Charles T. Resch

Stimulating the In Situ Activity of Geobacter Species To Remove Uranium from the Groundwater of a Uranium-Contaminated Aquifer

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 μM 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.

Sorption of divalent metals on calcite

The sorption of seven divalent metals (Ba, Sr, Cd, Mn, Zn, Co, and Ni) was measured on calcite over a large initial metal (Me) concentration range (10−8 to 10−4 mol/L) in constant ionic strength (I = 0.1), equilibrium CaCO3(s)-CaCO3(aq) suspensions that varied in pH. At higher initial Me concentrations (10−5 to 1−4 mol/L) geochemical calculations indicated that the equilibrium solutions were saturated with discrete solid phases of the sorbates: CdCO3(s), MnCO3(s), Zn5(OH)6(CO3)2(s), Co(OH)2(s), and Ni(OH)2(s), implying that aqueous concentrations were governed by solubility. However, significant sorption of all the metals except for Ba and Sr was observed at aqueous concentrations below saturation with Me-solid phases. Divalent metal ion sorption was dependent on aqueous Ca concentration, and the following selectivity sequence was observed: Cd > Zn ≥ Mn > Co > Ni > Ba = Sr. The metals varied in their sorption reversibility, which was correlated with the single-ion hydration energies of the metal sorbates. The strongly hydrated metals (Zn, Co, and Ni) were most desorbable. A sorption model that included aqueous speciation and Me2+-Ca2+ exchange on cation-specific surface sites was developed that described most of the data well. The chemical nature of the surface complex used in this model was unspecified and could represent either a hydrated or dehydrated surface complex, or a surface precipitate. A single exchange constant for Cd, Mn, Co, and Ni could describe the sorption of that metal over a wide range in pH, Ca concentration, and surface concentration. Zinc, however, exhibited nonlinear sorption behavior and required exchange constants that varied with surface coverage. Our data suggested that (i) Cd and Mn dehydrate soon after their adsorption to calcite and form a phase that behaves like a surface precipitate, and (ii) Zn, Co, and Ni form surface complexes that remain hydrated until the ions are incorporated into the structure by recrystallization.

Chromate adsorption on amorphous iron oxyhydroxide in the presence of major groundwater ions.

Chromate adsorption on amorphous iron oxyhydroxide was investigated in dilute iron suspensions as a single solute and in solutions of increasing complexity containing CO2(g), SO4S (aq), H4SiO4(aq), and cations (K , MgS , CaS (aq)). In paired-solute systems (e.g., CrO4S -H2CO3*), anionic cosolutes markedly reduce CrO4S adsorption through a combination of competitive and electrostatic effects, but cations exert no appreciable influence. Additionally, H4SiO4 exhibits a strong time-dependent effect: CrO4S adsorption is greatly decreased with increasing H4SiO4 contact time. In multiple-ion mixtures, each anion added to the mixture decreases CrO4S adsorption further. Adsorption constants for the individual reactive solutes were used in the triple-layer model. The model calculations are in good agreement with the CrO4S adsorption data for paired- and multiple-solute systems. However, the model calculations underestimate CrO4S adsorption when surface site saturation is approached. Questions remain regarding the surface interactions of both CO2(aq) and H4SiO4. The results have major implications for the adsorption behavior of CrO4S and other oxyanions in subsurface waters.

Microbiological and Geochemical Heterogeneity in an In Situ Uranium Bioremediation Field Site

ABSTRACT The geochemistry and microbiology of a uranium-contaminated subsurface environment that had undergone two seasons of acetate addition to stimulate microbial U(VI) reduction was examined. There were distinct horizontal and vertical geochemical gradients that could be attributed in large part to the manner in which acetate was distributed in the aquifer, with more reduction of Fe(III) and sulfate occurring at greater depths and closer to the point of acetate injection. Clone libraries of 16S rRNA genes derived from sediments and groundwater indicated an enrichment of sulfate-reducing bacteria in the order Desulfobacterales in sediment and groundwater samples. These samples were collected nearest the injection gallery where microbially reducible Fe(III) oxides were highly depleted, groundwater sulfate concentrations were low, and increases in acid volatile sulfide were observed in the sediment. Further down-gradient, metal-reducing conditions were present as indicated by intermediate Fe(II)/Fe(total) ratios, lower acid volatile sulfide values, and increased abundance of 16S rRNA gene sequences belonging to the dissimilatory Fe(III)- and U(VI)-reducing family Geobacteraceae. Maximal Fe(III) and U(VI) reduction correlated with maximal recovery of Geobacteraceae 16S rRNA gene sequences in both groundwater and sediment; however, the sites at which these maxima occurred were spatially separated within the aquifer. The substantial microbial and geochemical heterogeneity at this site demonstrates that attempts should be made to deliver acetate in a more uniform manner and that closely spaced sampling intervals, horizontally and vertically, in both sediment and groundwater are necessary in order to obtain a more in-depth understanding of microbial processes and the relative contribution of attached and planktonic populations to in situ uranium bioremediation.

Influence of humic substances on Co2+ sorption by a subsurface mineral separate and its mineralogic components

The sorption of Co2+ (10−6 mol/L) was measured on subsurface mineral materials in the absence and presence of a sorbed leonardite humic acid (LHA) to 1. (1) evaluate the sorptive role of mineral-bound humic substances, and 2. (2) establish approaches to model metal ion binding in composite materials. The subsurface materials were a < 2.0 μm size fraction of an ultisol saprolite (CP) and this same material treated with dithionite-citrate-bicarbonate (DCB) to remove Fe-oxides (DCP). Comparable experiments (with and without LHA) were also performed with mineral sorbents representing dominant phases in the CP separate (gibbsite, Al-goethite, and kaolinite) to evaluate their potential contributions to Co sorption. The mineral-bound LHA ranged in concentration between 0.1–0.4 mg-C/m2, representing approximately 0.7% of the subsurface isolate by mass. The sorption-desorption of LHA on the mineral surfaces, and its affinity for Co as a aqueous phase complexant were also determined. Batch measurements were employed (sorbents at 20–90 m2/L; LHA-DOC at ≈11 mg-C/L) over a range in pH and ionic strength (I) at I = 0.01 and 0.1 in NaClO4. The LHA strongly sorbed to the subsurface mineral isolates (CP and DCP), and to all the specimen sorbents except kaolinite. Maximum sorption of LHA occurred at lower pH (≈4.5). In solid-free suspensions, the affinity of LHA for Co increased with pH and decreasing I (Kd ranging 20–450 L/g). Mineral-bound LHA increased Co sorption on all the sorbents by factors of 10–60 %, with the greatest augmentation noted at pH values (4.5–6.5) where 1. (1) maximum LHA sorption occurred, and 2. (2) Co sorption to the mineral phase was weak and dominated by ion exchange. The LHA appeared simply to augment, rather than to change the intrinsic adsorption behavior of the mineral sorbents. Accordingly, predictions of the Kd for Co on the LHA-coated subsurface materials (DCP, CP) based on a linear additivity model agreed well with the experimental data, suggesting that the complex humic-mineral association acted as a noninteractive sorbent mixture at low aqueous Co concentrations.

Quinoline sorption to subsurface materials: role of pH and retention of the organic cation

The sorption of quinoline (pK/sub a/ = 4.94) was investigated on low-organic-carbon subsurface materials that varied in pH. Sorption isotherms were measured from 10/sup -7/ to 10/sup -4/ M quinoline and were found to be nonlinear. The resulting Freundlich constant (K/sub F/), based on total aqueous quinoline concentration, were poorly correlated with subsoil properties, including organic carbon. Higher sorption in the acidic subsoils and favorable coefficients for regression of K/sub F/, normalized to cation-exchange capacity vs. the ionization fraction (Q), point to the importance of ion exchange of the protonated compound. When the subsoil pH is adjusted, it is shown that sorption parallels the ionization fraction and retention of the organic cation far exceeds that of the neutral species. Calculations of surface speciation and thermodynamic parameters of sorption (..delta..H/sup 0/, ..delta..S/sup 0/) point to ion exchange and/or surface protonation at pH, levels exceeding pK/sub a/ by greater than 2 log units. It is suggested that in subsurface materials of low carbon content, quinoline sorption is controlled by pH the nature and capacity of the exchange complex, and groundwater ion composition. 46 references, 5 figures, 4 tables.

Uranium in Framboidal Pyrite from a Naturally Bioreduced Alluvial Sediment

Samples of a naturally bioreduced, U-contaminated alluvial sediment were characterized with various microscopic and spectroscopic techniques and wet chemical extraction methods. The objective was to investigate U association and interaction with minerals of the sediment. Bioreduced sediment comprises approximately 10% of an alluvial aquifer adjacent to the Colorado River, in Rifle, CO, that was the site of a former U milling operation. Past and ongoing research has demonstrated that bioreduced sediment is elevated in solid-associated U, total organic carbon, and acid-volatile sulfide, and depleted in bioavailable Fe(III) confirming that sulfate and Fe(III) reduction have occurred naturally in the sediment. SEM/EDS analyses demonstrated that framboidal pyrites (FeS(2)) of different sizes ( approximately 10-20 microm in diameter), and of various microcrystal morphology, degree of surface weathering, and internal porosity were abundant in the <53 microm fraction (silt + clay) of the sediment and absent in adjacent sediments that were not bioreduced. SEM-EMPA, XRF, EXAFS, and XANES measurements showed elevated U was present in framboidal pyrite as both U(VI) and U(IV). This result indicates that U may be sequestered in situ under conditions of microbially driven sulfate reduction and pyrite formation. Conversely, such pyrites in alluvial sediments provide a long-term source of U under conditions of slow oxidation, contributing to the persistence of U of some U plumes. These results may also help in developing remedial measures for U-contaminated aquifers.

Spatial and temporal dynamics of the microbial community in the Hanford unconfined aquifer.

Pyrosequencing analysis of 16S rRNA genes was used to study temporal dynamics of groundwater bacteria and archaea over 10 months within three well clusters separated by ∼30 m and located 250 m from the Columbia River on the Hanford Site, WA. Each cluster contained three wells screened at different depths ranging from 10 to 17 m that differed in hydraulic conductivities. Representative samples were selected for analyses of prokaryotic 16S and eukaryotic 18S rRNA gene copy numbers. Temporal changes in community composition occurred in all nine wells over the 10-month sampling period. However, there were particularly strong effects near the top of the water table when the seasonal rise in the Columbia River caused river water intrusion at the top of the aquifer. The occurrence and disappearance of some microbial assemblages (such as Actinobacteria ACK-M1) were correlated with river water intrusion. This seasonal impact on microbial community structure was greater in the shallow saturated zone than deeper zone in the aquifer. Spatial and temporal patterns for several 16S rRNA gene operational taxonomic units associated with particular physiological functions (for example, methane oxidizers and metal reducers) suggests dynamic changes in fluxes of electron donors and acceptors over an annual cycle. In addition, temporal dynamics in eukaryotic 18S rRNA gene copies and the dominance of protozoa in 18S clone libraries suggest that bacterial community dynamics could be affected not only by the physical and chemical environment but also by top-down biological control.

Groundwater–surface water mixing shifts ecological assembly processes and stimulates organic carbon turnover

Environmental transitions often result in resource mixtures that overcome limitations to microbial metabolism, resulting in biogeochemical hotspots and moments. Riverine systems, where groundwater mixes with surface water (the hyporheic zone), are spatially complex and temporally dynamic, making development of predictive models challenging. Spatial and temporal variations in hyporheic zone microbial communities are a key, but understudied, component of riverine biogeochemical function. Here, to investigate the coupling among groundwater–surface water mixing, microbial communities and biogeochemistry, we apply ecological theory, aqueous biogeochemistry, DNA sequencing and ultra-high-resolution organic carbon profiling to field samples collected across times and locations representing a broad range of mixing conditions. Our results indicate that groundwater–surface water mixing in the hyporheic zone stimulates heterotrophic respiration, alters organic carbon composition, causes ecological processes to shift from stochastic to deterministic and is associated with elevated abundances of microbial taxa that may degrade a broad suite of organic compounds.

Competitive reduction of pertechnetate (99TcO4-) by dissimilatory metal reducing bacteria and biogenic Fe(II).

The fate of pertechnetate ((99)Tc(VII)O(4)(-)) during bioreduction was investigated in the presence of 2-line ferrihydrite (Fh) and various dissimilatory metal reducing bacteria (DMRB) (Geobacter, Anaeromyxobacter, Shewanella) in comparison with TcO(4)(-) bioreduction in the absence of Fh. In the presence of Fh, Tc was present primarily as a fine-grained Tc(IV)/Fe precipitate that was distinct from the Tc(IV)O(2)·nH(2)O solids produced by direct biological Tc(VII) reduction. Aqueous Tc concentrations (<0.2 μm) in the bioreduced Fh suspensions (1.7 to 3.2 × 10(-9) mol L(-1)) were over 1 order of magnitude lower than when TcO(4)(-) was biologically reduced in the absence of Fh (4.0 × 10(-8) to 1.0 × 10(-7) mol L(-1)). EXAFS analyses of the bioreduced Fh-Tc products were consistent with variable chain length Tc-O octahedra bonded to Fe-O octahedra associated with the surface of the residual or secondary Fe(III) oxide. In contrast, biogenic TcO(2)·nH(2)O had significantly more Tc-Tc second neighbors and a distinct long-range order consistent with small particle polymers of TcO(2). In Fe-rich subsurface sediments, the reduction of Tc(VII) by Fe(II) may predominate over direct microbial pathways, potentially leading to lower concentrations of aqueous (99)Tc(IV).