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11

Electronic Resource

Analysis of bacterial and protozoan communities in an aquifer contaminated with monoaromatic hydrocarbons (1998)

Zarda, Boris ; Mattison, Geoffrey ; Hess, Annatina ; [et al.]

Oxford, UK : Blackwell Publishing Ltd

FEMS microbiology ecology 27 (1998), S. 0

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ISSN: 1574-6941

Source: Blackwell Publishing Journal Backfiles 1879-2005

Topics: Biology

Notes: Bacterial and protozoan communities were examined in three cores (A, B and C) from an aquifer located at an abandoned refinery near Hünxe, Germany. Cores were removed along a transect bordering a plume containing various monoaromatic hydrocarbons. Monoaromatic hydrocarbons could not be detected in the unsaturated zone in any core but were present in the saturated zones of core C (between 280 and 42 600 μmol kg−1 of core material [dry wt.]) and cores A and B (between 30 and 190 μmol kg−1 of core material [dry wt.]). Xylene isomers accounted for 50–70% of monoaromatic hydrocarbons in all cores. The number of DAPI-stained bacteria was found to increase from the low-contaminated cores A and B (approx. 0.1×108 cells and 0.2×108 cells g−1 of core material [dry wt.], respectively) to the high-contaminated core C (2.4×108 cells g−1 of core material [dry wt.]). The higher bacterial numbers in core C were found to coincide with a higher detection rate obtained by in situ hybridization using probe Eub338 to target the domain Bacteria (13–42% for core C as compared to 3–25% for cores A and B, respectively). Proteobacteria of the δ-subdivision (which includes many sulfate-reducing bacteria) were the most predominant of the groups investigated (7–15% of DAPI-stained bacteria) and were followed by Proteobacteria of the γ- and β-subdivisions (4% and 1% of DAPI-stained bacteria, respectively). The total numbers of protozoa and bacteria determined by direct counting occurred in a ratio of approx. 1:103, which was independent of depth or core examined. Most probable number analysis combined with a subsequent classification of the culturable protozoa revealed nanoflagellates as the major component of the protozoan community. Naked amoebae became increasingly more encysted with depth, except in the high-contaminated core C where vegetative trophozoites were present in the saturated zone. The co-occurrence of bacteria and protozoa in association with high concentrations of monoaromatic hydrocarbons suggests the involvement of trophic interactions in the process of biodegradation.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1111/j.1574-6941.1998.tb00532.x

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12

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Field-scale isotopic labeling of phospholipid fatty acids from acetate-degrading sulfate-reducing bacteria (2005)

Pombo, Silvina A. ; Kleikemper, Jutta ; Schroth, Martin H. ; [et al.]

Oxford, UK : Blackwell Publishing Ltd

FEMS microbiology ecology 51 (2005), S. 0

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ISSN: 1574-6941

Source: Blackwell Publishing Journal Backfiles 1879-2005

Topics: Biology

Notes: Isotopic labeling of biomarker molecules is a technique applied to link microbial community structure with activity. Previously, we successfully labeled phospholipid fatty acids (PLFA) of suspended nitrate-reducing bacteria in an aquifer. However, the application of the method to low energy-yielding processes such as sulfate reduction, and extension of the analysis to attached communities remained to be studied. To test the feasibility of the latter application, an anoxic test solution of 500 l of groundwater with addition of 0.5 mM Br− as a conservative tracer, 1.1 mM SO2−4, and 2.0 mM [2-13C]acetate was injected in the transition zone of a petroleum hydrocarbon-contaminated aquifer where sulfate-reducing and methanogenic conditions prevailed. Thousand liters of test solution/groundwater mixture were extracted in a stepwise fashion after 2–46 h incubation. Computed apparent first-order rate coefficients were 0.31 ± 0.04 day−1 for acetate and 0.34 ± 0.05 day−1 for SO2−4 consumption. The δ13C increased from −71.03‰ to +3352.50‰ in CH4 and from −16.15‰ to +32.13‰ in dissolved inorganic carbon (DIC). A mass balance suggested that 43% of the acetate-derived 13C appeared in DIC and 57% appeared in CH4. Thus, acetate oxidation coupled to sulfate reduction and acetoclastic methanogenesis occurred simultaneously. The δ13C of PLFA increased on average by 27‰ in groundwater samples and 4‰ in sediment samples. Hence, both suspended and attached communities actively degraded acetate. The PLFA labeling patterns and fluorescent in situ hybridization (FISH) analyses of sediment and groundwater samples suggested that the main sulfate-reducing bacteria degrading the acetate were Desulfotomaculum acetoxidans and Desulfobacter sp. in groundwater, and D. acetoxidans in sediment.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1016/j.femsec.2004.08.010

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13

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Tracing toluene-assimilating sulfate-reducing bacteria using 13C-incorporation in fatty acids and whole-cell hybridization (2001)

Pelz, Oliver ; Chatzinotas, Antonis ; Zarda-Hess, Annatina ; [et al.]

Oxford, UK : Blackwell Publishing Ltd

FEMS microbiology ecology 38 (2001), S. 0

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ISSN: 1574-6941

Source: Blackwell Publishing Journal Backfiles 1879-2005

Topics: Biology

Notes: Polar lipid-derived fatty acids (PLFA) commonly found in sulfate-reducing bacteria were detected in high abundance in the sediment harvested from a monitoring well of a petroleum-hydrocarbon (PHC)-contaminated aquifer. Aquifer microcosms were incubated under sulfate-reducing conditions with [methyl-14C]toluene to determine the 14C-mass balances and with [methyl-13C]toluene to follow the flow of carbon from toluene into biomarker fatty acids. An aliquot was used to establish an aquifer-derived toluene-degrading sulfate-reducing consortium, which grew well in liquid medium. Whole-cell hybridization using 16S rRNA-targeted oligonucleotide probes specific for different phylogenetic levels within the sulfate-reducing bacteria was applied in order to characterize the sulfate-reducing populations in the original sediment, the aquifer microcosms, and the aquifer-derived consortium. In the aquifer microcosms, the 14C quantification revealed that 61.6% of the [methyl-14C]toluene was mineralized and 2.7% was assimilated. Following [methyl-13C]toluene depletion (〈1 μM), the highest 13C-enrichment was found in PLFA 16:1ω5c. In addition, biomarker fatty acids characteristic for the genera Desulfobacter and Desulfobacula (cy17:0 and 10Me16:0) were also 13C-enriched, contrary to those of other sulfate-reducing genera, e.g. Desulfovibrio and Synthrophobacter (i17:1ω7c), Desulfobulbus and Desulforhabdus (15:1ω6c and 17:1ω6c). Although hybridization detection rates remained low, indicating low bacterial activities, 43% (aquifer sediment) and 30% (aquifer microcosm) of the total active bacteria belonged to the Desulfobacteriaceae thus supporting the PLFA-based results. Desulfobacter-species (42%), which belong to the Desulfobacteriaceae, dominated the community of the consortium. Our study showed that carbon stable isotope analysis in combination with whole-cell hybridization could link toluene degradation in aquifer microcosms to the metabolic activity of the Desulfobacter-like populations. These populations could play an important role in the clean up of aromatic PHC-contaminated aquifers.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1111/j.1574-6941.2001.tb00890.x

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14

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Anaerobic degradation of toluene by pure cultures of denitrifying bacteria (1991)

Schocher, Riet J. ; Seyfried, Birgit ; Vazquez, Francisco ; [et al.]

Springer

Archives of microbiology 157 (1991), S. 7-12

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ISSN: 1432-072X

Keywords: Pseudomonas spp. ; Toluene ; Aromatic hydrocarbons ; Anaerobic degradation

Source: Springer Online Journal Archives 1860-2000

Topics: Biology

Notes: Abstract Several denitrifying Pseudomonas spp., isolated with various aromatic compounds, were tested for the ability to degrade toluene in the absence of molecular oxygen. Four out of seven strains were able to degrade toluene in the presence of N2O. More than 50% of the 14C from ring-labelled toluene was released as CO2, and up to 37% was assimilated into cell material. Furthermore it was demonstrated for two strains that they were able to grow on toluene as the sole carbon and energy source in the presence of N2O. Suspensions of cells pre-grown on toluene degraded toluene, benzaldehyde or benzoate without a lag phase and without accumulation of intermediates. p-Cresol, p-hydroxybenzylalcohol, p-hydroxybenzaldehyde or p-hydroxybenzoate was degraded much slower or only after distinct lag times. In the presence of fluoroacetate [14C]toluene was transformed to [14C]benzoate, which suggests that anaerobic toluene degradation proceeds through oxidation of the methyl side chain to benzoate.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1007/BF00245327

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15

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Detection of mRNA of nprM in Bacillus megaterium ATCC 14581 grown in soil by whole-cell hybridization (1995)

Hönerlage, Wolfgang ; Hahn, Dittmar ; Zeyer, Josef

Springer

Archives of microbiology 163 (1995), S. 235-241

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ISSN: 1432-072X

Keywords: In situ detection of mRNA ; Bacillus megaterium ; Extracellular neutral protease ; nprM ; In vitro transcripts ; Whole-cell hybridization

Source: Springer Online Journal Archives 1860-2000

Topics: Biology

Notes: Abstract Transcripts of nprM, the gene encoding the major extracellular protease of Bacillus megaterium ATCC 14581, were detected by both Northern blot analysis and whole-cell hybridization with digoxigenin-labeled in vitro ranscripts throughout the exponential growth phase and the early stationary phase. In cells of the late stationary phase, only low amounts of transcripts were observed with the two techniques. No transcripts could be detected in spores. In soil the presence of mRNA of nprM could be demonstrated by whole-cell hybridization in growing cells germinated from heat-activated spores until they reached the late transition state. No transcripts of nprM were detected in cells containing forespores. Both cells grown in pure culture and in soil had to be permeabilized with lysozyme to allow hybridization with digoxigeninlabeled probes. These results demonstrate the applicability of nucleic-acid probing techniques to localize microbial processes in soil. The approach described of detecting mRNA in fixed bacterial cells should facilitate in situ studies of gene transcription and specific activities in individual cells in heterogeneous environmental systems.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1007/BF00393374

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16

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Detection of mRNA of nprM in Bacillus megaterium ATCC 14581 grown in soil by whole-cell hybridization (1995)

Hönerlage, Wolfgang ; Hahn, D. ; Zeyer, Josef

Springer

Archives of microbiology 163 (1995), S. 235-241

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ISSN: 1432-072X

Keywords: Key words In situ detection of mRNA ; Bacillus ; megaterium ; Extracellular neutral protease ; nprM ; In vitro transcripts ; Whole-cell hybridization

Source: Springer Online Journal Archives 1860-2000

Topics: Biology

Notes: Abstract Transcripts of nprM, the gene encoding the major extracellular protease of Bacillus megaterium ATCC 14581, were detected by both Northern blot analysis and whole-cell hybridization with digoxigenin-labeled in vitro transcripts throughout the exponential growth phase and the early stationary phase. In cells of the late stationary phase, only low amounts of transcripts were observed with the two techniques. No transcripts could be detected in spores. In soil the presence of mRNA of nprM could be demonstrated by whole-cell hybridization in growing cells germinated from heat-activated spores until they reached the late transition state. No transcripts of nprM were detected in cells containing forespores. Both cells grown in pure culture and in soil had to be permeabilized with lysozyme to allow hybridization with digoxigenin-labeled probes. These results demonstrate the applicability of nucleic-acid probing techniques to localize microbial processes in soil. The approach described of detecting mRNA in fixed bacterial cells should facilitate in situ studies of gene transcription and specific activities in individual cells in heterogeneous environmental systems.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1007/BF00393374

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17

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Analysis of bacterial community structure in bulk soil by in situ hybridization (1997)

Zarda, Boris ; Hahn, D. ; Chatzinotas, Antonis ; [et al.]

Springer

Archives of microbiology 168 (1997), S. 185-192

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ISSN: 1432-072X

Keywords: Key words Fluorescent oligonucleotide probes ; Planctomycetes ; rRNA ; Whole-cell hybridization

Source: Springer Online Journal Archives 1860-2000

Topics: Biology

Notes: Abstract In situ hybridization with rRNA-targeted, fluorescent (Cy3-labeled) oligonucleotide probes was used to analyze bacterial community structure in ethanol- or paraformaldehyde-fixed bulk soil after homogenization of soil samples in 0.1% pyrophosphate by mild ultrasonic treatment. In ethanol-fixed samples 37 ± 7%, and in paraformaldehyde 41 ± 8% of the 4′, 6-diamidino-2-phenylindole(DAPI)-stained cells were detected with the bacterial probe Eub338. The yield could not be increased by enzymatic and/or chemical pretreatments known to enhance the permeability of bacterial cells for probes. However, during storage in ethanol for 7 months, the detectability of bacteria increased in both ethanol- and paraformaldehyde-fixed samples to up to 47 ± 8% due to an increase in the detection yield of members of the α-subdivision of Proteobacteria from 2 ± 1% to 10 ± 3%. Approximately half of the bacteria detected by probe Eub338 could be affiliated to major phylogenetic groups such as the α-, β-, γ-, and δ-subdivisions of Proteobacteria, gram-positive bacteria with a high G+C DNA content, bacteria of the Cytophaga-Flavobacterium cluster of the CFB phylum, and the planctomycetes. The analysis revealed that bacteria of the α- and δ-subdivision of Proteobacteria and the planctomycetes were predominant. Here, members of the α-subdivision of Proteobacteria accounted for approximately 10 ± 3% of DAPI-stained cells, which corresponded to 44 ± 16 × 108 cells (g soil, dry wt.)–1, while members of the δ-subdivision of Proteobacteria made up 4 ± 2% of DAPI-stained cells [17 ± 9 × 108 cells (g soil, dry wt.)–1]. A large population of bacteria in bulk soil was represented by the planctomycetes, which accounted for 7 ± 3% of DAPI-stained cells [32 ± 12 × 108 cells (g soil, dry wt.)–1]. The detection of planctomycetes in soil confirms previous reports on the occurrence of planctomycetes in soil and indicates a yet unknown ecological significance of this group, which to date has never been isolated from terrestrial environments.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1007/s002030050486

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18

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Intrinsic bioremediation of a petroleum hydrocarbon-contaminated aquifer and assessment of mineralization based on stable carbon isotopes (1999)

Bolliger, Christof ; Höhener, Patrick ; Hunkeler, Daniel ; [et al.]

Springer

Biodegradation 10 (1999), S. 201-217

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ISSN: 1572-9729

Source: Springer Online Journal Archives 1860-2000

Topics: Biology , Energy, Environment Protection, Nuclear Power Engineering , Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition

Notes: Abstract This study presents a stepwise concept to assess the in situ microbial mineralization of petroleum hydrocarbons (PHC) in aquifers. A new graphical method based on stable carbon isotope ratios (δ13C) was developed to verify the origin of dissolved inorganic carbon (DIC). The concept and the isotope method were applied to an aquifer in Studen, Switzerland, in which more than 34,000 liters of heating oil were accidentally released. Chemical analyses of ground water revealed that in this aquifer locally, anaerobic conditions prevailed, and that PHC mineralization was linked to the consumption of oxidants such as O2, NO 2 - , and SO 4 2- and the production of reduced species such as Fe2+, Mn2+, HSS and CH4. However, alkalinity and DIC balances showed a quantitative disagreement in the link between oxidant consumption and DIC production, indicating that chemical data alone may not be a reliable assessment tool. δ13C ratios in DIC have been used before for bioremediation assessment, but results were reported to be negatively influenced by methanogenesis. Using the new graphical method to display δ13C data, it was possible to identify anomalies found in methanogenic monitoring wells. It could be shown that 88% of the DIC produced in the contaminated aquifer originated from microbial PHC mineralization. Thus, the new graphical method to display δ13C ratios appears to be a useful tool for the assessment of microbial hydrocarbon mineralization in a complex environment.

Type of Medium: Electronic Resource

URL: http://dx.doi.org/10.1023/A:1008375213687

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