Previously, it has been shown that not only is uranium reduction under fermentation condition common among clostridia species, but also the strains differed in the extent of their capability and the pH of the culture significantly affected uranium(VI) reduction. In this study, using HPLC and GC techniques, metabolic properties of those clostridial strains active in uranium reduction under fermentation conditions have been characterized and their effects on capability variance of uranium reduction discussed. Then, the relationship between hydrogen metabolism and uranium reduction has been further explored and the important role played by hydrogenase in uranium(VI) and iron(III) reduction by clostridia demonstrated. When hydrogen was provided as the headspace gas, uranium(VI) reduction occurred in the presence of whole cells of clostridia. This is in contrast to that of nitrogen as the headspace gas. Without clostridia cells, hydrogen alone could not result in uranium(VI) reduction. In alignment with this observation, it was also found that either copper(II) addition or iron depletion in the medium could compromise uranium reduction by clostridia. In the end, a comprehensive model was proposed to explain uranium reduction by clostridia and its relationship to the overall metabolism especially hydrogen (H2) production. 1. Introduction Subsurface contamination by radionuclides and toxic metals is a major problem throughout the U.S. Department of Energy’s (DOE’s) complex; uranium contamination evokes particular environmental concern due to the high solubility and mobility of its oxidized form, U(VI). As physically removing the contaminated material is financially prohibitive, we need innovative, cost-effective in situ stabilization technologies that exploit the processes of natural attenuation. Recently, researchers at some DOE sites have assessed the microbial stabilization of actinides (U, Pu, and Np) and fission products (Tc) in subsurface environments, as in the uranium mill tailing remedial action (UMTRA) site in Rifle, CO (http://www.gjem.energy.gov/moab/) and at the Oak Ridge Field Research Centre (ORFRC) at Oak Ridge, TN (http://www.esd.ornl.gov/orifrc/). A wide variety of bacteria, including Desulfovibrio, Geobacter, and Shewanella, can couple the oxidation of organic compounds to the reduction of U(VI) and thus reductively precipitate uranium in its reduced form U(IV) [1–3]. Investigators have explored the mechanisms of uranium(VI) reduction by anaerobic bacteria [4–6]. Clostridia are anaerobic fermenting bacteria. They are gram-positive,
References
[1]
E. Cardenas, W. M. Wu, B. M. Leigh et al., “Significant association between sulfate-reducing bacteria and uranium-reducing microbial communities as revealed by a combined massively parallel sequencing-indicator species approach,” Applied and Environmental Microbiology, vol. 76, no. 20, pp. 6778–6786, 2010.
[2]
Y. Liang, J. D. Van Nostrand, L. A. N'guessan, et al., “Microbial functional gene diversity with a shift of subsurface redox conditions during in situ uranium reduction,” Applied and Environmental Microbiology, vol. 76, no. 20, pp. 6778–6786, 2012.
[3]
J. D. Van Nostrand, L. Wu, W. M. Wu et al., “Dynamics of microbial community composition and function during in situ bioremediation of a uranium-contaminated aquifer,” Applied and Environmental Microbiology, vol. 77, no. 11, pp. 3860–3869, 2011.
[4]
J. C. Renshaw, L. J. C. Butchins, F. R. Livens, I. May, J. M. Charnock, and J. R. Lloyd, “Bioreduction of uranium: environmental implications of a pentavalent intermediate,” Environmental Science and Technology, vol. 39, no. 15, pp. 5657–5660, 2005.
[5]
J. D. Wall and L. R. Krumholz, “Uranium reduction,” Annual Review of Microbiology, vol. 60, pp. 149–166, 2006.
[6]
M. J. Wilkins, F. R. Livens, D. J. Vaughan, and J. R. Lloyd, “The impact of Fe(III)-reducing bacteria on uranium mobility,” Biogeochemistry, vol. 78, no. 2, pp. 125–150, 2006.
[7]
S. B. Leschine, “Cellulose degradation in anaerobic environments,” Annual Review of Microbiology, vol. 49, pp. 399–426, 1995.
[8]
D. R. Woods, “The genetic engineering of microbial solvent production,” Trends in Biotechnology, vol. 13, no. 7, pp. 259–264, 1995.
[9]
J. G. Zeikus, “Chemical and fuel production by anaerobic bacteria,” Annual Review of Microbiology, vol. 34, pp. 423–464, 1980.
[10]
J. S. Chen and J. L. Johnson, “Molecular biology of nitrogen fixation in the clostridia,” Biotechnology, vol. 25, pp. 371–392, 1993.
[11]
H. G. Wood and L. G. Ljungdahl, “Autotrophic character of acetogenic bacteria,” in Variations in Autotrophic Life, J. M. Shively and L. L. Barton, Eds., pp. 201–250, Academic Press, San Diego, Calif, USA, 1991.
[12]
A. Karnholz, K. Küsel, A. G？？ner, A. Schramm, and H. L. Drake, “Tolerance and metabolic response of acetogenicbacteria toward oxygen,” Applied and Environmental Microbiology, vol. 68, pp. 1005–1009, 2002.
[13]
K. Küsel, A. Karnholz, T. Trinkwalter, R. Devereux, G. Acker, and H. L. Drake, “Physiological ecology of Clostridium glycolicum RD-1, an aerotolerant acetogen isolated from sea grass roots,” Applied and Environmental Microbiology, vol. 67, no. 10, pp. 4734–4741, 2001.
[14]
A. J. Francis, C. J. Dodge, F. Lu, G. P. Halada, and C. R. Clayton, “XPS and XANES studies of uranium reduction by Clostridium sp.,” Environmental Science Technology, vol. 28, no. 4, pp. 636–639, 1994.
[15]
A. J. Francis, C. J. Dodge, and G. E. Meinken, “Biotransformation of pertechnetate by Clostridia,” Radiochimica Acta, vol. 90, no. 9–11, pp. 791–797, 2002.
[16]
A. J. Francis, G. Joshi-T, C. J. Dodge, and J. B. Gillow, “Biotransformation of uranium and transition metal citrate complexes by Clostridia,” Journal of Nuclear Science and Technology, vol. 3, supplement, pp. 935–938, 2002.
[17]
W. Gao and A. J. Francis, “Reduction of uranium(VI) to (IV) by Clostrdia,” Applied and Environmental Microbiology, vol. 74, pp. 4580–4584, 2008.
[18]
L. Petrie, N. N. North, S. L. Dollhopf, D. L. Balkwill, and J. E. Kostka, “Enumeration and characterization of iron(III)-reducing microbial communities from acidic subsurface sediments contaminated with uranium(VI),” Applied and Environmental Microbiology, vol. 69, no. 12, pp. 7467–7479, 2003.
[19]
Y. Suzuki, S. D. Kelly, K. M. Kemner, and J. F. Banfield, “Microbial populations stimulated for hexavalent uranium reduction in uranium mine sediment,” Applied and Environmental Microbiology, vol. 69, no. 3, pp. 1337–1346, 2003.
[20]
W. Dong, G. Xie, T. R. Miller et al., “Sorption and bioreduction of hexavalent uranium at a military facility by the Chesapeake Bay,” Environmental Pollution, vol. 142, no. 1, pp. 132–142, 2006.
[21]
A. S. Madden, A. C. Smith, D. L. Balkwill, L. A. Fagan, and T. J. Phelps, “Microbial uranium immobilization independent of nitrate reduction,” Environmental Microbiology, vol. 9, no. 9, pp. 2321–2330, 2007.
[22]
A. J. Francis and C. J. Dodge, “Anaerobic microbial dissolution of transition and heavy metal oxides,” Applied and Environmental Microbiology, vol. 54, pp. 1009–1014, 1988.
[23]
E. R. Weyer and L. F. Rettger, “A comparative study of six different strains of the organism commonly concerned in large-scale production of butyl alcohol and acetone by the biological process,” Journal of Bacteriology, vol. 14, pp. 399–424, 1927.
[24]
J. N？lling, G. Breton, M. V. Omelchenko et al., “Genome sequence and comparative analysis of the solvent-producing bacterium Clostridium acetobutylicum,” Journal of Bacteriology, vol. 183, no. 16, pp. 4823–4838, 2001.
[25]
A. J. Francis, S. Dobbs, and B. J. Nine, “Microbial activity of trench leachates from shallow-land, low-level radioactive waste disposal sites,” Applied and Environmental Microbiology, vol. 40, no. 1, pp. 108–113, 1980.
[26]
J. C. Cox, D. G. Nicholls, and W. J. Ingledew, “Transmembrane electrical potential and transmembrane pH gradient in the acidophile Thiobacillus ferrooxidans,” Biochemical Journal, vol. 178, no. 1, pp. 195–200, 1979.
[27]
P. D. Cotter and C. Hill, “Surviving the acid test: responses of gram-positive bacteria to low pH,” Microbiology and Molecular Biology Reviews, vol. 67, no. 3, pp. 429–453, 2003.
