the mycorrhizosphere (MR) of juvenile spruce and beech grown in a mixture under ambient and twice ambient ozone and inoculated with the root pathogen Phytophthora citricola. The phytotron experiment was performed over two vegetation periods, adding the pathogen at the end of the first growing season. Root biomass data suggest that the combined treatment affected spruce more than beech and that the negative influence of ozone on stress tolerance against the root pathogen P. citricola was greater for spruce than for beech. In contrast, beech was more affected when the pathogen was the sole stressor. The functional soil parameter chosen for studies of MR soil samples was activity of extracellular enzymes. After the first year of ozone exposure, MR soil samples of both species showed increased activity of almost all measured enzymes (acid phosphatase, chitinase, beta-glucosidase, cellobiohydrolase) in the O3 treatment. Species-specific differences were observed, with a stronger effect of P. citricola on beech MR and a stronger ozone effect on spruce MR. In the second year, the effects of the combined treatment (ozone and P. citricola) were a significant increase in the activity of most enzymes (except cellobiohydrolase) for both tree species. The results indicated that responsiveness of MR soils towards ozone and P. citricola was related to the severity of infection of the plants and the reduction of belowground biomass, suggesting a strong, direct influence of plant stress on MR soil enzyme activity. Additional research is needed using different species and combined stresses to determine the broader ecological relevance of shifts in rhizosphere enzymes.

Atmosphere↗
PubMed2005

Short term effects of ozone on the plant-rhizosphere-bulk soil system of young beech trees.

Plant growth largely depends on microbial community structure and function in the rhizosphere. In turn, microbial communities in the rhizosphere rely on carbohydrates provided by the host plant. This paper presents the first study on ozone effects in the plant-rhizosphere-bulk soil system of 4-year-old beech trees using outdoor lysimeters as a research platform. The lysimeters were filled with homogenized soil from the corresponding horizons of a forest site, thus minimizing field heterogeneity. Four lysimeters were treated with ambient ozone (1 x O3) and four with double ambient ozone concentrations (2 x O3; restricted to 150 ppb). In contrast to senescence, which was almost unaffected by ozone treatment, both the photochemical quantum yield of photosystem II (PSII) and leaf gas exchange were reduced (11 - 45 %) under the elevated O3 regime. However, due to large variation between the plants, no statistically significant O3 effect was found. Even though the amount of primary metabolites, such as sugar and starch, was not influenced by elevated O3 concentrations, the reduced photosynthetic performance was reflected in leaf biochemistry in the form of a reduction in soluble phenolic metabolites. The rhizosphere microbial community also responded to the O3 treatment. Both community structure and function were affected, with a tendency towards a lower diversity and a significant reduction in the potential nutrient turnover. In contrast, litter degradation was unaffected by the fumigation, indicating that in situ microbial functionality of the bulk soil did not change.

Carbohydrate Metabolism↗
PubMed2004

RNA fingerprinting of microbial community in the rhizosphere soil of grain legumes.

Microbial structural and expression profiles of the rhizospheres of three legumes, faba beans, peas and white lupin, were compared by RNA-arbitrarily primed PCR technique. Two different primers, M13 reverse and 10-mer primers, were used in the amplification and products resolved on non-denaturing polyacrylamide gel. With both DNA and RNA profiles Lupinus and Pisum rhizospheres were more similar to each other than to Vicia rhizosphere. The RAP-PCR products were also dot blotted and probed for bacterial peptidase transcripts. Plant-dependent rhizosphere effect was evident by the marked absence of transcripts for bacterial neutral metallopeptidase in Lupinus rhizosphere. The results of dot blot were further confirmed by RT-PCR for the expression of bacterial neutral metallopeptidase in the three rhizospheres.

Bacteria↗
PubMed2004

A rapid and highly sensitive method for measuring enzyme activities in single mycorrhizal tips using 4-methylumbelliferone-labelled fluorogenic substrates in a microplate system.

A microplate fluorimetric assay was developed for measuring potential activities of extracellular enzymes of individual ectomycorrhizal (EM) roots using methylumbelliferone (MU)-labelled fluorescent substrate analogues and microsieves to minimise damage due to manipulation of excised mycorrhizal roots. Control experiments revealed that enzyme activities remained stable over the whole time of the experiment suggesting a strong affinity of the studied enzymes to the fungal cell walls. The same mycorrhizal tips thus could be used repeatedly for enzyme detection and subsequently analysed for the projection area by automated image analysis. The developed system was evaluated on four different EM species measuring pH optimum and substrate saturation of phosphatase, chitinase and beta-glucosidase. The four EM species studied were Lactarius subdulcis, Russula ochroleuca, Cortinarius obtusus and Xerocomus cf. chrysenteron. Depending upon the enzyme, each species exhibited different levels of enzymatic activities as well as enzyme kinetics and showed also differences in pH optima.

Chitinases↗
PubMed2003

A TaqMan-PCR protocol for quantification and differentiation of the phytopathogenic Clavibacter michiganensis subspecies.

Real-time TaqMan-PCR assays were developed for detection, differentiation and absolute quantification of the pathogenic subspecies of Clavibacter michiganensis (Cm) in one single PCR run. The designed primer pair, targeting intergenic sequences of the rRNA operon (ITS) common in all subspecies, was suitable for the amplification of the expected 223-nt DNA fragments of all subspecies. Closely related bacteria were completely discriminated, except of Rathayibacter iranicus, from which weak PCR product bands appeared on agarose gel after 35 PCR cycles. Sufficient specificity of PCR detection was reached by introduction of the additional subspecies specific probes used in TaqMan-PCR. Only Cm species were detected and there was clear differentiation among the subspecies C. michiganensis sepedonicus (Cms), C. michiganensis michiganensis (Cmm), C. michiganensis nebraskensis (Cmn), C. michiganensis insidiosus (Cmi) and C. michiganensis tessellarius (Cmt). The TaqMan assays were optimized to enable a simultaneous quantification of each subspecies. Validity is shown by comparison with cell counts.

Actinomycetales↗
PubMed2002

Influence of nonylphenol on the microbial community of lake sediments in microcosms.

In this study the impact of nonylphenol, an estrogenic degradation product of alkylphenol polyethoxylates, on the microbial community structure in contaminated sediments of aquatic microcosms using in situ hybridization with fluorescently labeled oligonucleotides probes was investigated. A positive correlation between nonylphenol concentration and cell numbers of bacteria and microfungi as well as an increase in the numbers of active bacteria was found. However, the ratio between total microorganisms and active bacteria remained unchanged. A large fraction of the cells could be identified using group specific oligonucleotide probes. A slight change in the composition of the microbial community structure was observed, with Gram-positive bacteria with high DNA G + C-content becoming more abundant at higher concentrations of nonylphenol.

Bacteria↗
PubMed2002

Enumeration of total bacteria and bacteria with genes for proteolytic activity in pure cultures and in environmental samples by quantitative PCR mediated amplification.

Real-time quantitative PCR assays were developed for the absolute quantification of different groups of bacteria in pure cultures and in environmental samples. 16S rRNA genes were used as markers for eubacteria, and genes for extracellular peptidases were used as markers for potentially proteolytic bacteria. For the designed 16S rDNA TaqMan assay, specificity of the designed primer-probe combination for eubacteria, a high amplification efficiency over a wide range of starting copy numbers and a high reproducibility is demonstrated. Cell concentrations of Bacillus cereus, B. subtilis and Pseudomonas fluorescens in liquid culture were monitored by TaqMan-PCR using the 16S rDNA target sequence of Escherichia coli as external standard for quantification. Results agree with plate counts and microscopic counts of DAPI stained cells. The significance of 16S rRNA operon multiplicity to the quantification of bacteria is discussed.Furthermore, three sets of primer pair together with probe previously designed for targeting different classes of bacterial extracellular peptidases were tested for their suitability for TaqMan-PCR based quantification of proteolytic bacteria. Since high degeneracy of the probes did not allow accurate quantification, SybrGreen was used instead of molecular probes to visualize and quantify PCR products during PCR. The correlation between fluorescence and starting copy number was of the same high quality as for the 16S rDNA TaqMan assay for all the three peptidase gene classes. The detected amount of genes for neutral metallopeptidase of B. cereus, for subtilisin of B. subtilis and for alkaline metallopeptidase of P. fluorescens corresponded exactly to the numbers of bacteria investigated by the 16S rDNA targeting assay. The developed assays were applied for the quantification of bacteria in soil samples.

Bacillus cereus↗
PubMed2002

Development and characterization of murine monoclonal antibodies specific for dissimilatoric copper nitrite reductase.

Several hybridoma cell lines from mice were established, producing monoclonal antibodies (MAbs) directed against the dissimilatoric copper nitrite reductase (dNIR) to detect actual denitrifying bacteria at the single cell level under nondestructive conditions in the environment. The mice were immunized with native or recombinant enzyme gained from two different bacteria, Ochrobactrum anthropi and Alcaligenes faecalis. The antibodies obtained could be divided into two groups according to their different specificities for dNIRs of different bacteria: One group of MAbs had a broad specificity for dissimilatoric copper nitrite reductases from bacteria of different phylogenetic taxa; the other group gave only a clear signal with the corresponding immunogen. None of the raised MAbs showed a cross reactivity with the dissimilatoric heme nitrite reductase. One MAb from each group (MAb dNIR1a and MAb dNIR29) has been selected for further investigation. Data of enzyme-linked immunosorbent assay (ELISA), Western blotting, and immunofluorescence-microscopy are presented and compared with phylogenetic data. Furthermore, results of Western blotting experiments with cells, grown without nitrate under aerobic conditions, and cells cultivated with nitrate under anaerobiosis, are shown.

Alcaligenes↗
PubMed2001

PCR primers and functional probes for amplification and detection of bacterial genes for extracellular peptidases in single strains and in soil.

A set of primers and functional probes was developed for the detection of peptidase gene fragments of proteolytic bacteria. Based on DNA sequence data, degenerate PCR primers and internal DIG-labeled probes specific for genes encoding alkaline metallopeptidases (apr) (E.3.4.24), neutral metallopeptidases (npr) (E.3.4.24) and serine peptidases (sub) (E.3.4.21) were derived by multiple sequence alignments. Type strains with known peptidase genes and proteolytic bacteria from a grassland rhizosphere soil, a garden soil and an arable field were investigated for their genotypic proteolytic potential. For 52 out of 53 proteolytic bacterial isolates, at least one of the three peptidase classes could be identified by this approach. The amplified gene fragments were of the expected sizes with each of the three primer sets. The functional probes APR, NPR and SUB have been shown to hybridize specifically to the corresponding gene fragments. sub and npr genes were mainly found in Bacillus species. apr genes were only found in the Pseudomonas fluorescens biotypes and in two morphologically identical Flavobacterium-Cytophaga strains from two different sites. In most of the Bacillus spp., both sub and the npr and in the Flavobacterium-Cytophaga strains even all the three genes could be detected. PCR with DNA isolated from soil led to one main product of the expected size with each primer pair whose identity was additionally confirmed by Southern blot hybridization with the corresponding probes.

Bacteria↗
PubMed2000

Taxonomic characterization of Ochrobactrum sp. isolates from soil samples and wheat roots, and description of Ochrobactrum tritici sp. nov. and Ochrobactrum grignonense sp. nov.

A large collection of bacterial strains, immunotrapped from soil and from the wheat rhizoplane, was subjected to polyphasic taxonomy by examining various pheno- and genotypic parameters. Strains were grouped on (inter) repetitive extragenic palindromic DNA (REP) PCR profiles at the intraspecies level. Pheno- and genotypic characters were assessed for representatives from 13 different REP groups. Strains of nine REP groups constituting two physiological BIOLOG clusters fell in the coherent DNA-DNA reassociation group of Ochrobactrum anthropi. Strains of two REP groups constituting a separate BIOLOG cluster fell in the coherent DNA-DNA reassociation group of Ochrobactrum intermedium. Additional phenotypic characters differentiating O. anthropi and O. intermedium were found. REP group K strains constituted a different BIOLOG cluster, a separate DNA-DNA reassociation group and a distinct phylogenetic lineage in 165 rDNA homology analysis, indicating that REP group K strains represent a new species. Diagnostic phenotypic characters were found. Closest relatives were Ochrobactrum species. The name Ochrobactrum grignonense sp. nov. is proposed (type strain OgA9aT = LMG 18954T = DSM 13338T). REP group J strains again constituted a different BIOLOG cluster, a separate DNA-DNA reassociation group and showed, as a biological particularity, a strict preference for the rhizoplane as habitat. Diagnostic phenotypic characters were found. This indicated that REP group J strains represent a further new species, although phylogenetic analyses using 16S rDNA homology were not able to separate the cluster of REP group J sequences significantly from 16S rDNA sequences of Ochrobactrum anthropi. The name Ochrobactrum tritici sp. nov. is proposed (type strain SCII24T = LMG 18957T = DSM 13340T).

Alphaproteobacteria↗
PubMed2000

Ecology and evolution of bacterial microdiversity.

Using high resolution molecular fingerprinting techniques like random amplification of polymorphic DNA, repetitive extragenic palindromic PCR and multilocus enzyme electrophoresis, a high bacterial diversity below the species and subspecies level (microdiversity) is revealed. It became apparent that bacteria of a certain species living in close association with different plants either as associated rhizosphere bacteria or as plant pathogens or symbiotic organisms, typically reflect this relationship in their genetic relatedness. The strain composition within a population of soil bacterial species at a given field site, which can be identified by these high resolution fingerprinting techniques, was markedly influenced by soil management and soil features. The observed bacterial microdiversity reflected the conditions of the habitat, which select for better adapted forms. In addition, influences of spatial separation on specific groupings of bacteria were found, which argue for the occurrence of isolated microevolution. In this review, examples are presented of bacterial microdiversity as influenced by different ecological factors, with the main emphasis on bacteria from the natural environment. In addition, information available from some of the first complete genome sequences of bacteria (Helicobacter pylori and Escherichia coli) was used to highlight possible mechanisms of molecular evolution through which mutations are created; these include mutator enzymes. Definitions of bacterial species and subspecies ranks are discussed in the light of detailed information from whole genome typing approaches.

Bacteria↗
PubMed2000

Molecular analysis of ammonia oxidation and denitrification in natural environments.

This review summarizes aspects of the current knowledge about the ecology of ammonia-oxidizing and denitrifying bacteria. The development of molecular techniques has contributed enormously to the rapid recent progress in the field. Different techniques for doing so are discussed. The characterization of ammonia-oxidizing and -denitrifying bacteria by sequencing the genes encoding 16S rRNA and functional proteins opened the possibility of constructing specific probes. It is now possible to monitor the occurrence of a particular species of these bacteria in any habitat and to get an estimate of the relative abundance of different types, even if they are not culturable as yet. These data indicate that the composition of nitrifying and denitrifying communities is complex and apparently subject to large fluctuations, both in time and in space. More attempts are needed to enrich and isolate those bacteria which dominate the processes, and to characterize them by a combination of physiological, biochemical and molecular techniques. While PCR and probing with nucleotides or antibodies are primarily used to study the structure of nitrifying and denitrifying communities, studies of their function in natural habitats, which require quantification at the transcriptional level, are currently not possible.

Ammonia↗
PubMed1997

Characterization and application of a strain-specific monoclonal antibody against the rhizosphere bacterium Azospirillum brasilense Wa5.

A hybridoma cell line producing a rat monoclonal antibody (MAb Bo-33) directed against lipopolysaccharide of Azospirillum brasilense Wa5 has been established and characterized. Whole bacteria were used as immunogens. The number of antigens per cell was about 1500. The number of antigens per cell of reisolates from the rhizosphere of what was similar to the number of antigens of bacteria cultivated in rich medium. The sensitivity of detection using MAb Bo-33 was about 100 bacteria/ml. Therefore, the MAb was suitable for in situ immunofluorescence detection and a sensitive direct quantification of Azospirillum brasilense Wa5 in rhizosphere extracts.

Animals↗
PubMed1997

Root colonization of different plants by plant-growth-promoting Rhizobium leguminosarum bv. trifolii R39 studied with monospecific polyclonal antisera.

Monospecific polyclonal antisera raised against Rhizobium leguminosarum bv. trifolii R39, a bacterium which was isolated originally from red clover nodules, were used to study the colonization of roots of leguminous and nonleguminous plants (Pisum sativum, Lupinus albus, Triticúm aestivum, and Zea mays) after inoculation. Eight weeks after inoculation of soil-grown plants, between 0.1 and 1% of the total bacterial population in the rhizospheres of all inoculated plants were identified as R. leguminosarum bv. trifolii R39. To characterize the associative colonization of the nonleguminous plants by R.leguminosarum bv. trifolii R39 in more detail, a time course study was performed with inoculated roots of Z. mays. R. leguminosarum bv. trifolii R39 was found almost exclusively in the rhizosphere soil and on the rhizoplane 4 weeks after inoculation. Colonization of inner root tissues was detected only occasionally at this time. During the process of attachment of R. leguminosarum bv. trifolii R39 to the rhizoplane, bacterial lipopolysaccharides were overexpressed, and this may be important for plant-microbe interaction. Fourteen weeks after inoculation, microcolonies of R. leguminosarum bv. trifolii R39 were detected in lysed cells of the root cortex as well as in intracellular space of central root cylinder cells. At the beginning of flowering (18 weeks after inoculation), the number of R. leguminosarum bv. trifolii R39 organisms decreased in the rhizosphere soil, rhizoplane, and inner root tissue.

Antigens, Bacterial↗
PubMed1997

Direct examination of subgingival plaque from a diseased periodontal site using confocal laser scanning microscopy (CLSM).

Material taken directly from a periodontal site was investigated using immunofluorescence, acridine orange staining and confocal laser scanning microscopy (CLSM). Porphyromonas gingivalis was tracked by a specific polyclonal antibody and its pronounced occurrence in inflamed as compared to non-inflamed areas was demonstrated. Further accompanying microorganisms were counterstained with acridine orange which could provide information on the viability of individual cells. Optical sections by laser microscopy revealed the spatial arrangement of the investigated material. The combination of specific staining and CLSM allows a detailed microbiological investigation of clinical material obtained directly without cultivation.

Antibodies, Bacterial↗