Multiple sets of chemotaxis genes including three cheA homologs were identified in the genome sequence of the anaerobic bacterium Desulfovibrio vulgaris Hildenborough. Each CheA is a histidine kinase (HK) and part of a two component signal transduction system. Knock out mutants in the three cheA genes were created using single cross-over homologous recombination insertion. We studied the phenotypes of the cheA mutants in detail and discovered that ?cheA-3 has a non swarming/swimming phenotype both in the soft agar plates and Palleroni chamber assays. CheA-3 shows similarity to the Shewanella oneidensis CheA-3 and the Vibrio cholerae CheA-2 that are responsible for chemotaxis in the respective organisms. We did not find any morphological or structural differences between the three Delta cheA mutants and the wild type cells in electron microscopy. Our results from these studies are presented.

Due to increasing atmospheric CO2 concentrations causing the global energy and environmental crises, geological sequestration of carbon dioxide is now being actively considered as an attractive option to mitigate greenhouse gas emissions. One of the important strategies is to use deep unminable coal seams, for those generally contain significant quantities of coal bed methane that can be recovered by CO2 injection through enhanced coal bed natural gas production, as a method to safely store CO2. It has been well known that the adsorbing CO2 molecules introduce structural deformation, such as distortion, shrinkage, or swelling, of the adsorbent of coal organic matrix. The accurate investigations of CO2 sorption capacity as well as of adsorption behavior need to be performed under the conditions that coals deform. The U.S. Department of Energy-National Energy Technology Laboratory and Regional University Alliance are conducting carbon dioxide sorption isotherm experiments by using manometric analysis method for estimation of CO2 sorption capacity of various coal samples and are constructing a gravimetric apparatus which has a visual window cell. The gravimetric apparatus improves the accuracy of carbon dioxide sorption capacity and provides feasibility for the observation of structural deformation of coal sample while carbon dioxide molecules interact with coal …

Our goal is to use isotopic measurements to understand how contaminants are introduced to and stored in the vadose zone, and what processes control migration from the vadose zone to groundwater and then to surface water. We have been using the Hanford Site in south-central Washington as our field laboratory, and our investigations are often stimulated by observations made as part of the groundwater monitoring program and vadose zone characterization activities. Understanding the transport of contaminants at Hanford is difficult due to the presence of multiple potential sources within small areas, the long history of activities, the range of disposal methods, and the continuing evolution of the hydrological system. Observations often do not conform to simple models, and cannot be adequately understood with standard characterization approaches, even though the characterization activities are quite extensive. One of our objectives is to test the value of adding isotopic techniques to the characterization program, which has the immediate potential benefit of addressing specific remediation issues, but more importantly, it allows us to study fundamental processes at the scale and in the medium where they need to be understood. Here we focus on two recent studies at the waste management area (WMA) T-TX-TY, which …

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For the in situ reductive immobilization of U to be an acceptable strategy for the removal of that element from groundwater, the long-term stability of U(IV) must be determined. Rates of biotransformation of Fe species influence the mineralogy of the resulting products (Fredrickson et al., 2003; Senko et al., 2005), and we hypothesize that the rate of U(VI) reduction influences the mineralogy of resultant U(IV) precipitates. We hypothesize that slower rates of U(VI) reduction will yield U(IV) phases that are more resistant to reoxidation, and will therefore be more stable upon cessation of electron donor addition. U(IV) phases formed by relatively slow reduction may be more crystalline or larger in comparison to their relatively rapidly-formed counterparts (Figure 1), thus limiting the reactivity of slowly-formed U(IV) phases toward various oxidants. The physical location of U(IV) precipitates relative to bacterial cells may also limit the reactivity of biogenic U(IV) phases. In this situation, we expect that precipitation of U(IV) within the bacterial cell may protect U(IV) from reoxidation by limiting physical contact between U(IV) and oxidants (Figure 1). We assessed the effect of U(VI) reduction rate on the subsequent reoxidation of biogenic U(IV) and are currently conducting column scale studies to determine …

D. vulgaris Hildenborough has 72 response regulators. The Desulfovibrio are sulfate reducing bacteria that are important in the sulfur and carbon cycles in anoxic habitats. Its large number of two componenent systems are probably critical to its ability to sense and respond to its environment. Our goal is to map these RRs to the genes they regulate using a DNA-affinity-purification-chip (DAP-chip) protocol. First target determined usuing EMSA. A positive target was determined for as many RRs as possible using EMSA. Targets were selected based on gene proximity, regulon predictions and/or predicted sigma54 dependent promoters. qPCR was used to ensure that the target was enriched from sheared genomic DNA before proceeding to the DAP-chip.

Effective environmental management of DOE sites requires reliable prediction of reactive transport phenomena. A central issue in prediction of subsurface reactive transport is the impact of multiscale physical, chemical, and biological heterogeneity. Heterogeneity manifests itself through incomplete mixing of reactants at scales below those at which concentrations are explicitly defined (i.e., the numerical grid scale). This results in a mismatch between simulated reaction processes (formulated in terms of average concentrations) and actual processes (controlled by local concentrations). At the field scale, this results in apparent scale-dependence of model parameters and inability to utilize laboratory parameters in field models. Accordingly, most field modeling efforts are restricted to empirical estimation of model parameters by fitting to field observations, which renders extrapolation of model predictions beyond fitted conditions unreliable. The objective of this project is to develop a theoretical and computational framework for (1) connecting models of coupled reactive transport from pore-scale processes to field-scale bioremediation through a hierarchy of models that maintain crucial information from the smaller scales at the larger scales; and (2) quantifying the uncertainty that is introduced by both the upscaling process and uncertainty in physical parameters. One of the challenges of addressing scale-dependent effects of coupled processes in …

Remediation and long-term stewardship of uranium-contaminated sediments and groundwaters are critical problems at a number of DOE facilities and mining sites. Some remediation strategies based on in-situ bioreduction of U are potentially effective in significantly decreasing U concentrations in groundwaters. However, a number of basic processes require understanding in order to identify conditions more conducive to success of reduction-based U stabilization. Our current research targets several of these issues including: (1) effects of organic carbon (OC) forms and supply rates on stability of bioreduced U, (2) the roles of Fe(III)- and Mn(III,IV)-oxides as potential U oxidants in sediments, and (3) microbial community changes in relation to U redox changes. These issues were identified in our previous study on U bioreduction and reoxidation (Wan et al., 2005). Most of our studies are being conducted on historically U-contaminated sediments from Area 2 of the Field Research Center, Oak Ridge National Laboratory, in flow-through columns simulating in-situ field remediation.

A 15 h ozonation was performed on bioremediated soil to remove recalcitrant residual oil. To monitor the survival of indigenous microorganisms in the soil during in-situ chemical oxidation(ISCO) culturing and a functional genearray, GeoChip, was used to examine the functional genes and structure of the microbial community during ozonation (0h, 2h, 4h, 6h, 10hand15h). Breakthrough ozonation decreased the population of cultivable heterotrophic bacteria by about 3 orders of magnitude. The total functional gene abundance and diversity decreased during ozonation, as the number of functional genes was reduced by 48percent after 15 h. However, functional genes were evenly distributed during ozonation as judged by the Shannon-Weaver Evenness index. A sharp decrease in gene number was observed in the first 6 h of ozonation followed by a slower decrease in the next 9 h, which was consistent with microbial populations measured by a culture based method. Functional genes involved in carbon, nitrogen, phosphors and sulfur cycling, metal resistance and organic remediation were detected in all samples. Though the pattern of gene categories detected was similar for all time points, hierarchica lcluster of all functional genes and major functional categories all showed a time-serial pattern. Bacteria, archaea and fungi decreased by 96.1percent, 95.1percent …

Environmental nanoparticles are often poorly-crystalline or metastable structures, whose kinetics of formation and growth are poorly understood. Further, the sorption or growth of nanoparticles on mineral surfaces may control the mineral surface's reactivity and modify its ability to influence contaminant transport. Due to the characteristic length scale, a holistic understanding of the nucleation mechanisms and kinetics of nanoparticle formation on mineral surfaces is difficult to achieve with traditional methodology. In this work, our intent is to determine the molecular nature of nucleation on surfaces, the kinetics of surface nucleation and growth, and the effect of crystal surface topology using new synchrotron-based techniques. We have approached these objectives by: (1) combining state-of-the-art crystal-truncation rod diffraction (CTR) and grazing incidence x-ray absorption fine structure spectroscopy (GIXAS) techniques to investigate the three-dimensional molecular-scale geometry of silicate monomer sorption on the r-plane of hematite; and (2) developing a new grazing-incidence small angle x-ray scattering (GISAXS) setup at SSRL (0.08 nm{sup -1} < q < 8 nm{sup -1}) to explore the initial development of environmental nanoparticles on various mineral surfaces. This study also includes complementary techniques such as atomic force microscopy (AFM), bulk SAXS, dynamic light scattering (DLS), XRD, and TEM.

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Recent advancements in utility-scale wind turbine technology and pricing have vastly increased the potential land area where turbines can be deployed in the United States. This presentation quantifies the new developable land potential (e.g., capacity curves), visually identifies new areas for possible development (e.g., new wind resource maps), and begins to address deployment barriers to wind in new areas for modern and future turbine technology.

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Parabolic trough receivers, or heat collection elements (HCEs), absorb sunlight focused by the mirrors and transfer that thermal energy to a fluid flowing within them. Thje absorbing tube of these receivers typically operates around 400 C (752 F). HCE manufacturers prevent thermal loss from the absorbing tube to the environment by using sputtered selective Cermet coatings on the absorber and by surrounding the absorber with a glass-enclosed evacuated annulus. This work quantifies the heat loss of the Solel UVAC2 and Schott PTR70 HCEs. At 400 C, the HCEs perform similarly, losing about 400 W/m of HCE length. To put this in perspective, the incident beam radiation on a 5 m mirror aperture is about 4500 W/m, with about 75% of that energy ({approx} 3400 W/m) reaching the absorber surface. Of the 3400 W/m on the absorber, about 3000 W/m is absorbed into the working fluid while 400 W/m is lost to the environment.

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Phylogenetic analyses were done for the Shewanella strains isolated from Baltic Sea (38 strains), US DOE Hanford Uranium bioremediation site [Hanford Reach of the Columbia River (HRCR), 11 strains], Pacific Ocean and Hawaiian sediments (8 strains), and strains from other resources (16 strains) with three out group strains, Rhodopseudomonas palustris, Clostridium cellulolyticum, and Thermoanaerobacter ethanolicus X514, using DNA relatedness derived from WCGA-based DNA-DNA hybridizations, sequence similarities of 16S rRNA gene and gyrB gene, and sequence similarities of 6 loci of Shewanella genome selected from a shared gene list of the Shewanella strains with whole genome sequenced based on the average nucleotide identity of them (ANI). The phylogenetic trees based on 16S rRNA and gyrB gene sequences, and DNA relatedness derived from WCGA hybridizations of the tested Shewanella strains share exactly the same sub-clusters with very few exceptions, in which the strains were basically grouped by species. However, the phylogenetic analysis based on DNA relatedness derived from WCGA hybridizations dramatically increased the differentiation resolution at species and strains level within Shewanella genus. When the tree based on DNA relatedness derived from WCGA hybridizations was compared to the tree based on the combined sequences of the selected functional genes (6 loci), we …

The overall objective of this project is to examine the activity of nonspecific phosphohydrolases present in naturally occurring subsurface microorganisms for the purpose of promoting the immobilization of radionuclides through the production of uranium [U(VI)] phosphate precipitates. Specifically, we hypothesize that the precipitation of U(VI) phosphate minerals may be promoted through the microbial release and/or accumulation of PO4 3- as a means to detoxify radionuclides and heavy metals. An experimental approach was designed to determine the extent of phosphatase activity in bacteria previously isolated from contaminated subsurface soils collected at the ERSP Field Research Center (FRC) in Oak Ridge, TN. Screening of 135 metal resistant isolates for phosphatase activity indicated the majority (75 of 135) exhibited a phosphatase-positive phenotype. During this phase of the project, a PCR based approach has also been designed to assay FRC isolates for the presence of one or more classes of the characterized non-specific acid phophastase (NSAP) genes likely to be involved in promoting U(VI) precipitation. Testing of a subset of Pb resistant (Pbr) Arthrobacter, Bacillus and Rahnella strains indicated 4 of the 9 Pbr isolates exhibited phosphatase phenotypes suggestive of the ability to bioprecipitate U(VI). Two FRC strains, a Rahnella sp. strain Y9602 and …

Dissolved inorganic carbon (DIC) is commonly measured in water and is an important parameter for understanding carbonate equilibrium, carbon cycling, and water-rock interaction. While accurate measurements can be made in the analytical laboratory, we have developed a rapid, portable technique that can be used to obtain accurate and precise data in the field as well.

in this study, we carried out large-scale comparative genomics analysis of regulatory interactions in Desulfovibrio vulgaris and 12 related genomes from Desulfovibrionales order using our recently developed web server RegPredict (http://regpredict.lbl.gov). An overall reference collection of 26 Desulfovibrionales regulogs can be accessed through RegPrecise database (http://regpredict.lbl.gov).

Reconstruction of transcriptional regulatory networks is one of the major challenges facing the bioinformatics community in view of constantly growing number of complete genomes. The comparative genomics approach has been successfully used for the analysis of the transcriptional regulation of many metabolic systems in various bacteria taxa. The key step in this approach is given a position weight matrix, find an optimal threshold for the search of potential binding sites in genomes. In our previous work we proposed an approach for automatic selection of TFBS score threshold coupled with inference of regulon content. In this study we developed two modifications of this approach providing two additional levels of sensitivity.

The role of tetraheme cytochrome c3 (CycA) in the metabolism of the sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough (DvH) was investigated by deletion of the cycA gene using a marker-exchange deletion strategy. A highly abundant periplasmic cytochrome, CycA has the important function of transferring electrons from periplasmic hydrogenases (Hyd, Hyn, Hys) to transmembrane complexes which transport the electrons to the cytoplasm where sulfate is reduced. Previous studies have indicated that during its interaction with periplasmic hydrogenases, CycA is also involved in the reduction of toxic metals. Growth of the cycA mutant strain on lactate as the electron donor and sulfate as the terminal electron acceptor showed that, despite its abundance, CycA is not essential for DvH growth. However, the rate of growth of the mutant strain was significantly lower, and the extent of growth less, than rates and extents of growth of the wild type and complement strains on lactate/sulfate medium. This indicates that a portion of the electrons generated from cytoplasmic lactate oxidation are transported by CycA for energy production, possibly in a hydrogen cycling mechanism employed to generate ATP. Failure of the mutant strain to grow on either formate or H2, with sulfate or sulfite as electron acceptors, further …

Plutonium has a long half-life (2.4 x 104 years) and is of concern because of its chemical and radiological toxicity, high-energy alpha radioactive decay. A full understanding of its speciation and interactions with environmental processes is required in order to predict, contain, or remediate contaminated sites. Under aerobic conditions Pu is sparingly soluble, existing primarily in its tetravalent oxidation state. To the extent that pentavalent and hexavalent complexes and small colloidal species form they will increase the solubility and resultant mobility from contamination sources. There is evidence that in both marine environments and brines substantial fractions of the plutonium in solution is present as hexavalent plutonyl, PuO2 2+.