Dataset Information

Environmental whole-genome amplification to access microbial populations in contaminated sediments.

ABSTRACT: Low-biomass samples from nitrate and heavy metal contaminated soils yield DNA amounts that have limited use for direct, native analysis and screening. Multiple displacement amplification (MDA) using phi29 DNA polymerase was used to amplify whole genomes from environmental, contaminated, subsurface sediments. By first amplifying the genomic DNA (gDNA), biodiversity analysis and gDNA library construction of microbes found in contaminated soils were made possible. The MDA method was validated by analyzing amplified genome coverage from approximately five Escherichia coli cells, resulting in 99.2% genome coverage. The method was further validated by confirming overall representative species coverage and also an amplification bias when amplifying from a mix of eight known bacterial strains. We extracted DNA from samples with extremely low cell densities from a U.S. Department of Energy contaminated site. After amplification, small-subunit rRNA analysis revealed relatively even distribution of species across several major phyla. Clone libraries were constructed from the amplified gDNA, and a small subset of clones was used for shotgun sequencing. BLAST analysis of the library clone sequences showed that 64.9% of the sequences had significant similarities to known proteins, and "clusters of orthologous groups" (COG) analysis revealed that more than half of the sequences from each library contained sequence similarity to known proteins. The libraries can be readily screened for native genes or any target of interest. Whole-genome amplification of metagenomic DNA from very minute microbial sources, while introducing an amplification bias, will allow access to genomic information that was not previously accessible. The reported SSU rRNA sequences and library clone end sequences are listed with their respective GenBank accession numbers, DQ 404590 to DQ 404652, DQ 404654 to DQ 404938, and DX 385314 to DX 389173.

SUBMITTER: Abulencia CB

PROVIDER: S-EPMC1472342 | biostudies-literature | 2006 May

REPOSITORIES: biostudies-literature

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Environmental whole-genome amplification to access microbial populations in contaminated sediments.

Abulencia Carl B CB Wyborski Denise L DL Garcia Joseph A JA Podar Mircea M Chen Wenqiong W Chang Sherman H SH Chang Hwai W HW Watson David D Brodie Eoin L EL Hazen Terry C TC Keller Martin M

Applied and environmental microbiology 20060501 5

Low-biomass samples from nitrate and heavy metal contaminated soils yield DNA amounts that have limited use for direct, native analysis and screening. Multiple displacement amplification (MDA) using phi29 DNA polymerase was used to amplify whole genomes from environmental, contaminated, subsurface sediments. By first amplifying the genomic DNA (gDNA), biodiversity analysis and gDNA library construction of microbes found in contaminated soils were made possible. The MDA method was validated by an ...[more]

PMID: 16672469

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Project description:Environmental management often aims at sustainability and at implementing goals such as the United Nations Sustainable Development Goals (SDGs). Several of these goals rely upon healthy sediments, implying that contaminated sediments need to be addressed in environmental management. In Sweden, the SDGs and national environmental objectives are meant to be central in environmental management. However, in environmental risk assessment (ERA) for contaminated sediments, it is not clear to assessors or authorities in Sweden which goals to consider and how. In the present study, we explore which environmental goals ERAs of contaminated sediment should consider, according to representatives from Swedish regulatory and advisory authorities, and to what extent ERAs consider those goals in practice in Sweden. The identified goals indicated that protection of aquatic ecosystems and associated services is a main priority for the authorities. The ERAs analyzed showed that most identified goals were addressed, albeit not consistently. Furthermore, risk assessors seem to put less weight on ecological aspects than authorities do. This indicates that authorities need to stress the importance of environmental aspects and provide guidance on how to address them. Several environmental goals were found to be directly relevant to contaminated sediments and ERAs can contribute to their fulfillment, given that ERA is a basis for management decisions. However, practitioners need clearer guidance on which goals to address and how. Finally, there seems to be good potential for implementation of the SDGs in environmental management in Sweden, due to existing legislation based on sustainable development and well-anchored national environmental objectives that overlap with the SDGs. Integr Environ Assess Manag 2019;00:1-12. © 2019 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals, Inc. on behalf of Society of Environmental Toxicology & Chemistry (SETAC).

Project description:BackgroundRadionuclide- and heavy metal-contaminated subsurface sediments remain a legacy of Cold War nuclear weapons research and recent nuclear power plant failures. Within such contaminated sediments, remediation activities are necessary to mitigate groundwater contamination. A promising approach makes use of extant microbial communities capable of hydrolyzing organophosphate substrates to promote mineralization of soluble contaminants within deep subsurface environments.Methodology/principal findingsUranium-contaminated sediments from the U.S. Department of Energy Oak Ridge Field Research Center (ORFRC) Area 2 site were used in slurry experiments to identify microbial communities involved in hydrolysis of 10 mM organophosphate amendments [i.e., glycerol-2-phosphate (G2P) or glycerol-3-phosphate (G3P)] in synthetic groundwater at pH 5.5 and pH 6.8. Following 36 day (G2P) and 20 day (G3P) amended treatments, maximum phosphate (PO4(3-)) concentrations of 4.8 mM and 8.9 mM were measured, respectively. Use of the PhyloChip 16S rRNA microarray identified 2,120 archaeal and bacterial taxa representing 46 phyla, 66 classes, 110 orders, and 186 families among all treatments. Measures of archaeal and bacterial richness were lowest under G2P (pH 5.5) treatments and greatest with G3P (pH 6.8) treatments. Members of the phyla Crenarchaeota, Euryarchaeota, Bacteroidetes, and Proteobacteria demonstrated the greatest enrichment in response to organophosphate amendments and the OTUs that increased in relative abundance by 2-fold or greater accounted for 9%-50% and 3%-17% of total detected Archaea and Bacteria, respectively.Conclusions/significanceThis work provided a characterization of the distinct ORFRC subsurface microbial communities that contributed to increased concentrations of extracellular phosphate via hydrolysis of organophosphate substrate amendments. Within subsurface environments that are not ideal for reductive precipitation of uranium, strategies that harness microbial phosphate metabolism to promote uranium phosphate precipitation could offer an alternative approach for in situ sequestration.

Dataset Information

Environmental whole-genome amplification to access microbial populations in contaminated sediments.

Publications

Environmental whole-genome amplification to access microbial populations in contaminated sediments.

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