Jim K. Fredrickson

Effect of electron donor and solution chemistry on products of dissimilatory reduction of technetium by Shewanella putrefaciens

ABSTRACT To help provide a fundamental basis for use of microbial dissimilatory reduction processes in separating or immobilizing99Tc in waste or groundwaters, the effects of electron donor and the presence of the bicarbonate ion on the rate and extent of pertechnetate ion [Tc(VII)O4−] enzymatic reduction by the subsurface metal-reducing bacterium Shewanella putrefaciens CN32 were determined, and the forms of aqueous and solid-phase reduction products were evaluated through a combination of high-resolution transmission electron microscopy, X-ray absorption spectroscopy, and thermodynamic calculations. When H2served as the electron donor, dissolved Tc(VII) was rapidly reduced to amorphous Tc(IV) hydrous oxide, which was largely associated with the cell in unbuffered 0.85% NaCl and with extracellular particulates (0.2 to 0.001 μm) in bicarbonate buffer. Cell-associated Tc was present principally in the periplasm and outside the outer membrane. The reduction rate was much lower when lactate was the electron donor, with extracellular Tc(IV) hydrous oxide the dominant solid-phase reduction product, but in bicarbonate systems much less Tc(IV) was associated directly with the cell and solid-phase Tc(IV) carbonate may have been present. In the presence of carbonate, soluble (<0.001 μm) electronegative, Tc(IV) carbonate complexes were also formed that exceeded Tc(VII)O4− in electrophoretic mobility. Thermodynamic calculations indicate that the dominant reduced Tc species identified in the experiments would be stable over a range of Eh and pH conditions typical of natural waters. Thus, carbonate complexes may represent an important pathway for Tc transport in anaerobic subsurface environments, where it has generally been assumed that Tc mobility is controlled by low-solubility Tc(IV) hydrous oxide and adsorptive, aqueous Tc(IV) hydrolysis products.

Temporal Shifts in the Geochemistry and Microbial Community Structure of an Ultradeep Mine Borehole Following Isolation

A borehole draining a water-bearing dyke fracture at 3.2-km depth in a South African Au mine was isolated from the open mine environment. Geochemical, stable isotopic, nucleic acid-based, and phospholipid fatty acid (PLFA) analyses were employed as culture-independent means for assessing shifts in the microbial community and habitat as the system equilibrated with the native rock-water environment. Over a two-month period, the pH increased from 5.5 to 7.4, concurrent with a drop in pe from −2 to −3. Whereas rDNAs related to Desulfotomaculum spp. represented the major clone type encountered throughout, lipid biomarker profiling along with 16S rDNA clone library and terminal restriction fragment length polymorphism (T-RFLP) analyses indicated the emergence of other Gram-positive and deeply-branching lineages in samples during the later stages of the equilibration period. A biofilm that formed on the mine wall below the borehole produced abundant rDNAs related to the α Proteobacteria. β- and γ −Proteobacteria appeared to transiently bloom in the borehole shortly after isolation. Chemical modeling and sulfur isotope analyses of the borehole effluent indicated that microbial sulfate reduction was the major terminal electron-accepting process shortly after isolation, whereas Fe+3 reduction dominated towards the end of the experiment. The persistence of Desulfotomaculum-like bacteria throughout suggests that these organisms adapted to changing geochemical conditions as the redox decreased and pH increased following the isolation of the borehole from the mine atmosphere. The restoration of anaerobic aquatic chemistry to this borehole environment may have allowed microbiota indigenous to the local basalt aquifer to become more dominant among the diverse collection of bacterial lineages present in the borehole.

A Global Perspective on the Microbial Abundance and Activity in the Deep Subsurface

Reports that microorganisms and even microbial communities exist at great depths below the continental surface and below the sea floor have steadily increased in the past several years. The best evidence for the presence and diversity of microorganisms at hundreds to thousands of meters below the land surface (mbls.) lies with cores taken using tracers to detect contamination and processed by aseptic methods (Griffin et al., 1997). Other sources of information on the subterranean biosphere include ground water samples and microbial investigations of recently excavated mine faces. This review compares results from these studies with those of the marine subsurface in order to place constraints on the global subsurface biomass and rate of biogeochemical cycling.

Biosolubilization of low-rank coal by a Trametes versicolor siderophore-like product and other complexing agents.

SummaryA heat stable, low molecular weight (<1000) extracellular product inTrametes versicolor (=Coriolus versicolor=Polyporous versicolor) cultures was demonstrated to be a principal factor in the solubilization of leonardite and other low-rank coals. The solubilization of leonardite byT. versicolor cell-free cultures and active fractions was inhibited by Fe3+ and was mimicked by the siderophore desferal mesylate and the iron chelating agents EDTA and 8-hydroxyquinoline. Leonardite solubilization by these later compounds was also inhibited by Fe3+. The ferrated and unferrated form of the partially purified active component fromT. versicolor cultures demonstrated absorption spectra that were similar to the ferrated and unferrated form of desferal mesylate.

Colonization and degradation of oxidized bituminous and lignite coals by fungi

SummaryAPenicillium sp. previously shown to grow on lignite coals degraded an air-oxidized bituminous coal (Illinois #6) to a material that was more than 80% soluble in 0.5 N NaOH. Scanning electron microscopy of the oxidized Illinois #6 revealed colonization of the surface by thePenicillium sp., production of conidia, and erosion of the coal surface. The average molecular weight (MW) of Illinois #6 degraded by the fungus and base-solubilized was approximately 1000 Da. The average MW for base-solubilized Illinois #6 that was not exposed to the fungus was 6000 Da, suggesting solubilizing mechanisms other than base catalysis. A spectrophotometric assay to quantify the microbial conversion of biosolubilized coal was developed. Standard curves were constructed based on the absorbance at 450 nm of different quantities of microbe-solubilized coal. An acid precipitation step was necessary to remove medium and/or microbial metabolites from solubilized coal to prevent overestimation of the extent of coal biosolubilization. Furthermore, the absorption spectra for different coal products varied, necessitating construction of standard curves for individual coals.

Multi-Scale Mass Transfer Processes Controlling Natural Attenuation and Engineered Remediation: An IFRC Focused on Hanford’s 300 Area Uranium Plume

The Integrated Field-Scale Subsurface Research Challenge (IFRC) at the Hanford Site 300 Area uranium (U) plume addresses multi-scale mass transfer processes in a complex hydrogeologic setting where groundwater and riverwater interact. A series of forefront science questions on mass transfer are posed for research which relate to the effect of spatial heterogeneities; the importance of scale; coupled interactions between biogeochemical, hydrologic, and mass transfer processes; and measurements and approaches needed to characterize and model a mass-transfer dominated system. The project was initiated in February 2007, with CY 2007 and CY 2008 progress summarized in preceding reports. The site has 35 instrumented wells, and an extensive monitoring system. It includes a deep borehole for microbiologic and biogeochemical research that sampled the entire thickness of the unconfined 300 A aquifer. Significant, impactful progress has been made in CY 2009 with completion of extensive laboratory measurements on field sediments, field hydrologic and geophysical characterization, four field experiments, and modeling. The laboratory characterization results are being subjected to geostatistical analyses to develop spatial heterogeneity models of U concentration and chemical, physical, and hydrologic properties needed for reactive transport modeling. The field experiments focused on: (1) physical characterization of the groundwater flow field during amorexa0» period of stable hydrologic conditions in early spring, (2) comprehensive groundwater monitoring during spring to characterize the release of U(VI) from the lower vadose zone to the aquifer during water table rise and fall, (3) dynamic geophysical monitoring of salt-plume migration during summer, and (4) a U reactive tracer experiment (desorption) during the fall. Geophysical characterization of the well field was completed using the down-well Electrical Resistance Tomography (ERT) array, with results subjected to robust, geostatistically constrained inversion analyses. These measurements along with hydrologic characterization have yielded 3D distributions of hydraulic properties that have been incorporated into an updated and increasingly robust hydrologic model. Based on significant findings from the microbiologic characterization of deep borehole sediments in CY 2008, down-hole biogeochemistry studies were initiated where colonization substrates and spatially discrete water and gas samplers were deployed to select wells. The increasingly comprehensive field experimental results, along with the field and laboratory characterization, are leading to a new conceptual model of U(VI) flow and transport in the IFRC footprint and the 300 Area in general, and insights on the microbiological community and associated biogeochemical processes. A significant issue related to vertical flow in the IFRC wells was identified and evaluated during the spring and fall field experimental campaigns. Both upward and downward flows were observed in response to dynamic Columbia River stage. The vertical flows are caused by the interaction of pressure gradients with our heterogeneous hydraulic conductivity field. These impacts are being evaluated with additional modeling and field activities to facilitate interpretation and mitigation. The project moves into CY 2010 with ambitious plans for a drilling additional wells for the IFRC well field, additional experiments, and modeling. This research is part of the ERSP Hanford IFRC at Pacific Northwest National Laboratory.«xa0less

Microbial life in the deep terrestrial subsurface

The distribution and function of microorganisms is a vital issue in microbial ecology. The US Department of Energy`s Program, ``Microbiology of the Deep Subsurface,`` concentrates on establishing fundamental scientific information about organisms at depth, and the use of these organisms for remediation of contaminants in deep vadose zone and groundwater environments. This investigation effectively extends the Biosphere hundreds of meters into the Geosphere and has implications to a variety of subsurface activities.

Multi-Scale Mass Transfer Processes Controlling Natural Attenuation and Engineered Remediation: An IFRC Focused on Hanford’s 300 Area Uranium Plume January 2010 to January 2011

The Integrated Field Research Challenge (IFRC) at the Hanford Site 300 Area uranium (U) plume addresses multi-scale mass transfer processes in a complex subsurface hydrogeologic setting where groundwater and riverwater interact. A series of forefront science questions on reactive mass transfer focus research. These questions relate to the effect of spatial heterogeneities; the importance of scale; coupled interactions between biogeochemical, hydrologic, and mass transfer processes; and measurements and approaches needed to characterize and model a mass-transfer dominated system. The project was initiated in February 2007, with CY 2007, CY 2008, and CY 2009 progress summarized in preceding reports. A project peer review was held in March 2010, and the IFRC project has responded to all suggestions and recommendations made in consequence by reviewers and SBR/DOE. These responses have included the development of “Modeling” and “Well-Field Mitigation” plans that are now posted on the Hanford IFRC web-site. The site has 35 instrumented wells, and an extensive monitoring system. It includes a deep borehole for microbiologic and biogeochemical research that sampled the entire thickness of the unconfined 300 A aquifer. Significant, impactful progress has been made in CY 2010 including the quantification of well-bore flows in the fully screened wells and the testing of means to mitigate them; the development of site geostatistical models of hydrologic and geochemical properties including the distribution of U; developing and parameterizing a reactive transport model of the smear zone that supplies contaminant U to the groundwater plume; performance of a second passive experiment of the spring water table rise and fall event with a associated multi-point tracer test; performance of downhole biogeochemical experiments where colonization substrates and discrete water and gas samplers were deployed to the lower aquifer zone; and modeling of past injection experiments for model parameterization, deconvolution of well-bore flow effects, system understanding, and publication. We continued efforts to assimilate geophysical logging and 3D ERT characterization data into our site wide geophysical model, and have now implemented a new strategy for this activity to bypass an approach that was found unworkable. An important focus of CY 2010 activities has been infrastructure modification to the IFRC site to eliminate vertical well bore flows in the fully screened wells. The mitigation procedure was carefully evaluated and is now being implementated. A new experimental campaign is planned for early spring 2011 that will utilize the modified well-field for a U reactive transport experiment in the upper aquifer zone. Preliminary geophysical monitoring experiments of rainwater recharge in the vadose zone have been initiated with promising results, and a controlled infiltration experiment to evaluate U mobilization from the vadose zone is now under planning for the September 2011. The increasingly comprehensive field experimental results, along with the field and laboratory characterization, are leading to a new conceptual model of U(VI) flow and transport in the IFRC footprint and the 300 Area in general, and insights on the microbiological community and associated biogeochemical processes.