@article {3145, title = {Enrichment of specific protozoan populations during in situ bioremediation of uranium-contaminated groundwater.}, journal = {ISME J}, volume = {7}, year = {2013}, month = {2013 Jul}, pages = {1286-98}, abstract = {

The importance of bacteria in the anaerobic bioremediation of groundwater polluted with organic and/or metal contaminants is well recognized and in some instances so well understood that modeling of the in situ metabolic activity of the relevant subsurface microorganisms in response to changes in subsurface geochemistry is feasible. However, a potentially significant factor influencing bacterial growth and activity in the subsurface that has not been adequately addressed is protozoan predation of the microorganisms responsible for bioremediation. In field experiments at a uranium-contaminated aquifer located in Rifle, CO, USA, acetate amendments initially promoted the growth of metal-reducing Geobacter species, followed by the growth of sulfate reducers, as observed previously. Analysis of 18S rRNA gene sequences revealed a broad diversity of sequences closely related to known bacteriovorous protozoa in the groundwater before the addition of acetate. The bloom of Geobacter species was accompanied by a specific enrichment of sequences most closely related to the ameboid flagellate, Breviata anathema, which at their peak accounted for over 80\% of the sequences recovered. The abundance of Geobacter species declined following the rapid emergence of B. anathema. The subsequent growth of sulfate-reducing Peptococcaceae was accompanied by another specific enrichment of protozoa, but with sequences most similar to diplomonadid flagellates from the family Hexamitidae, which accounted for up to 100\% of the sequences recovered during this phase of the bioremediation. These results suggest a prey-predator response with specific protozoa responding to increased availability of preferred prey bacteria. Thus, quantifying the influence of protozoan predation on the growth, activity and composition of the subsurface bacterial community is essential for predictive modeling of in situ uranium bioremediation strategies.

}, keywords = {Acetates, Biodegradation, Environmental, Eukaryota, Geobacter, Groundwater, Molecular Sequence Data, Oxidation-Reduction, Phylogeny, RNA, Ribosomal, 16S, RNA, Ribosomal, 18S, Uranium}, issn = {1751-7370}, doi = {10.1038/ismej.2013.20}, author = {Holmes, Dawn E and Giloteaux, Ludovic and Williams, Kenneth H and Wrighton, Kelly C and Wilkins, Michael J and Thompson, Courtney A and Roper, Thomas J and Long, Philip E and Lovley, Derek R} } @article {3138, title = {Field evidence of selenium bioreduction in a uranium-contaminated aquifer.}, journal = {Environ Microbiol Rep}, volume = {5}, year = {2013}, month = {2013 Jun}, pages = {444-52}, abstract = {

Removal of selenium from groundwater was documented during injection of acetate into a uranium-contaminated aquifer near Rifle, Colorado (USA). Bioreduction of aqueous selenium to its elemental form (Se0) concentrated it within mineralized biofilms affixed to tubing used to circulate acetate-amended groundwater. Scanning and transmission electron microscopy revealed close association between Se0 precipitates and cell surfaces, with Se0 aggregates having a diameter of 50-60 nm. Accumulation of Se0 within biofilms occurred over a three-week interval at a rate of c. 9 mg Se0 m(-2) tubing day(-1). Removal was inferred to result from the activity of a mixed microbial community within the biofilms capable of coupling acetate oxidation to the reduction of oxygen, nitrate and selenate. Phylogenetic analysis of the biofilm revealed a community dominated by strains of Dechloromonas sp. and Thauera sp., with isolates exhibiting genetic similarity to the latter known to reduce selenate to Se0. Enrichment cultures of selenate-respiring microorganisms were readily established using Rifle site groundwater and acetate, with cultures dominated by strains closely related to D. aromatica (96-99\% similarity). Predominance of Dechloromonas sp. in recovered biofilms and enrichments suggests this microorganism may play a role in the removal of selenium oxyanions present in Se-impacted groundwaters and sediments.

}, keywords = {Acetates, Betaproteobacteria, Biodegradation, Environmental, Biofilms, Colorado, Groundwater, Humans, Microbial Consortia, Oxidation-Reduction, Oxygen, Phylogeny, RNA, Ribosomal, 16S, Selenic Acid, Selenium, Selenium Compounds, Thauera, Uranium, Water Pollutants, Chemical}, issn = {1758-2229}, doi = {10.1111/1758-2229.12032}, author = {Williams, Kenneth H and Wilkins, Michael J and N{\textquoteright}Guessan, A Lucie and Arey, Bruce and Dodova, Elena and Dohnalkova, Alice and Holmes, Dawn and Lovley, Derek R and Long, Philip E} } @article {3143, title = {Fluctuations in species-level protein expression occur during element and nutrient cycling in the subsurface.}, journal = {PLoS One}, volume = {8}, year = {2013}, month = {2013}, pages = {e57819}, abstract = {

While microbial activities in environmental systems play a key role in the utilization and cycling of essential elements and compounds, microbial activity and growth frequently fluctuates in response to environmental stimuli and perturbations. To investigate these fluctuations within a saturated aquifer system, we monitored a carbon-stimulated in situ Geobacter population while iron reduction was occurring, using 16S rRNA abundances and high-resolution tandem mass spectrometry proteome measurements. Following carbon amendment, 16S rRNA analysis of temporally separated samples revealed the rapid enrichment of Geobacter-like environmental strains with strong similarity to G. bemidjiensis. Tandem mass spectrometry proteomics measurements suggest high carbon flux through Geobacter respiratory pathways, and the synthesis of anapleurotic four carbon compounds from acetyl-CoA via pyruvate ferredoxin oxidoreductase activity. Across a 40-day period where Fe(III) reduction was occurring, fluctuations in protein expression reflected changes in anabolic versus catabolic reactions, with increased levels of biosynthesis occurring soon after acetate arrival in the aquifer. In addition, localized shifts in nutrient limitation were inferred based on expression of nitrogenase enzymes and phosphate uptake proteins. These temporal data offer the first example of differing microbial protein expression associated with changing geochemical conditions in a subsurface environment.

}, keywords = {Biomass, Carbon, Environment, Gene Expression Regulation, Bacterial, Geobacter, Groundwater, Humic Substances, Iron, Oxidation-Reduction, Phosphates, Plankton, Proteomics, RNA, Ribosomal, 16S, Tandem Mass Spectrometry, Uranium, Vanadium, Water Microbiology}, issn = {1932-6203}, doi = {10.1371/journal.pone.0057819}, author = {Wilkins, Michael J and Wrighton, Kelly C and Nicora, Carrie D and Williams, Kenneth H and McCue, Lee Ann and Handley, Kim M and Miller, Chris S and Giloteaux, Ludovic and Montgomery, Alison P and Lovley, Derek R and Banfield, Jillian F and Long, Philip E and Lipton, Mary S} } @article {413, title = {Microbial functional gene diversity with a shift of subsurface redox conditions during In Situ uranium reduction.}, journal = {Appl Environ Microbiol}, volume = {78}, year = {2012}, month = {2012 Apr}, pages = {2966-72}, abstract = {To better understand the microbial functional diversity changes with subsurface redox conditions during in situ uranium bioremediation, key functional genes were studied with GeoChip, a comprehensive functional gene microarray, in field experiments at a uranium mill tailings remedial action (UMTRA) site (Rifle, CO). The results indicated that functional microbial communities altered with a shift in the dominant metabolic process, as documented by hierarchical cluster and ordination analyses of all detected functional genes. The abundance of dsrAB genes (dissimilatory sulfite reductase genes) and methane generation-related mcr genes (methyl coenzyme M reductase coding genes) increased when redox conditions shifted from Fe-reducing to sulfate-reducing conditions. The cytochrome genes detected were primarily from Geobacter sp. and decreased with lower subsurface redox conditions. Statistical analysis of environmental parameters and functional genes indicated that acetate, U(VI), and redox potential (E(h)) were the most significant geochemical variables linked to microbial functional gene structures, and changes in microbial functional diversity were strongly related to the dominant terminal electron-accepting process following acetate addition. The study indicates that the microbial functional genes clearly reflect the in situ redox conditions and the dominant microbial processes, which in turn influence uranium bioreduction. Microbial functional genes thus could be very useful for tracking microbial community structure and dynamics during bioremediation.}, keywords = {Biodegradation, Environmental, Biota, Environmental Microbiology, Environmental Pollutants, Genetic Variation, Microarray Analysis, Oxidation-Reduction, Uranium}, issn = {1098-5336}, doi = {10.1128/AEM.06528-11}, author = {Liang, Yuting and Van Nostrand, Joy D and N{\textquoteright}guessan, Lucie A and Peacock, Aaron D and Deng, Ye and Long, Philip E and Resch, C Tom and Wu, Liyou and He, Zhili and Li, Guanghe and Hazen, Terry C and Lovley, Derek R and Zhou, Jizhong} } @article {457, title = {Proteogenomic monitoring of Geobacter physiology during stimulated uranium bioremediation.}, journal = {Appl Environ Microbiol}, volume = {75}, year = {2009}, month = {2009 Oct}, pages = {6591-9}, abstract = {Implementation of uranium bioremediation requires methods for monitoring the membership and activities of the subsurface microbial communities that are responsible for reduction of soluble U(VI) to insoluble U(IV). Here, we report a proteomics-based approach for simultaneously documenting the strain membership and microbial physiology of the dominant Geobacter community members during in situ acetate amendment of the U-contaminated Rifle, CO, aquifer. Three planktonic Geobacter-dominated samples were obtained from two wells down-gradient of acetate addition. Over 2,500 proteins from each of these samples were identified by matching liquid chromatography-tandem mass spectrometry spectra to peptides predicted from seven isolate Geobacter genomes. Genome-specific peptides indicate early proliferation of multiple M21 and Geobacter bemidjiensis-like strains and later possible emergence of M21 and G. bemidjiensis-like strains more closely related to Geobacter lovleyi. Throughout biostimulation, the proteome is dominated by enzymes that convert acetate to acetyl-coenzyme A and pyruvate for central metabolism, while abundant peptides matching tricarboxylic acid cycle proteins and ATP synthase subunits were also detected, indicating the importance of energy generation during the period of rapid growth following the start of biostimulation. Evolving Geobacter strain composition may be linked to changes in protein abundance over the course of biostimulation and may reflect changes in metabolic functioning. Thus, metagenomics-independent community proteogenomics can be used to diagnose the status of the subsurface consortia upon which remediation biotechnology relies.}, keywords = {Amino Acid Sequence, Bacterial Proteins, Biodegradation, Environmental, Genomics, Geobacter, Molecular Sequence Data, Oxidation-Reduction, Peptide Mapping, Plankton, Proteomics, Uranium, Water Microbiology, Water Pollutants, Radioactive}, issn = {1098-5336}, doi = {10.1128/AEM.01064-09}, author = {Wilkins, Michael J and VerBerkmoes, Nathan C and Williams, Kenneth H and Callister, Stephen J and Mouser, Paula J and Elifantz, Hila and N{\textquoteright}guessan, Lucie A and Thomas, Brian C and Nicora, Carrie D and Shah, Manesh B and Abraham, Paul and Lipton, Mary S and Lovley, Derek R and Hettich, Robert L and Long, Philip E and Banfield, Jillian F} } @article {482, title = {Sustained removal of uranium from contaminated groundwater following stimulation of dissimilatory metal reduction.}, journal = {Environ Sci Technol}, volume = {42}, year = {2008}, month = {2008 Apr 15}, pages = {2999-3004}, abstract = {Previous field studies on in situ bioremediation of uranium-contaminated groundwater in an aquifer in Rifle, Colorado identified two distinct phases following the addition of acetate to stimulate microbial respiration. In phase I, Geobacter species are the predominant organisms, Fe(III) is reduced, and microbial reduction of soluble U(VI) to insoluble U(IV) removes uranium from the groundwater. In phase II, Fe(III) is depleted, sulfate is reduced, and sulfate-reducing bacteria predominate. Long-term monitoring revealed an unexpected third phase during which U(VI) removal continues even after acetate additions are stopped. All three of these phases were successfully reproduced in flow-through sediment columns. When sediments from the third phase were heat sterilized, the capacity for U(VI) removal was lost. In the live sediments U(VI) removed from the groundwater was recovered as U(VI) in the sediments. This contrasts to the recovery of U(IV) in sediments resulting from the reduction of U(VI) to U(IV) during the Fe(III) reduction phase in acetate-amended sediments. Analysis of 16S rRNA gene sequences in the sediments in which U(VI) was being adsorbed indicated that members of the Firmicutes were the predominant organisms whereas no Firmicutes sequences were detected in background sediments which did not have the capacity to sorb U(VI), suggesting that the U(VI) adsorption might be due to the presence of these living organisms or at least their intact cell components. This unexpected enhanced adsorption of U(VI) onto sediments following the stimulation of microbial growth in the subsurface may potentially enhance the cost effectiveness of in situ uranium bioremediation.}, keywords = {Acetates, Adsorption, Bacteria, Colorado, Geologic Sediments, Oxidation-Reduction, RNA, Ribosomal, 16S, Sulfates, Uranium, Water Pollutants, Radioactive, Water Supply}, issn = {0013-936X}, author = {N{\textquoteright}guessan, Lucie A and Vrionis, Helen A and Resch, Charles T and Long, Philip E and Lovley, Derek R} } @article {493, title = {Subsurface clade of Geobacteraceae that predominates in a diversity of Fe(III)-reducing subsurface environments.}, journal = {ISME J}, volume = {1}, year = {2007}, month = {2007 Dec}, pages = {663-77}, abstract = {There are distinct differences in the physiology of Geobacter species available in pure culture. Therefore, to understand the ecology of Geobacter species in subsurface environments, it is important to know which species predominate. Clone libraries were assembled with 16S rRNA genes and transcripts amplified from three subsurface environments in which Geobacter species are known to be important members of the microbial community: (1) a uranium-contaminated aquifer located in Rifle, CO, USA undergoing in situ bioremediation; (2) an acetate-impacted aquifer that serves as an analog for the long-term acetate amendments proposed for in situ uranium bioremediation and (3) a petroleum-contaminated aquifer in which Geobacter species play a role in the oxidation of aromatic hydrocarbons coupled with the reduction of Fe(III). The majority of Geobacteraceae 16S rRNA sequences found in these environments clustered in a phylogenetically coherent subsurface clade, which also contains a number of Geobacter species isolated from subsurface environments. Concatamers constructed with 43 Geobacter genes amplified from these sites also clustered within this subsurface clade. 16S rRNA transcript and gene sequences in the sediments and groundwater at the Rifle site were highly similar, suggesting that sampling groundwater via monitoring wells can recover the most active Geobacter species. These results suggest that further study of Geobacter species in the subsurface clade is necessary to accurately model the behavior of Geobacter species during subsurface bioremediation of metal and organic contaminants.}, keywords = {Biodegradation, Environmental, Ecosystem, Ferric Compounds, Geobacter, Hydrocarbons, Aromatic, Molecular Sequence Data, Oxidation-Reduction, Petroleum, Phylogeny, Polymerase Chain Reaction, RNA, Ribosomal, 16S, Sequence Analysis, DNA, Uranium}, issn = {1751-7362}, doi = {10.1038/ismej.2007.85}, author = {Holmes, Dawn E and O{\textquoteright}Neil, Regina A and Vrionis, Helen A and N{\textquoteright}guessan, Lucie A and Ortiz-Bernad, Irene and Larrahondo, Maria J and Adams, Lorrie A and Ward, Joy A and Nicoll, Julie S and Nevin, Kelly P and Chavan, Milind A and Johnson, Jessica P and Long, Philip E and Lovley, Derek R} } @article {563, title = {Stimulating the in situ activity of Geobacter species to remove uranium from the groundwater of a uranium-contaminated aquifer.}, journal = {Appl Environ Microbiol}, volume = {69}, year = {2003}, month = {2003 Oct}, pages = {5884-91}, abstract = {The potential for removing uranium from contaminated groundwater by stimulating the in situ activity of dissimilatory metal-reducing microorganisms was evaluated in a uranium-contaminated aquifer located in Rifle, Colo. Acetate (1 to 3 mM) was injected into the subsurface over a 3-month period via an injection gallery composed of 20 injection wells, which was installed upgradient from a series of 15 monitoring wells. U(VI) concentrations decreased in as little as 9 days after acetate injection was initiated, and within 50 days uranium had declined below the prescribed treatment level of 0.18 micro M in some of the monitoring wells. Analysis of 16S ribosomal DNA (rDNA) sequences and phospholipid fatty acid profiles demonstrated that the initial loss of uranium from the groundwater was associated with an enrichment of Geobacter species in the treatment zone. Fe(II) in the groundwater also increased during this period, suggesting that U(VI) reduction was coincident with Fe(III) reduction. As the acetate injection continued over 50 days there was a loss of sulfate from the groundwater and an accumulation of sulfide and the composition of the microbial community changed. Organisms with 16S rDNA sequences most closely related to those of sulfate reducers became predominant, and Geobacter species became a minor component of the community. This apparent switch from Fe(III) reduction to sulfate reduction as the terminal electron accepting process for the oxidation of the injected acetate was associated with an increase in uranium concentration in the groundwater. These results demonstrate that in situ bioremediation of uranium-contaminated groundwater is feasible but suggest that the strategy should be optimized to better maintain long-term activity of Geobacter species.}, keywords = {Acetates, DNA, Ribosomal, Ecosystem, Fatty Acids, Ferric Compounds, Fresh Water, Geobacter, Mining, Oxidation-Reduction, Phospholipids, RNA, Ribosomal, 16S, Sulfates, Uranium, Water Pollution, Chemical}, issn = {0099-2240}, author = {Anderson, Robert T and Vrionis, Helen A and Ortiz-Bernad, Irene and Resch, Charles T and Long, Philip E and Dayvault, Richard and Karp, Ken and Marutzky, Sam and Metzler, Donald R and Peacock, Aaron and White, David C and Lowe, Mary and Lovley, Derek R} }