Sustained removal of uranium from contaminated groundwater following stimulation of dissimilatory metal reduction

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.     

In-situ evidence for uranium immobilization and remobilization

The in-situ microbial reduction and immobilization of uranium was assessed as a means of preventing the migration of this element in the terrestrial subsurface. Uranium immobilization (putatively identified as reduction) and microbial respiratory activities were evaluated in the presence of exogenous electron donors and acceptors with field push-pull tests using wells installed in an anoxic aquifer contaminated with landfill leachate. Uranium(VI) amended at 1.5 microM was reduced to less than 1 nM in groundwater in less than 8 d during all field experiments. Amendments of 0.5 mM sulfate or 5 mM nitrate slowed U(VI) immobilization and allowed for the recovery of 10% and 54% of the injected element, respectively, as compared to 4% in the unamended treatment. Laboratory incubations confirmed the field tests and showed that the majority of the U(VI) immobilized was due to microbial reduction. In these tests, nitrate treatment (7.5 mM) inhibited U(VI) reduction, and nitrite was transiently produced. Further push-pull tests were performed in which either 1 or 5 mM nitrate was added with 1.0 uM U(VI) to sediments that already contained immobilized uranium. After an initial loss of the amendments, the concentration of soluble U(VI) increased and eventually exceeded the injected concentration, indicating that previously immobilized uranium was remobilized as nitrate was reduced. Laboratory experiments using heat-inactivated sediment slurries suggested that the intermediates of dissimilatory nitrate reduction (denitrification or dissimilatory nitrate reduction to ammonia), nitrite, nitrous oxide, and nitric oxide were all capable of oxidizing and mobilizing U(IV). These findings indicate that in-situ subsurface U(VI) immobilization can be expected to take place under anaerobic conditions, but the permanence of the approach can be impaired by disimilatory nitrate reduction intermediates that can mobilize previously reduced uranium.     

Remediation and recovery of uranium from contaminated subsurface environments with electrodes

Previous studies have demonstrated that Geobacter species can effectively remove uranium from contaminated groundwater by reducing soluble U(VI) to the relatively insoluble U(IV) with organic compounds serving as the electron donor. Studies were conducted to determine whether electrodes might serve as an alternative electron donor for U(VI) reduction by a pure culture of Geobacter sulfurreducens and microorganisms in uranium-contaminated sediments. Electrodes poised at -500 mV (vs a Ag/AgCl reference) rapidly removed U(VI) from solution in the absence of cells. However, when the poise at the electrode was removed, all of the U(VI) returned to solution, demonstrating that the electrode did not reduce U(VI). If G. sulfurreducens was present on the electrode, U(VI) did not return to solution until the electrode was exposed to dissolved oxygen. This suggeststhat G. sulfurreducens on the electrode reduced U(VI) to U(IV) which was stably precipitated until reoxidized in the presence of oxygen. When an electrode was placed in uranium-contaminated subsurface sediments, U(VI) was removed and recovered from groundwater using poised electrodes. Electrodes emplaced in flow-through columns of uranium-contaminated sediments readily removed U(VI) from the groundwater, and 87% of the uranium that had been removed was recovered from the electrode surface after the electrode was pulled from the sediments. These results suggest that microorganisms can use electrons derived from electrodes to reduce U(VI) and that it may be possible to remove and recover uranium from contaminated groundwater with poised electrodes.     

Oxidative Dissolution of Biogenic Uraninite in Groundwater at Old Rifle, CO

Reductive bioremediation is currently being explored as a possible strategy for uranium-contaminated aquifers such as the Old Rifle site (Colorado). The stability of U(IV) phases under oxidizing conditions is key to the performance of this procedure. An in situ method was developed to study oxidative dissolution of biogenic uraninite (UO?), a desirable U(VI) bioreduction product, in the Old Rifle, CO, aquifer under different variable oxygen conditions. Overall uranium loss rates were 50-100 times slower than laboratory rates. After accounting for molecular diffusion through the sample holders, a reactive transport model using laboratory dissolution rates was able to predict overall uranium loss. The presence of biomass further retarded diffusion and oxidation rates. These results confirm the importance of diffusion in controlling in-aquifer U(IV) oxidation rates. Upon retrieval, uraninite was found to be free of U(VI), indicating dissolution occurred via oxidation and removal of surface atoms. Interaction of groundwater solutes such as Ca2? or silicate with uraninite surfaces also may retard in-aquifer U loss rates. These results indicate that the prolonged stability of U(IV) species in aquifers is strongly influenced by permeability, the presence of bacterial cells and cell exudates, and groundwater geochemistry.     

Chromium(VI) reduction kinetics by zero-valent iron in moderately hard water with humic acid: iron dissolution and humic acid adsorption

In zerovalent iron treatment systems, the presence of multiple solution components may impose combined effects that differ from corresponding individual effects. The copresence of humic acid and hardness (Ca2+/Mg2+) was found to influence Cr(VI) reduction by Feo and iron dissolution in a way different from their respective presence in batch kinetics experiments with synthetic groundwater at initial pH 6 and 9.5. Cr(VI) reduction rate constants (k(obs)) were slightly inhibited by humic acid adsorption on iron filings (decreases of 7-9% and 10-12% in the presence of humic acid alone and together with hardness, respectively). The total amount of dissolved Fe steadily increased to 25 mg L(-1) in the presence of humic acid alone because the formation of soluble Fe-humate complexes appeared to suppress iron precipitation. Substantial amounts of soluble and colloidal Fe-humate complexes in groundwater may arouse aesthetic and safety concerns in groundwater use. In contrast, the coexistence of humic acid and Ca2+/Mg2+ significantly promoted aggregation of humic acid and metal hydrolyzed species, as indicated by XPS and TEM analyses, which remained nondissolved (>0.45 microm) in solution. These metal-humate aggregates may impose long-term impacts on PRBs in subsurface settings.     

In situ bioreduction of uranium (VI) to submicromolar levels and reoxidation by dissolved oxygen

Groundwater within Area 3 of the U.S. Department of Energy (DOE) Environmental Remediation Sciences Program (ERSP) Field Research Center at Oak Ridge, TN (ORFRC) contains up to 135 microM uranium as U(VI). Through a series of experiments at a pilot scale test facility, we explored the lower limits of groundwater U(VI) that can be achieved by in-situ biostimulation and the effects of dissolved oxygen on immobilized uranium. Weekly 2 day additions of ethanol over a 2-year period stimulated growth of denitrifying, Fe(III)-reducing, and sulfate-reducing bacteria, and immobilization of uranium as U(IV), with dissolved uranium concentrations decreasing to low levels. Following sulfite addition to remove dissolved oxygen, aqueous U(VI) concentrations fell below the U.S. Environmental Protection Agengy maximum contaminant limit (MCL) for drinking water (< 30/microg L(-1) or 0.126 microM). Under anaerobic conditions, these low concentrations were stable, even in the absence of added ethanol. However, when sulfite additions stopped, and dissolved oxygen (4.0-5.5 mg L(-1)) entered the injection well, spatially variable changes in aqueous U(VI) occurred over a 60 day period, with concentrations increasing rapidly from < 0.13 to 2.0 microM at a multilevel sampling (MLS) well located close to the injection well, but changing little at an MLS well located further away. Resumption of ethanol addition restored reduction of Fe(III), sulfate, and U(VI) within 36 h. After 2 years of ethanol addition, X-ray absorption near-edge structure spectroscopy (XANES) analyses indicated that U(IV) comprised 60-80% of the total uranium in sediment samples. Atthe completion of the project (day 1260), U concentrations in MLS wells were less than 0.1 microM. The microbial community at MLS wells with low U(VI) contained bacteria that are known to reduce uranium, including Desulfovibrio spp. and Geobacter spp., in both sediment and groundwater. The dominant Fe(III)-reducing species were Geothrix spp.     

Bioreduction of uranium: environmental implications of a pentavalent intermediate

The release of uranium and other transuranics into the environment, and their subsequent mobility, are subjects of intense public concern. Uranium dominates the inventory of most medium- and low-level radioactive waste sites and under oxic conditions is highly mobile as U(VI), the soluble uranyl dioxocation (UO2)2+. Specialist anaerobic bacteria are, however, able to reduce U(VI)to insoluble U(IV), offering a strategy for the bioremediation of uranium-contaminated groundwater and a potential mechanism for the biodeposition of uranium ores. Despite the environmental importance of U(VI) bioreduction, there is little information on the mechanism of this transformation. In the course of this study we used X-ray absorption spectroscopy (XAS) to show that the subsurface metal-reducing bacterium Geobacter sulfurreducens reduces U(VI) by a one-electron reduction, forming an unstable (UO2)+ species. The final, insoluble U(IV) product could be formed either through further reduction of U(V) or through its disproportionation. When G. sulfurreducens was challenged with the chemically analogous (NpO2)+, which is stable with respect to disproportionation, it was not reduced, suggesting that it is disproportionation of U(V) which leads to the U(IV) product. This surprising discrimination between U and Np illustrates the need for mechanistic understanding and care in devising in situ bioremediation strategies for complex wastes containing other redox-active actinides, including plutonium.     

Dissolution of technetium(IV) oxide by natural and synthetic organic ligands under both reducing and oxidizing conditions

Technetium-99 (Tc) in nuclear waste is a significant environmental concern due to its long half-life and high mobility in the subsurface. Reductive precipitation of technetium(IV) oxides [TcO(2)(s)] is an effective means of immobilizing Tc, thereby impeding its migration in groundwater. However, technetium(IV) oxides are subject to dissolution by oxidants and/or complexing agents. In this study we ascertain the effects of a synthetic organic ligand, ethylenediaminetetraacetate (EDTA), and two natural humic isolates on the dissolution and solubility of technetium(IV) oxides. Pure synthetic technetium(IV) oxide (0.23 mM) was used in batch experiments to determine dissolution kinetics at pH ～6 under both reducing and oxidizing conditions. All organic ligands were found to enhance the dissolution of technetium(IV) oxides, increasing their solubility from ～10(-8) M (without ligands) to 4 × 10(-7) M under strictly anoxic conditions. Reduced Tc(IV) was also found to reoxidize rapidly under oxic conditions, with an observed oxidative dissolution rate approximately an order of magnitude higher than that of ligand-promoted dissolution under reducing conditions. Significantly, oxidative dissolution was inhibited by EDTA but enhanced by humic acid compared to experiments without any complexing agents. The redox functional properties of humics, capable of facilitating intramolecular electron transfer, may account for this increased oxidation rate under oxic conditions. Our results highlight the importance of complex interactions for the stability and mobility of Tc and thus for the long-term fate of Tc in contaminated environments.     

Solution and microbial controls on the formation of reduced U(IV) species

Reduction of U(VI) to U(IV) as the result of direct or indirect microbial activity is currently being explored for in situ remediation of subsurface U plumes, under the assumption that U(IV) solubility is controlled by the low-solubility mineral uraninite (U(IV)-dioxide). However, recent characterizations of U in sediments from biostimulated field sites, as well as laboratory U(VI) bioreduction studies, report on the formation of U(IV) species that lack the U═O(2)═U coordination of uraninite, suggesting that phases other than uraninite may be controlling U(IV) solubility in environments with complexing surfaces and ligands. To determine the controls on the formation of such nonuraninite U(IV) species, the current work studied the reduction of carbonate-complexed U(VI) by (1) five Gram-positive Desulfitobacterium strains, (2) the Gram-negative bacteria Anaeromyxobacter dehalogenans 2CP-C and Shewanella putrefaciens CN32, and (3) chemically reduced 9,10-anthrahydroquinone-2,6-disulfonate (AH(2)QDS, a soluble reductant). Further, the effects of 0.3 mM dissolved phosphate on U(IV) species formation were explored. Extended X-ray absorption fine structure (EXAFS) spectroscopy analysis demonstrated that the addition of phosphate causes the formation of a nonuraninite, phosphate-complexed U(IV) species, independent of the biological or abiotic mode of U(VI) reduction. In phosphate-free medium, U(VI) reduction by Desulfitobacterium spp. and by AH(2)QDS resulted in nonuraninite, carbonate-complexed U(IV) species, whereas reduction by Anaeromyxobacter or Shewanella yielded nanoparticulate uraninite. These findings suggest that the Gram-positive Desulfitobacterium strains and the Gram-negative Anaeromyxobacter and Shewanella species use distinct mechanisms to reduce U(VI).     

In situ bioreduction of technetium and uranium in a nitrate-contaminated aquifer

The potential to stimulate an indigenous microbial community to reduce a mixture of U(VI) and Tc(VII) in the presence of high (120 mM) initial NO3- co-contamination was evaluated in a shallow unconfined aquifer using a series of single-well, push-pull tests. In the absence of added electron donor, NO3-, Tc(VII), and U(VI) reduction was not detectable. However, in the presence of added ethanol, glucose, or acetate to serve as electron donor, rapid NO3- utilization was observed. The accumulation of NO2-, the absence of detectable NH4+ accumulation, and the production of N2O during in situ acetylene-block experiments suggest that NO3- was being consumed via denitrification. Tc(VII) reduction occurred concurrently with NO3- reduction, but U(VI) reduction was not observed until two or more donor additions resulted in iron-reducing conditions, as detected by the production of Fe(II). Reoxidation/remobilization of U(IV) was also observed in tests conducted with high (approximately 120 mM) but not low (approximately 1 mM) initial NO3- concentrations and not during acetylene-block experiments conducted with high initial NO3-. These results suggest that NO3(-)-dependent microbial U(IV) oxidation may inhibit or reverse U(VI) reduction and decrease the stability of U(IV) in this environment. Changes in viable biomass, community composition, metabolic status, and respiratory state of organisms harvested from down-well microbial samplers deployed during these tests were consistent with the conclusions that electron donor additions resulted in microbial growth, the creation of anaerobic conditions, and an increase in activity of metal-reducing organisms (e.g., Geobacter). The results demonstrate that it is possible to stimulate the simultaneous bioreduction of U(VI) and Tc(VII) mixtures commonly found with NO3- co-contamination at radioactive waste sites.     

Uranium reduction in sediments under diffusion-limited transport of organic carbon

Costly disposal of uranium (U) contaminated sediments is motivating research on in situ U(VI) reduction to insoluble U(IV) via directly or indirectly microbially mediated pathways. Delivery of organic carbon (OC) into sediments for stimulating U bioreduction is diffusion-limited in less permeable regions of the subsurface. To study OC-based U reduction in diffusion-limited regions, one slightly acidic and another calcareous sediment were treated with uranyl nitrate, packed into columns, then hydrostatically contacted with tryptic soy broth solutions. Redox potentials, U oxidation state, and microbial communities were well correlated. At average supply rates of 0.9 micromol OC (g sediment)(-1) day(-1), the U reduction zone extended to only about35-45 mm into sediments. The underlying unreduced U(VI) zone persisted over 600 days because the supply of OC was diffusion-limited and metabolized within a short distance. These results also suggestthat low U concentrations in groundwater samples from OC-treated sediments are not necessarily indicative of pervasive U reduction because interior and exterior regions of such sediment blocks can contain primarily U(VI) and U(IV), respectively.     

Pilot-scale in situ bioremedation of uranium in a highly contaminated aquifer. 2. Reduction of u(VI) and geochemical control of u(VI) bioavailability

In situ microbial reduction of soluble U(VI) to sparingly soluble U(IV) was evaluated at the site of the former S-3 Ponds in Area 3 of the U.S. Department of Energy Natural and Accelerated Bioremediation Research Field Research Center, Oak Ridge, TN. After establishing conditions favorable for bioremediation (Wu, et al. Environ. Sci. Technol. 2006, 40, 3988-3995), intermittent additions of ethanol were initiated within the conditioned inner loop of a nested well recirculation system. These additions initially stimulated denitrification of matrix-entrapped nitrate, but after 2 months, aqueous U levels fell from 5 to approximately 1 microM and sulfate reduction ensued. Continued additions sustained U(VI) reduction over 13 months. X-ray near-edge absorption spectroscopy (XANES) confirmed U(VI) reduction to U(IV) within the inner loop wells, with up to 51%, 35%, and 28% solid-phase U(IV) in sediment samples from the injection well, a monitoring well, and the extraction well, respectively. Microbial analyses confirmed the presence of denitrifying, sulfate-reducing, and iron-reducing bacteria in groundwater and sediments. System pH was generally maintained at less than 6.2 with low bicarbonate level (0.75-1.5 mM) and residual sulfate to suppress methanogenesis and minimize uranium mobilization. The bioavailability of sorbed U(VI) was manipulated by addition of low-level carbonate (< 5 mM) followed by ethanol (1-1.5 mM). Addition of low levels of carbonate increased the concentration of aqueous U, indicating an increased rate of U desorption due to formation of uranyl carbonate complexes. Upon ethanol addition, aqueous U(VI) levels fell, indicating that the rate of microbial reduction exceeded the rate of desorption. Sulfate levels simultaneously decreased, with a corresponding increase in sulfide. When ethanol addition ended but carbonate addition continued, soluble U levels increased, indicating faster desorption than reduction. When bicarbonate addition stopped, aqueous U levels decreased, indicating adsorption to sediments. Changes in the sequence of carbonate and ethanol addition confirmed that carbonate-controlled desorption increased bioavailability of U(VI) for reduction.     

Reduction of U(VI) Incorporated in the Structure of Hematite

U(VI) doped hematite was synthesized and exposed to two different organic reductants with E(0) of 0.23 and 0.70 V. A combination of HAADF-TEM and EXAFS provided evidence that uranium was incorporated in hematite in uranate, likely octahedral coordination. XPS indicated that structurally incorporated U(VI) was reduced to U(V), whereas non-incorporated U(VI) was reduced to U(IV). Specifically, the experiments indicate that U(V) was the dominant oxidation state of uranium in hematite around Eh -0.24 to -0.28 V and pH 7.7-8.6 for at least up to 5 weeks of reaction time. U(V), but not U(IV), was also detected in hematite at Eh +0.21 V (pH 7.1-7.3). The results support the hypothesis, based on previous experimental and theoretical work, that the stability field of U(V) is widened relative to U(IV) and U(VI) in uranate coordination environments where the coordination number of U is less than 8.     

Photodegradation of a ternary iron(III)-uranium(VI)-citric acid complex

The mechanisms of photodegradation of binary iron- and uranium-citrate and ternary iron-uranium-citrate complexes were elucidated. Citric acid degradation products were identified by HPLC and GC, and the metal precipitates were identified by XRD and EXAFS. Photodegradation of a binuclear iron-citrate complex occurred as a result of two one-electron oxidations of citric acid with the formation of 3-oxoglutarate and two ferrous ions. The ferrous ions were reoxidized by a photo-Fenton reaction, resulting in the precipitation of iron as two-line ferrihydrite Fe(OH)3. The citric acid in the uranium-citrate complex underwent a two-electron oxidation to acetoacetate with the concomitant reduction of U(VI) to U(IV). The U(IV) was subsequently photooxidized in the presence of dioxygen with precipitation of uranium as the mineral schoepite (UO3 x 2H2O). A two-step electron reduction of two ferric ions to two ferrous ions wasthe primary mechanism for photodegradation of the ternary iron-uranium-citrate complex with oxidation of citric acid to 3-oxoglutarate; reduction of uranium was not observed. The iron precipitated as ferrihydrite and the uranyl ion as a uranyl hydroxide species. These results show the potential application of photochemical treatment of wastewater and decontamination solutions containing binary and ternary iron- and uranium-citrate complexes.     

Mechanisms of uranium interactions with hydroxyapatite: implications for groundwater remediation

The speciation of U(VI) sorbed to synthetic hydroxyapatite was investigated using a combination of U LIII-edge XAS, synchrotron XRD, batch uptake measurements, and SEM-EDS. The mechanisms of U(VI) removal by apatite were determined in order to evaluate the feasibility of apatite-based in-situ permeable reactive barriers (PRBs). In batch U(VI) uptake experiments with synthetic hydroxyapatite (HA), near complete removal of dissolved uranium (>99.5%) to <0.05 microM was observed over a range of total U(VI) concentrations up to equimolar of the total P in the suspension. XRD and XAS analyses of U(VI)-reacted HA at sorbed concentrations < or = 4,700 ppm U(VI) suggested that uranium(VI) phosphate, hydroxide, and carbonate solids were not present at these concentrations. Fits to EXAFS spectra indicate the presence of Ca neighbors at 3.81 A. U-Ca separation, suggesting that U(VI) adsorbs to the HA surfaces as an inner-sphere complex. Uranium(VI) phosphate solid phases were not detected in HA with 4700 ppm sorbed U(VI) by backscatter SEM or EDS, in agreement with the surface complexation process. In contrast, U(VI) speciation in samples that exceeded 7000 ppm sorbed U(VI) included a crystalline uranium(VI) phosphate solid phase, identified as chernikovite by XRD. At these higher concentrations, a secondary, uranium(VI) phosphate solid was detected by SEM-EDS, consistent with chernikovite precipitation. Autunite formation occurred at total U:P molar ratios > or = 0.2. Our findings provide a basis for evaluating U(VI) sorption mechanisms by commercially available natural apatites for use in development of PRBs for groundwater U(VI) remediation.     

Identification of simultaneous U(VI) sorption complexes and U(IV) nanoprecipitates on the magnetite (111) surface

Sequestration of uranium (U) by magnetite is a potentially important sink for U in natural and contaminated environments. However, molecular-scale controls that favor U(VI) uptake including both adsorption of U(VI) and reduction to U(IV) by magnetite remain poorly understood, in particular, the role of U(VI)-CO(3)-Ca complexes in inhibiting U(VI) reduction. To investigate U uptake pathways on magnetite as a function of U(VI) aqueous speciation, we performed batch sorption experiments on (111) surfaces of natural single crystals under a range of solution conditions (pH 5 and 10; 0.1 mM U(VI); 1 mM NaNO(3); and with or without 0.5 mM CO(3) and 0.1 mM Ca) and characterized surface-associated U using grazing incidence extended X-ray absorption fine structure spectroscopy (GI-EXAFS), grazing incidence X-ray diffraction (GI-XRD), and scanning electron microscopy (SEM). In the absence of both carbonate ([CO(3)](T), denoted here as CO(3)) and calcium (Ca), or in the presence of CO(3) only, coexisting adsorption of U(VI) surface species and reduction to U(IV) occurs at both pH 5 and 10. In the presence of both Ca and CO(3), only U(VI) adsorption (VI) occurs. When U reduction occurs, nanoparticulate UO(2) forms only within and adjacent to surface microtopographic features such as crystal boundaries and cracks. This result suggests that U reduction is limited to defect-rich surface regions. Further, at both pH 5 and 10 in the presence of both CO(3) and Ca, U(VI)-CO(3)-Ca ternary surface species develop and U reduction is inhibited. These findings extend the range of conditions under which U(VI)-CO(3)-Ca complexes inhibit U reduction.     

Non-uraninite products of microbial U(VI) reduction

A promising remediation approach to mitigate subsurface uranium contamination is the stimulation of indigenous bacteria to reduce mobile U(VI) to sparingly soluble U(IV). The product of microbial uranium reduction is often reported as the mineral uraninite. Here, we show that the end products of uranium reduction by several environmentally relevant bacteria (Gram-positive and Gram-negative) and their spores include a variety of U(IV) species other than uraninite. U(IV) products were prepared in chemically variable media and characterized using transmission electron microscopy (TEM) and X-ray absorption spectroscopy (XAS) to elucidate the factors favoring/inhibiting uraninite formation and to constrain molecular structure/composition of the non-uraninite reduction products. Molecular complexes of U(IV) were found to be bound to biomass, most likely through P-containing ligands. Minor U(IV)-orthophosphates such as ningyoite [CaU(PO(4))(2)], U(2)O(PO(4))(2), and U(2)(PO(4))(P(3)O(10)) were observed in addition to uraninite. Although factors controlling the predominance of these species are complex, the presence of various solutes was found to generally inhibit uraninite formation. These results suggest a new paradigm for U(IV) in the subsurface, i.e., that non-uraninite U(IV) products may be found more commonly than anticipated. These findings are relevant for bioremediation strategies and underscore the need for characterizing the stability of non-uraninite U(IV) species in natural settings.     

Speciation-dependent microbial reduction of uranium within iron-coated sands

Transport of uranium within surface and subsurface environments is predicated largely on its redox state. Uranyl reduction may transpire through either biotic (enzymatic) or abiotic pathways; in either case, reduction of U(VI) to U(IV) results in the formation of sparingly soluble UO2 precipitates. Biological reduction of U(VI), while demonstrated as prolific under both laboratory and field conditions, is influenced by competing electron acceptors (such as nitrate, manganese oxides, or iron oxides) and uranyl speciation. Formation of Ca-UO2-CO3 ternary complexes, often the predominate uranyl species in carbonate-bearing soils and sediments, decreases the rate of dissimilatory U(VI) reduction. The combined influence of uranyl speciation within a mineralogical matrix comparable to natural environments and under hydrodynamic conditions, however, remains unresolved. We therefore examined uranyl reduction by Shewanella putrefaciens within packed mineral columns of ferrihydrite-coated quartz sand under conditions conducive or nonconducive to Ca-UO2-CO3 species formation. The results are dramatic. In the absence of Ca, where uranyl carbonato complexes dominate, U(VI) reduction transpires and consumes all of the U(VI) within the influent solution (0.166 mM) over the first 2.5 cm of the flow field for the entirety of the 54 d experiment. Over 2 g of U is deposited during this reaction period, and despite ferrihydrite being a competitive electron acceptor, uranium reduction appears unabated for the duration of our experiments. By contrast, in columns with 4 mM Ca in the influent solution (0.166 mM uranyl), reduction (enzymatic or surface-bound Fe(III) mediated) appears absent and breakthrough occurs within 18 d (at a flow rate of 3 pore volumes per day). Uranyl speciation, and in particular the formation of ternary Ca-UO2-CO3 complexes, has a profound impact on U(VI) reduction and thus transport within anaerobic systems.