Susan Glasauer

Controls on Fe reduction and mineral formation by a subsurface bacterium

Abstract The reductive dissolution of FeIII (hydr)oxides by dissimilatory iron-reducing bacteria (DIRB) could have a large impact on sediment genesis and Fe transport. If DIRB are able to reduce FeIII in minerals of high structural order to carry out anaerobic respiration, their range could encompass virtually every O2-free environment containing FeIII and adequate conditions for cell growth. Previous studies have established that Shewanella putrefaciens CN32, a known DIRB, will reduce crystalline Fe oxides when initially grown at high densities in a nutrient-rich broth, conditions that poorly model the environments where CN32 is found. By contrast, we grew CN32 by batch culture solely in a minimal growth medium. The stringent conditions imposed by the growth method better represent the conditions that cells are likely to encounter in their natural habitat. Furthermore, the expression of reductases necessary to carry out dissimilatory Fe reduction depends on the method of growth. It was found that under anaerobic conditions CN32 reduced hydrous ferric oxide (HFO), a poorly crystalline FeIII mineral, and did not reduce suspensions containing 4 mM FeIII in the form of poorly ordered nanometer-sized goethite (α-FeOOH), well-ordered micron-sized goethite, or nanometer-sized hematite (α-Fe2O3) crystallites. Transmission electron microscopy (TEM) showed that all minerals but the micron-sized goethite attached extensively to the bacteria and appeared to penetrate the outer cellular membrane. In the treatment with HFO, new FeII and FeIII minerals formed during reduction of HFO-Fe in culture medium containing 4.0 mmol/L Pi (soluble inorganic P), as observed by TEM with energy-dispersive X-ray spectroscopy, selected area electron diffraction, and X-ray diffraction. The minerals included magnetite (Fe3O4), goethite, green rust, and vivianite [Fe3(PO4)2 · 8H2O]. Vivianite appeared to be the stable end product and the mean coherence length was influenced by the rate of FeIII reduction. When Pi was 0.4 mol/L under otherwise identical conditions, goethite was the only mineral observed to form, and less Fe2+ was produced overall. Hence, the ability of DIRB to reduce Fe (hydr)oxides may be limited when the bacteria are grown under nutrient-limited conditions, and the minerals that result depend on the vigor of FeIII reduction.

Sorption of Fe (Hydr)Oxides to the Surface of Shewanella putrefaciens: Cell-Bound Fine-Grained Minerals Are Not Always Formed De Novo

ABSTRACT Shewanella putrefaciens, a gram-negative, facultative anaerobe, is active in the cycling of iron through its interaction with Fe (hydr)oxides in natural environments. Fine-grained Fe precipitates that are attached to the outer membranes of many gram-negative bacteria have most often been attributed to precipitation and growth of the mineral at the cell surface. Our study of the sorption of nonbiogenic Fe (hydr)oxides revealed, however, that large quantities of nanometer-scale ferrihydrite (hydrous ferric oxide), goethite (α-FeOOH), and hematite (α-Fe2O3) adhered to the cell surface. Attempts to separate suspensions of cells and minerals with an 80% glycerin cushion proved that the sorbed minerals were tightly attached to the bacteria. The interaction between minerals and cells resulted in the formation of mineral-cell aggregates, which increased biomass density and provided better sedimentation of mineral Fe compared to suspensions of minerals alone. Transmission electron microscopy observations of cells prepared by whole-mount, conventional embedding, and freeze-substitution methods confirmed the close association between cells and minerals and suggested that in some instances, the mineral crystals had even penetrated the outer membrane and peptidoglycan layers. Given the abundance of these mineral types in natural environments, the data suggest that not all naturally occurring cell surface-associated minerals are necessarily formed de novo on the cell wall.

Reduction of Actinides and Fission Products by Fe(III)-Reducing Bacteria

Microbial metabolism plays a pivotal role in controlling the solubility and mobility of radionuclides in waters contaminated by nuclear waste. The distribution and activity of dissimilatory Fe(III)-reducing bacteria are of particular importance because they can alter the solubility of radionuclides via direct enzymatic reduction or by indirect mechanisms catalyzed via a range of electron shuttling compounds. Using a combination of the techniques of microbiology, biochemistry, and molecular biology, we have characterized the mechanisms of electron transfer to key radionuclides by Fe(III)-reducing bacteria. The mechanisms of enzyme-mediated reduction of problematic actinides, principally U(VI) but including Pu(IV) and Np(V), are described in this review. In addition, the mechanisms by which the fission product Tc(VII) is reduced are also discussed. Direct enzymatic reductions of Tc(VII), catalyzed by microbial hydrogenases, are described along with indirect mechanisms catalyzed by microbially produced Fe(II). Finally, we describe new results that demonstrate the transfer of electrons from biogenic U(IV) to Tc(VII), leading to coprecipitation of Tc(IV) and U(IV), and opening the way for treatment of liquid wastes cocontaminated with both uranium and technetium in one step.

INHIBITION OF SINTERING BY Si DURING THE CONVERSION OF Si-RICH FERRIHYDRITE TO HEMATITE

The distribution and chemical state of Si in a synthetic 2-line ferrihydrite with a Si/(Si + Fe) molar ratio of 0.11 was studied. Heat treatment under oxidizing conditions shows that Si-rich ferrihydrite is stable to 400°C. The transformation to hematite and the formation of a polymerized amorphous-silica phase occur at 850°C. At this temperature, the specific surface area decreases greatly and the average pore diameter increases, which is indicative of sintering. Heating under severe reducing conditions causes a segregation of Si from Fe and results in a mixture of elemental Fe and SiO2. Surface and structural data suggest that Si is located near the particle surface where it limits the rearrangement of Fe octahedra to form hematite.

Mixed-Valence Cytoplasmic Iron Granules Are Linked to Anaerobic Respiration

ABSTRACT Intracellular granules containing ferric and ferrous iron formed in Shewanella putrefaciens CN32 during dissimilatory reduction of solid-phase ferric iron. It is the first in situ detection at high resolution (150 nm) of a mixed-valence metal particle residing within a prokaryotic cell. The relationship of the internal particles to Fe(III) reduction may indicate a respiratory role.

The dynamic nature of bacterial surfaces: implications for metal-membrane interaction.

Bacterial envelopes are chemically complex, diverse structures. Chemical and physical influences from cellular microenvironments force lipids, proteins, and sugars to organize dynamically. This constant reorganization serves to maintain compartmentalization and function, but also affects the influence of charged functional groups that drive electrochemical interactions with metal ions. The interactions of metal species with cell walls are of particular interest because (i) metals must be taken up or excluded to maintain cell function, and (ii) electrochemical interactions between charged metals and anionic ligands are inevitable. In this review we explore the associations of metals with metal-reactive ligands found within bacterial envelopes, and outward to include those within biofilm matrics. The mechanisms that underpin metal binding to these ligands have not been well considered with respect to the dynamic organization of the biological structures themselves. Bacteria respond sensitively and rapidly to growth environment with de novo syntheses of chemical constituents, which can impact metal interactions. We discuss causes of membrane chemical variability as observed in laboratory experiments, and offer consequences for this adaptability in natural settings. The structural impacts of metal ion associations with bacterial envelopes are often overlooked. This review explores how dynamic bacterial surface chemistry influences metal binding and, in turn, how metal ions impact membrane organization in laboratory and natural conditions.

Constraints on the Preservation of Ferriferous Microfossils

The taphonomy of ancient microbial communities is understood via rare windows of microbes that are successfully entombed and preserved in the fossil record. Laboratory investigations using live bacterial cultures, combined with observations on microfossils preserved in ferriferous oolitic beds formed around 161 million years ago, were used to constrain conditions that are conducive to preservation by mineralization. Lab experiments confirmed redox conditions in conjunction with bacterial Fe metabolism as the critical parameter. When Fe was mobilized during the reductive dissolution of ferric hydroxide in laboratory studies, bacteria did not sorb sufficient amounts of ferriferous phases to result in casts. By contrast, iron-oxidizing bacteria sorbed sufficient ferric hydroxide to entomb the cells, although there was variation in Fe oxide sorption within a single culture. The comparison of information from lab and field investigations opens an avenue to infer processes of preservation by mineralization in a geological time frame.

Changes in Shewanella putrefaciens CN32 Membrane Stability upon Growth in the Presence of Soluble Mn(II), V(IV), and U(VI)

In natural reducing environments, such as anoxic sediments and soils, bacteria may be exposed to high concentrations of soluble transition metals. The aim of this study was to identify physiological and biochemical adaptations of Shewanella putrefaciens CN32 membranes to soluble Mn(II), V(IV), and U(VI). We assessed responses of CN32 to these metals, in aerobic and anaerobic cultures, by means of membrane fluidity and fatty acid composition assays. During aerobic growth, all metals had a stabilizing effect on fluidity, while under anoxic conditions this was only observed for bacteria treated with U(VI). Membrane gel-to-fluid phase transition temperatures were higher under anaerobic conditions and were not affected by the metal treatments. Fatty acid desaturation demonstrated linear correlation with significant increases in membrane fluidity, despite metal treatments that did not significantly alter fatty acid chemistry. Scanning transmission X-ray microscopy (STXM) and near-edge X-ray absorption fine structure spectroscopy (NEXAFS) at Mn 2p- and V 2p-edges revealed that both Mn(II) and V(IV) were associated with CN32 membranes, with V(IV) associating as VO2+ under anoxic conditions only. The results of this study indicate that the bacterial growth environment greatly impacts membrane chemistry and stability, with overall implications for in vitro as well as in situ studies. Supplemental materials are available for this article. Please go to the publishers online edition of Geomicrobiology Journal to view the supplemental file.

Removal of sewage phosphorus by adsorption and mineral precipitation, with recovery as a fertilizing soil amendment

Clear sand adsorbs 15-35% total phosphorus (P) from septic tank effluent, but P is mobilized when low-P effluent is applied. Amorphous P compounds formed by alkali aluminate chemical addition may also be subject to leaching. Crystalline mineralization is the desired end effect that isolates P thoroughly from the water resource. Using new low-energy iron electrochemistry (EC-P process), dissolved ferrous iron reacts with sewage phosphate ions (PO4) and precipitates onto filtration medium as vivianite [Fe3(PO4)2·8H2O], as identified by scanning electron microscopy and X-ray diffraction and predicted from Eh-pH-aHPO42- phase relations. Removal rates of 90-99% in sand, soil and synthetic foam filters are obtained. The precipitation of vivianite demonstrates that P can be immobilized quickly and without intermediary adsorption phases, as with Fe-rich soils. Vitreous silicate material (VSM) or rockwool that traps and precipitates mineral P after EC-P treatment was investigated as a means of P reuse as a fertilizing soil amendment. Comparative soil leaching and growth studies using corn plants demonstrate that the VSM alone reduces P losses from soils, and that VSM which has received EC-P effluent is equivalent to or better than commercial superphosphate fertilizer.

Petrology, Elemental and Isotope Geochemistry and Geomicrobiology of Carbonate Infillings and Biofilms Lining Cracks Below the Neoproterozoic (Sturtian) Cap Carbonate in the Mirbat Inlier, Southernmost Oman

Cryptic biofilms line cracks in granite basement below a Neoproterozoic cap carbonate in southern Oman. Their depth in the narrow cracks indicates that they grew in a dark non-photic environment, and signs of upward flow within the cracks suggest that they may have grown in waters expelled from depth, which may not have been in isotopic equilibrium with contemporary ocean waters. This study tested this hypothesis and disproved it. The biofilms and associated detrital carbonate were precipitated in isotopic equilibrium with contemporary seawater. The study also confirms the 13C secular trends previously observed and attributed to seawater changes during post-glacial transgression and accumulation of the cap carbonate.