2003 RESEARCH PROJECTS Biotransformation PROJECT: Anaerobic U(IV) Bio-oxidation and the Resultant Remobilization of Uranium in Contaminated Sediments PRINCIPAL INVESTIGATOR: John Coates Biotransformation A proposed strategy for the remediation of uranium (U) contaminated sites is based on immobilizing U by reducing the oxidized soluble hexavalent U [U(VI)], to form a reduced insoluble end product, U(IV). Due to the use of nitric acid in the processing of nuclear fuels, nitrate is often a co-contaminant found in many of the environments contaminated with uranium. Recent studies indicate that nitrate inhibits U(VI) reduction in sediment slurries. However, the mechanism responsible for the apparent inhibition of U(VI) reduction is unknown, i.e. preferential utilization of nitrate as an electron acceptor, direct biological oxidation of U(IV) coupled to nitrate reduction, and/or abiotic oxidation by intermediates of nitrate reduction. A recent pure culture study of a model U(VI)-reducing bacterium, Geobacter metallireducens , indicated that this organism was alternatively capable of coupling the oxidation of U(IV) to the reduction of nitrate. The oxidation of U(IV) coupled to the reduction of nitrate at circumneutral pH would yield enough energy to support growth (DG∞’=-384.8kJ/mole). Recent studies examining a similar anaerobic bio-oxidative process, nitrate-dependent Fe(II) oxidation, have suggested that this microbial process plays crucial role in iron and nitrate geochemistry. The same relationship may exist between U(IV) and nitrate. Co-contamination with U and nitrate or subsequent transport of nitrate into reduced zones could result in the bio-oxidation of U(IV) coupled to NO3- reduction potentially impeding or reversing remediation efforts based on a reductive remediation strategy leading to a remobilization of immobilized uranium. Therefore to predictably model remediation efforts, it is essential to understand the potential for bio-oxidation of U(IV) and the potential impact of this microbial process on the long term sequestration of U in the environment. The objective of the current study is to investigate the microbiology of anaerobic nitrate-dependent U(IV) oxidation in sedimentary environments, to identify the ubiquity and diversity of organisms responsible for this metabolism, and to determine the environmental parameters that affect their ability to mediate U(IV) bio-oxidation. PROJECT: Influence of Mass Transfer on Bioavailability and Kinetic Rate of Uranium(VI) Biotransformation PRINCIPAL INVESTIGATOR: Chongxuan Liu Biotransformation The bioavailability and kinetic rate of bioreduction of metals and radionuclides associated with intraparticle regions of porous media will be constrained by mass transfer processes in long-term contaminated sediments. Recent characterization of U(VI) speciation and physical location in 30-year contaminated Hanford Site sediments demonstrated that U(VI) primarily resides as a U(VI)-silicate microprecipitate in small fractures within sediment particle grains exhibiting pore sizes of a few microns or less. This research will: 1) identify and characterize biogeochemical strategies for accessing intraparticle U(VI) by representative dissimilatory metal reducing bacteria (Shewanella and Geobacter); 2) evaluate the influence of mass transfer on U(VI) bioavailability, microbiologic reduction rate and location, and long-term stability in contaminated sediments; and 3) develop overall kinetic models of the involved processes supported by fundamental experiments. The proposed research will utilize DOE pristine and U(VI) contaminated sediments (Hanford, Oak Ridge), and synthetic U(VI)-bearing mineral fragments and silica nanoparticle aggregates for controlled process identification. Carefully formulated experiments will determine: 1) the influence of U(VI) diffusion out of the particles and diffusion of biogenic electron transducing compounds into the particles on U(VI) bioreduction rate and extent and 2) the speciation and physical location of U(IV)/U(VI) and their influences on the reoxidation rate and long-term stability of precipitated U(IV). Spectroscopic and microscopic techniques will be used to determine intraparticle porosity, diffusivity, and tortuosity, and characterize microscopic mass transfer properties and models. The microscopic and macroscopic measurements will define the linkage of mass transfer models with biokinetic and uranium equilibrium/kinetic reactions.   PROJECT: Towards a More Complete Picture: Dissimilatory Metal Reduction by Anaeromyxobacter Species PRINCIPAL INVESTIGATOR: Frank E. Loeffler Biotransformation Although substantial knowledge accrued over the past decade on metal-reducing microbial populations, our understanding of the key players involved in metal reduction at contaminated sites is incomplete. Recent findings suggest that Anaeromyxobacter populations play relevant roles in toxic metal reduction and immobilization at contaminated DOE sites. This research effort will characterize Anaeromyxobacter isolates in hand, and assess their contribution towards metal detoxification and plume stabilization under environmentally relevant conditions. Further, the distribution, abundance, and diversity of Anaeromyxobacter species at uranium-impacted DOE sites will be explored using 16S rRNA gene-based approaches. The proposed research effort will enhance our understanding of the microbially-catalyzed metal reduction process, and will provide information for the rational design and operation of both intrinsic and engineered approaches for enhanced in situ bioremediation of toxic metals at contaminated DOE sites. This research will be conducted as a collaborative project at the Georgia Institute of Technology (Dr. F. Loeffler) and the University of Illinois (Dr. R. Sanford).   PROJECT: Biotransformations Involved in Sustained Reductive Removal of Uranium in Contaminated Aquifers PRINCIPAL INVESTIGATOR: Derek Lovley  Biotransformation Our previously funded NABIR Biotransformations research conclusively demonstrated that microbial U(VI) reduction in uranium-contaminated aquifers can be stimulated with the simple addition of acetate and that stimulating this process can effectively remove uranium from contaminated groundwater. A field experiment confirmed the results of the laboratory sediment incubations, but also revealed that long-term injection of acetate may deplete Fe(III) oxides from the site of acetate injection and promote the growth of acetate-oxidizing sulfate reducers, which appeared to be less effective than Geobacter species in reducing U(VI). These results from the field experiment demonstrate a need for further study of subsurface biotransformations in order to optimize the long-term in situ uranium bioremediation process. Therefore, the objective of this research is to evaluate previously unconsidered aspects of the competition for electron donors near injection wells during in situ bioremediation, the implications of this for U(VI) reduction, and strategies for manipulating biotransformations to promote long-term U(VI) reduction. Studies in which sediments and groundwater from uranium-contaminated sites will be incubated in flow-through columns, as well as field studies, will examine the effect of providing different organic electron donors at various concentrations on long-term U(VI) reduction. It is hypothesized that addition of acetate in concentrations sufficient to consume all of the sulfate near the point of injection will provide enough acetate to maintain an active, downgradient population of Geobacter species, which will continue to remove U(VI) from the groundwater. One alternative strategy is to diminish the competitiveness of sulfate reducers near the point of injection by increasing Fe(III) concentrations in the sediments. Another strategy is to add lactate, instead of acetate, because lactate may stimulate the growth of lactate-oxidizing sulfate-reducing microorganisms which can effectively reduce U(VI) as well as provide acetate for Geobacter species. The potential for dissimilatory metal-reducing microorganisms to reduce U(VI) adsorbed onto sediments will also be examined as this is significant pool of U(VI) at many subsurface sites. Furthermore, the kinetics of U(IV) oxidation with nitrate and oxygen once addition of electron donor is terminated will be evaluated. These studies are expected to expand the basic understanding of biotransformation processes in uranium-contaminated subsurface sediments and help identify successful strategies for long-term in situ bioremediation.   [Back to Award Recipients Page]