Peyton and Sani and the team in general reported in a major series of reports that the understanding of subsurface microbiology is hindered by the inaccessibility of this environment, particularly when the hydrogeologic medium is contaminated with toxic substances. Research in their labs indicated that the composition of the growth medium (e.g., bicarbonate complexation of U(VI)) and the underlying mineral phase (e.g., hematite) significantly affects the rate and extent of U(VI) reduction and immobilization through a variety of effects.
The authors research was aimed at elucidating those effects to a much greater extent, while exploring the potential for U(IV) reoxidation and subsequent re mobilization, which also appears to depend on the mineral phases present in the system. In situ coupons with a variety of mineral phases were placed in monitoring wells at the NABIR FRC. These coupons showed that the mineral phase composition significantly affected the resulting attached phase microbial community. Their comparative use of both batch and open flow reactors (more representative of field conditions) indicates that hydrodynamics and continual influx of substrate and contaminants can also yield significantly different results than those obtained with closed serum bottles.
To this end, the following overall experimental hypothesis tested was the following: On a mineral surface under anaerobic conditions, accumulations of secondary inorganic precipitates are controlled by:
a) the bacteria associated with the mineral surface,
b) the electron acceptors available for anaerobic bacterial respiration, and
c) local hydrodynamics and pH buffers govern micro- and meso-scale interaction of U in the presence of electron donors and acceptors, and nutrients.
The combined laboratory and field research extended fundamental results based on U(VI) reduction and immobilization with Desulfovibrio desulfuricans G20 as a model for the sulfate reducing bacteria (SRB) on hematite (found in FRC Nolichucky shale) and on other mineral phases. Using techniques developed to specifically probe biogeochemical processes, we expanded the current understanding of the roles played by mineral surfaces and aqueous chemistry on the reduction and immobilization of uranium by SRB.
Further, the authors expanded this understanding into the FRC field site through the emplacement of in situ biofilm coupons. At the FRC site, surrogate geological media contained in a porous receptacle were incubated in a well within the saturated zone of a pristine region of an aquifer to capture populations from the extant communities. Minerals included micaceous, specular hematite (1-3 mm diameter) from Minas Gerais, Brazil (kindly provided by K. Rosso, Pacific Northwest National Laboratory), illite shale (Ward’s Natural Science, cat no. 46 V 0315), uncontaminated saprolite from the FRC subsurface (a gift from P. Jardine, Oak Ridge National Laboratory), and course quartz sand.
Biofilm reactors were used to test uranium immobilization by D. desulfuricans G20 biofilms. These reactors included a flat plate flow reactor and 3 column reactors filled with hematite, dolomite and calcite. All reactors were fed with uranium in an SRB metal toxicity medium (MTM) developed in our laboratory that eliminates the formation of metal precipitates and minimizes metal complexation. The reactors were operated for approximately 4 months and were
continuously monitored for microbial activity and also uranium concentration in the outlet solutions. Over the four-month period, the uranium immobilization efficiency was 88% for the flat plate reactor, 99% for the dolomite column, 99% for the calcite column, and 85 % for the
hematite column.
In addition to mineral type, the effects of pH buffer on U(VI) reduction by D. desulfuricans G20 were studied using MTM, which showed that U(VI) is toxic to G20. Toxicity depended on the medium buffer and was observed in terms of longer lag times and in some cases, no measurable Peyton et al. Final Report - Biogeochemistry of Uranium … growth. The minimum inhibiting concentration (MIC=lowest concentration that utterly inhibits growth) of U(VI) in medium containing PIPES buffer was 140 μM, which is 40 times lower than previously reported for D. desulfuricans.
The effects of Fe(III)-(hydr)oxides (hematite, goethite, and ferrihydrite) on microbial reduction and reoxidation of uranium (U) were also evaluated under lactate-limited sulfate-reducing conditions. With lactate present, G20 reduced U(VI) in both PIPES and bicarbonate buffer. Once lactate was depleted, however, microbially reduced U served as an electron donor to reduce Fe(III) present in Fe(III)-(hydr)oxides. With the same initial amount of Fe(III) (10 mmol/L) for each Fe(III)-(hydr)oxide, reoxidation of U(IV) was greater with hematite than with goethite or ferrihydrite. As the initial mass loading of hematite increased from 0 – 20 mmol Fe(III)/L, the rate and extent of U(IV) reoxidation increased. Subsequent addition of hematite (15 mmol Fe(III)/L) to stationary phase cultures containing microbially reduced U(IV) also resulted in rapid reoxidation to U(VI). Analysis by U L3-edge XANES spectroscopy of microbially reduced U particles yielded spectra similar to that of natural uraninite.
Observations by high-resolution transmission electron microscopy, selected area electron diffraction, and energy-dispersive X-ray spectroscopic analysis confirmed that precipitated U associated with cells was uraninite with particle diameters of 3 – 5 nm. Our results suggest that in the presence of Fe(III)-(hydr)oxides, Fe(III) may act as an oxidant for U(IV) under electron donor-limited conditions, whereas Fe(II) may act as a reductant for U(VI) under other conditions.
This research has made significant advances in the effort to determine the nature of iron mineral surfaces in microcosm and flow reactors. In addition, subsurface “coupons” emplaced at the FRC have presented well-defined mineral surfaces to indigenous microflora for subsequent characterization and assessment of the resulting mineral-associated bacterial communities. Data obtained from this combination of field characterization, in-well coupons, and laboratory reactors are being used to address issues connected with accumulation of surface-associated bacteria, secondary inorganic precipitates, and ultimate stability of reduced uranium.
Characterizing, understanding, and finally controlling these combinations of biological and inorganic solid phases is likely the key to success for in situ bio-immobilization of radionuclides in fractured aquifer systems.
The results have been published in a number of peer reviewed journal articles. The abstracts and citations to those articles make up the bulk of our final report. Our interdisciplinary team members include:
Brent Peyton and Rajesh Sani (Washington State University),
Gill Geesey, Zbigniew Lewandowski, and Haluk Beyenal (Montana State University),
David Cummings (Idaho National Engineering and Environmental Laboratory) and
James Amonette (Pacific Northwest National Laboratory).
Managed & Built by I2M Consulting, LLC | © 2010-
