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2011 Annual Science Report

University of Wisconsin Reporting  |  SEP 2010 – AUG 2011

Project 4A: Characterization of Novel Solid-Phase Fe(II)-Oxidizing Chemolithotrophic Bacteria From Subsurface Environments

Project Summary

Ferrous iron (Fe(II)) can serve as an energy source for a wide variety of chemolithotrophic microorganisms (organisms that gain energy from metabolism of inorganic compounds). Fe(II) oxidation may have played a role in past (and possibly, present) life on Mars, whose crust is rich in primary Fe(II)-bearing silicate minerals, as well as Fe-bearing clay minerals formed during weathering of primary silicates. This project examined the physiological and phylogenetic properties of novel solid-phase Fe(II)-oxidizing bacteria (FeOB) isolated from subsurface sediments from the Columbia River basin in eastern Washington, as well as clay-rich subsoils from Madison, WI. The organisms were enriched using biotite (a Fe(II)-bearing primary silicate mineral) as an energy source, and purified on organics-containing medium. The capacity of the isolates to oxidize soluble and solid-phase Fe(II) compounds was assessed, and the 16S rRNA gene sequence for each of the isolate was determined. The results revealed that a wide variety of Proteobacteria are capable of catalyzing solid-phase Fe(II) oxidation, including several groups of organisms not previously known as FeOB. These results confirm and expand our knowledge of solid-phase chemolithotrophic Fe(II) oxidation on Earth, and bolster the concept of mineral-associated Fe(II) oxidation as a potential basis for microbial life on other terrestrial planets.

4 Institutions
3 Teams
2 Publications
0 Field Sites
Field Sites

Project Progress

Aqueous and certain solid-phase ferrous iron (Fe(II)) compounds can serve as an electron donor for chemolithotrophic iron-oxidizing microorganisms under both oxic and anoxic conditions (Emerson et al., 2010). It may also have played a role in past (and possibly, present) life on Mars, whose crust has a relatively rich content of Fe(II)-bearing silicate minerals (e.g. ultramafic basalt rocks) (McSween et al., 2009), as well as Fe(II)-bearing phyllosilicate minerals formed during weathering of primary silicates (Bishop et al., 2008; Carter et al., 2010; Fairen et al., 2010). Phyllosilicates have a unique potential to preserve organic biosignatures on Mars (Michalski et al., 2010; Orofino et al., 2010), and organisms capable of gaining metabolic energy from oxidation of Fe(II)-bearing phyllosilicates would be logical candidates for preservation in clay mineral assemblages. To date, with a few notable exceptions (Shelobolina et al., 2003; Weber et al., 2001), virtually all studies of chemolithotrophic Fe(II)-oxidizing microorganisms have focused on oxidation of aqueous Fe(II) under low oxygen conditions, or with nitrate as the electron acceptor (Emerson et al., 2010). Recent NAI-supported studies at UW have revealed the potential for chemolithotrophic oxidation of primary and secondary Fe(II)-bearing phyllosilicates (Shelobolina et al., 2011c), as well as Fe(II)-containing basalt glass (Honma and Roden, 2010). Understanding the physiological and phylogenetic properties of terrestrial Fe(II)-phyllosilicate oxidizing organisms is a prerequisite for evaluating whether or not this mode of energy metabolism could have been active on Mars.

This study examined novel Fe(II)-phyllosilicate oxidizing microorganisms isolated from two different subsurface sedimentary environments: Miocene-age Columbia River basin sediments (Ringold Formation) near Hanford in eastern Washington; and subsoils from Shoveler’s Sink, a nature preserve west of Madison in southern Wisconsin.

Miocene-age subsurface deposits near Hanford in eastern WA (left), and hydromorphic subsoils at Shoveler’s Sink near Madison, WI (right).

The Fe(II)-oxidizing bacteria (FeOB) were recovered using the Fe(II)-bearing primary phyllosilicate mineral biotite as an energy source. Both conventional enrichment culturing and a novel in situ incubation technique (“i-chips”) (Bollmann et al., 2010; Nichols et al., 2010) were employed for this purpose (Shelobolina et al., 2011a; Shelobolina et al., 2011b). Ten different mixotrophic FeOB capable of growth with either organic compounds or Fe(II) were isolated from the initial mixed cultures. Near complete 16SrDNA sequences were obtained for each culture, and their ability to oxidize Fe(II) was confirmed. The isolated FeOB all belong to the Proteobacteria, including Alphaproteobacteria from the genera Bosea, Bradyrhizobium, and Brevundimonas; Betaproteobacteria from the genera Dechloromonas, Leptothrix, and Ralstonia; and Gammaproteobacteria from the genus Serratia. With the exception of Leptothrix, none of the new isolates has been previously identified as a Fe(II)-oxidizing chemolithotroph, although Bradyrhizobium is known to be capable of chemolithotrophic growth with hydrogen (Neal et al., 1983) or thiosulfate (Masuda et al., 2010). These results suggest that FeOB which specialize in oxidation of insoluble Fe(II)-bearing phyllosilicates are distinct from well-recognized aqueous Fe(II)-oxidizing FeOB such as Gallionella, Sideroxydans, Leptothrix, and Sphaerotilus (Emerson et al., 2010). These results significantly expand our knowledge of the diversity of FeOB, and provide a wealth of new targets for genomics-based analysis of the evolutionary relationships, molecular mechanisms, and environmental regulation of solid-phase Fe(II)-oxidizing chemolithotrophs.


Bishop, J.L., Dobrea, E.Z.N., McKeown, N.K., Parente, M., Ehlmann, B.L., Michalski, J.R., Milliken, R.E., Poulet, F., Swayze, G.A., Mustard, J.F., Murchie, S.L., Bibring, J.P., 2008. Phyllosilicate diversity and past aqueous activity revealed at Mawrth Vallis, Mars. Science 321, 830-833.
Bollmann, A., Palumbo, A.V., Lewis, K., Epstein, S.S., 2010. Isolation and physiology of bacteria from contaminated subsurface sediments. Appl. Environ. Microbiol. 76, 7413-7419.
Carter, J., Poulet, F., Bibring, J.P., Murchie, S., 2010. Detection of hydrated silicates in crustal outcrops in the Northern Plains of Mars. Science 328, 1682-1686.
Emerson, D., Fleming, E.J., McBeth, J.M., 2010. Iron-oxidizing bacteria: an environmental and genomic perspective. Annu. Rev. Microbiol. 64, 561-583.
Fairen, A.G., Chevrier, V., Abramov, O., Marzo, G.A., Gavin, P., Davila, A.F., Tornabene, L.L., Bishop, J.L., Roush, T.L., Gross, C., Kneissl, T., Uceda, E.R., Dohm, J.M., Schulze-Makuch, D., Rodriguez, J.A.P., Amils, R., McKay, C.P., 2010. Noachian and more recent phyllosilicates in impact craters on Mars. Proc. Natl. Acad. Sci. U.S.A. 107, 12095-12100.
Honma, A., Roden, E.E., 2010. Chemolithotrophic microbial oxidation of basalt glass. Astrobiology Science Conference 2010, Abstract 5367.
Masuda, S., Eda, S., Ikeda, S., Mitsui, H., Minamisawa, K., 2010. Thiosulfate-dependent chemolithoautotrophic growth of Bradyrhizobium japonicum. Appl. Environ. Microbiol. 76, 2402-2409.
McSween, H.Y., Taylor, G.J., Wyatt, M.B., 2009. Elemental composition of the Martian crust. Science 324, 736-739.
Michalski, J.R., Bibring, J.P., Poulet, F., Loizeau, D., Mangold, N., Dobrea, E.N., Bishop, J.L., Wray, J.J., McKeown, N.K., Parente, M., Hauber, E., Altieri, F., Carrozzo, F.G., Niles, P.B., 2010. The Mawrth Vallis region of Mars: A potential landing site for the Mars Science Laboratory (MSL) mission. Astrobiology 10, 687-703.
Neal, J.L., Allen, G.C., Morse, R.D., Wolf, D.D., 1983. Anaerobic nitrate-dependent chemolithotrophic growth by Rhizobium japonicum. Can. J. Microbiol. 29, 316-320.
Nichols, D., Cahoon, N., Trakhtenberg, E.M., Pham, L., Mehta, A., Belanger, A., Kanigan, T., Lewis, K., Epstein, S.S., 2010. Use of i-chip for high-rhroughput in situ cultivation of “uncultivable” microbial species. Appl. Environ. Microbiol. 76, 2445-2450.
Orofino, V., Blanco, A., D’Elia, M., Licchelli, D., Fonti, S., Marzo, G.A., 2010. Study of terrestrial fossils in phyllosilicate-rich soils: Implication in the search for biosignatures on Mars. Icarus 208, 202-206.
Shelobolina, E., Xiong, M.Y., Kennedy, D.W., Roden, E.E., 2011a. Microbial agents of Fe-phyllosilicate redox metabolism in Hanford 300 Area sediments. Appl. Environ. Microbiol. Manuscript in preparation.
Shelobolina, E., Xiong, M.Y., Roden, E.E., 2011b. Isolation of iron-cycling organisms from an illite-smectite rich subsoil. Frontiers in Chemical Microbiology Manuscript in preparation.
Shelobolina, E.S., Blothe, M., Xu, H., Konishi, H., Roden, E., 2011c. Microbial oxidation of Fe(II)-bearing phyllosilicate minerals by a chemolithoautotrophic Fe(II)-oxidizing, nitrate-reducing enrichment culture Manuscript in prep.
Shelobolina, E.S., Gaw-VanPraagh, C., Lovley, D.R., 2003. Use of ferric and ferrous iron containing minerals for respiration by Desulfitobacterium frappieri. Geomicrobiol. J. 20, 143-156.
Weber, K.A., Picardal, F.W., Roden, E.E., 2001. Microbially-catalyzed nitrate-dependent oxidation of biogenic solid-phase Fe(II) compounds. Environ. Sci. Technol. 35, 1644-1650.

    Evgenya Shelobolina Evgenya Shelobolina
    Project Investigator
    Eric Roden

    Mai Yia Xiong
    Graduate Student

    Objective 3.2
    Origins and evolution of functional biomolecules

    Objective 5.1
    Environment-dependent, molecular evolution in microorganisms