2009. "Electron donor-dependent radionuclide reduction and nanoparticle formation by Anaeromyxobacter dehalogenans strain 2CP-C." Environmental Microbiology 11(2):534-543. Abstract Anaeromyxobacter dehalogenans strain 2CP-C can rapidly reduce U(VI) or Tc(VII) to U(IV)O2(s) or Tc(IV)O2(S) using either acetate or H2 as an electron donor source. Kinetic studies reveal that the H2-driven reduction of either U(VI) or Tc(VII) is faster than the acetate-driven reduction. The sub-cellular localization of reduced UO2 is extracellular while TcO2 nanoparticles are both periplasmic and extracellular. While electron donor-specific differences in UO2 nanoparticle aggregate size were observed, the association of UO2 nanoparticles with an exopolymeric substance (EPS) was observed and found to be independent of electron donor source. Electron donor-specific localization differences were not observed in cells incubated with Tc(VII). These finding have direct implications on immobilization strategies for highly soluble radionuclide contaminants in subsurface waters and the development of microbially assisted biostimulation efforts.
2009. "Copper isotope fractionation in acid mine drainage." Geochimica et Cosmochimica Acta 73(5):1247-1263. Abstract We surveyed the Cu isotopic composition of primary minerals and stream water affected by acid mine drainage in a mineralized watershed located in southwestern Colorado, USA. The 65Cu values (based on 65Cu/63Cu) of local enargite (65Cu = -0.01 ± 0.10‰) and chalcopyrite (65Cu = 0.16 ± 0.10‰) are within the general range of previously reported values for terrestrial primary Cu sulfides (-1 < 65Cu < 1). These mineral samples show lower 65Cu values than stream waters (65Cu = 1.36 - 1.74 ± 0.10‰), with an average isotopic fractionation (quantified as ∆aq-mino = 65Cuaq – 65Cu mino, where Cuaq is leached Cu and Cu mino is the original mineral) of 1.60 ± 0.14‰ and 1.43 ± 0.14‰ for enargite and chalcopyrite, respectively. To interpret this field survey, we simulated enargite and chalcopyrite leaching in batch experiments and found that, as in the field, leached Cuaq is isotopically enriched relative to primary minerals when microorganisms are absent (average ∆aq-min = 0.94 ± 0.14‰ for enargite, 1.18 ± 0.14‰ for chalcopyrite). Leaching of minerals in the presence of A. ferrooxidans results in smaller average fractionation in the opposite direction for chalcopyrite (∆aq-min = -0.57 ± 0.14‰) and no apparent fractionation for enargite (∆aq-min = 0.10 ± 0.14‰). The isotope effect during release of Cu from leaching minerals is inferred to be the same under both abiotic and biotic conditions. However, preferential association of isotopically enriched Cuaq with A. ferrooxidans cells, observed under TEM to occur as both localized precipitates around cells and Cu inside cells, is inferred to cause isotopic depletion of Cuaq in biotic experiments relative to abiotic experiments. Our results show indications of isotopic signatures of both abiotic chalcopyrite and enargite dissolution. Such signatures will be useful for AMD remediation and ore prospecting purposes.
2008. " Direct Involvement of Type II Secretion System in Extracellular Translocation of Shewanella Oneidensis Outer Membrane Cytochromes MtrC and OmcA." Journal of Bacteriology 190(15):5512-5516. doi:10.1128/JB.00514-08 Abstract Outer membrane decaheme c-type cytochromes MtrC and OmcA of Shewanella oneidensis MR-1 are extracellular lipoproteins important for dissimilatory reduction of solid metal (hydr)oxides during anaerobic respiration. To investigate the roles of type II secretion system (T2S) in translocation of MtrC and OmcA across outer membrane, we measured the effects of deleting two T2S genes, gspD and gspG, on the secretion of MtrC and OmcA when cells were grown under anaerobic conditions. Deletion of gspD or gspG resulted in slightly yellowish supernatants, different from the pink supernatant of wild type (wt). Comparative proteomic analyses revealed that, although MtrC, OmcA and NrfA, a periplasmic nitrite reductase, were present the supernatants of wt and ΔgspD mutant, their peptides counts were much lower in ΔgspD than in wt. Subsequent analyses with heme-staining and Western blot not only confirmed that deletion of gspD or gspG reduced the abundances of MtrC and OmcA in the supernatants, but also revealed that the deletions consequently increased their abundances inside the cells. Complementation of ΔgspG mutant with functional GspG could reverse the effects of deleting gspG on the colors of the supernatants and the abundances of MtrC and OmcA. In contrast, Western results showed that the abundance of NrfA was reduced in the supernatant and the cells of ΔgspD mutant, suggesting that reduced NrfA in the periplasm, where MtrC and OmcA were accumulated, contributed to its reduction in the supernatant. Thus, our results demonstrate at the first time that T2S facilitates translocation of MtrC and OmcA across outer membrane.
2008. "Comparison of uranium(VI) removal by Shewanella oneidensis MR-1 in flow and batch reactors." Water Research 42(12):2993-3002. doi:10.1016/j.watres.2008.04.003 Abstract To better understand the interactions among metal contaminants, nutrients, and microorganisms in subsurface under fracture-flow conditions, iron-reducing biofilms (pure cultures of Shewanella oneidensis MR-1) were grown in six fracture flow reactors (FFRs) of different geometries. The spatial and temporal distribution of nutrients, contaminant, and bacteria were examined using a tracer dye (brilliant blue FCF) and microscopy. The results showed that plugging by bacterial cells depended on the geometry of the reactor; and iron-reducing biofilms grown in FFRs had a definite U(VI)-reduction capacity. To find out the U(VI)-reduction capacity of iron-reducing biofilms, batch experiments of U(VI) reduction were performed in repetitive addition mode. U(VI)-reduction rates of stationary phase grown iron-reducing cultures with and without spent medium decreased after each U(VI) addition. At the end of the fourth U(VI)-addition, stationary phase iron-reducing cultures treated with U(VI) with and without spent medium yielded grey and black precipitates, respectively. These grey and black U precipitates were analyzed using High Resolution-Transmission Electron Microscopy, Energy-dispersive X-ray spectroscopy, and X-ray diffraction. Data for randomly selected area of black and grey U precipitates showed that reduced U particles (3-6 nm) were crystalline and amorphous in nature, respectively. This information obtained in this study could be used to develop substrate addition strategies for metal immobilization in subsurface fracture flow systems.
2008. "Isolation and characterization of an NAD+-degrading bacterium PTX1 and its role in chromium biogeochemical cycle." Biodegradation 19(3):417-424. doi:10.1007/s10532-007-9147-1 Abstract Microorganisms can reduce toxic chromate to less toxic trivalent chromium [Cr(III)]. Besides Cr(OH)3 precipitates, some soluble organo-Cr(III) complexes are readily formed upon microbial, enzymatic, and chemical reduction of chromate. However, the biotransformation of the organo-Cr(III) complexes has not been characterized. We have previously reported the formation of a nicotinamide adenine dinucleotide (NAD+)-Cr(III) complex after enzymatic reduction of chromate. Although the NAD+-Cr(III) complex was stable under sterile conditions, microbial cells were identified as precipitates in a non-sterile NAD+-Cr(III) solution after extended incubation. The most dominant bacterium PTX1 was isolated and assigned to Leifsonia genus by phylogenetic analysis of 16S rRNA gene sequence. PTX1 grew slowly on NAD+ with a doubling time of 17 h, and even more slowly on the NAD+-Cr(III) complex with an estimated doubling time of 35 days. The slow growth suggests that PTX1 passively grew on trace NAD+ dissociated from the NAD+-Cr(III) complex, facilitating further dissociation of the complex and formation of Cr(III) precipitates. Thus, organo-Cr(III) complexes might be an intrinsic link of the chromium biogeochemical cycle; they can be produced during chromate reduction and then further mineralized by microorganisms.
2008. "Hydrogenase- and Outer Membrane c-Type Cytochrome-Facilitated Reduction of Technetium(VII) by Shewanella oneidensis MR-1." Environmental Microbiology 10(1):125-136. doi:10.1111/j.1462-2920.2007.01438.x Abstract Pertechnetate, 99Tc(VII)O4-, is a highly mobile radionuclide contaminant at U.S. Department of Energy sites that can be enzymatically reduced by a range of anaerobic and facultatively anaerobic microorganisms, including Shewanella oneidensis MR-1, to poorly soluble Tc(IV)O2(s). In other microorganisms, Tc(VII)O4- reduction is generally considered to be catalyzed by hydrogenase. Here, we provide evidence that although the NiFe hydrogenase of MR-1 was involved in the H2-driven reduction of Tc(VII)O4- (presumably through a direct coupling of H2 oxidation and Tc(VII) reduction), the deletion of both hydrogenase genes did not completely eliminate the ability of MR-1 to reduce Tc(VII). With lactate as the electron donor, mutants lacking the outer membrane c-type cytochromes MtrC and OmcA or the proteins required for the maturation of c-type cytochromes were defective in reducing Tc(VII) to nanoparticulate TcO2·nH2O(s) relative to MR-1 or a NiFe hydrogenase mutant. In addition, reduced MtrC and OmcA were oxidized by Tc(VII)O4-, confirming the capacity for direct electron transfer from these OMCs to TcO4-. c-Type cytochrome-catalyzed Tc(VII) reduction could be a potentially important mechanism in environments where organic electron donor concentrations are sufficient to allow this reaction to dominate.
2008. "Characterization of uraninite nanoparticles produced by Shewanella oneidensis MR-1 ." Geochimica et Cosmochimica Acta 72(20):4901-4915. doi:10.1016/j.gca.2008.07.016 Abstract The reduction of uranium(VI) by Shewanella oneidensis MR-1 was studied to examine the effects of bioreduction kinetics and background electrolyte on the physical properties and reactivity to re-oxidation of the biogenic uraninite, UO2(s). Bioreduction experiments were conducted with uranyl acetate as the electron acceptor and sodium lactate as the electron donor under resting cell conditions in a 30 mM NaHCO3 buffer, and in a PIPES-buffered artificial groundwater (PBAGW). MR-1 was cultured in batch mode in a defined minimal medium with a specified air-to-medium volume ratio such that electron acceptor (O2) limiting conditions were reached just when cells were harvested for subsequent experiments. The rate of U(VI) bioreduction was manipulated by varying the cell density and the incubation temperature (1.0 _ 108 cell ml_1 at 20 _C or 2.0 _ 108 cell ml_1 at 37 _C) to generate U(IV) solids at ‘‘fast” and ‘‘slow” rates in the two different buffers. The presence of Ca in PBAGW buffer altered U(VI) speciation and solubility, and significantly decreased U(VI) bioreduction kinetics. High resolution transmission electron microscopy was used to measure uraninite particle size distributions produced under the four different conditions. The most common primary particle size was 2.9–3.0 nm regardless of U(VI) bioreduction rate or background electrolyte. Extended X-ray absorption fine-structure spectroscopy was also used to estimate uraninite particle size and was consistent with TEM results. The reactivity of the biogenic uraninite products with dissolved oxygen was tested, and neither U(VI) bioreduction rate nor background electrolyte had any statistical effect on oxidation rates. With MR-1, uraninite particle size was not controlled by the bioreduction rate of U(VI) or the background electrolyte. These results for MR-1, where U(VI) bioreduction rate had no discernible effect on uraninite particle size or oxidation rate, contrast with our recent research with Shewanella putrefaciens CN32, where U(VI) bioreduction rate strongly influenced both uraninite particle size and oxidation rate. These two studies with Shewanella species can be viewed as consistent if one assumes that particle size controls oxidation rates, so the similar uraninite particle sizes produced by MR-1 regardless of U(VI) bioreduction rate would result in similar oxidation rates. Factors that might explain why U(VI) bioreduction rate was an important control on uraninite particle size for CN32 but not for MR-1 are discussed.
2007. "Reduction of Pertechnetate [Tc(VII)] by Aqueous Fe(II) and the Nature of Solid Phase Redox Products." Geochimica et Cosmochimica Acta 71(9):2137-2157. doi:doi:10.1016/j.gca.2006.10.025 Abstract The subsurface behaviour of 99Tc, a contaminant resulting from nuclear fuels reprocessing, is strongly dependent on its valence (e.g., IV or VII). Abiotic reduction of soluble Tc(VII) by Fe(II)(aq) in pH 6-8 solutions was investigated under strictly anoxic conditions using an oxygen trap (<7.5 10-9atm O2) in the absence of atmospheric or aqueous carbonate. The reduction kinetics were strongly pH dependent. Complete and rapid reduction of Tc(VII) to a precipitated Tc(IV)/Fe form was observed when 11 µmol/L of Tc(VII) was reacted with 0.4 mmol/L Fe(II) at pH 7.0 and 8.0, while no significant reduction was observed over 1 month at pH 6.0. Experiments conducted at pH 7.0 with Fe(II)(aq) = 0.05-0.8 mmol/L further revealed that Tc(VII) reduction was a combination of homogeneous and heterogeneous reaction. The heterogeneous reaction was more rapid, but not quantified. The kinetics of homogeneous reduction were slow at pH 7, but increased dramatically at pH 8, and correlated with the concentration of Fe(OH)+ and Fe(OH)2o(aq). Wet chemical and Fe-x-ray absorption near edge spectroscopy measurements (XANES) indicated that both Fe(II) and Fe(III) were present in the Fe/Tc(IV) redox reaction products. 57Fe- Mössbauer, extended x-ray adsorption fine structure (EXAFS), and transmission electron microscopy (TEM) measurements on the solid phase redox products that contained 13-15% Tc indicated that they were poorly ordered and dominated by Fe(II)-containing ferrihydrite with minor magnetite. Tc(IV) exhibited homogeneous spatial distribution within the precipitates. According to Tc-EXAFS measurements and structural modeling, its molecular environment was consistent with a Tc2O10 octahedral dimer bound in bidentate edge-sharing mode to an octahedral FeO6 surface or vacancy site in ferrihydrite. The precipitate maintained Tc(IV)aq concentrations that were slightly below those in equilibrium with amorphous Tc(IV)O2•nH2O(s). The oxidation rate of sorbed Tc(IV) in the Fe/Tc precipitate was considerably slower than Tc(IV)O2•nH2O(s) as a result of its intraparticle/intragrain residence. It is suggested that precipitates of this nature may form in anoxic sediments or groundwaters, and that the intraparticle residence of sorbed/precipitated Tc(IV) may limit 99Tc remobilization upon the return of oxidizing conditions.
2007. "In Vitro Cell Culture Infectivity Assay for Human Noroviruses." Emerging Infectious Diseases 13(3):396-403. Abstract Human noroviruses (NoV) cause severe, self-limiting gastroenteritis that typically lasts 24 - 48 hours. The true nature of NoV pathogenesis remains unknown due to the lack of suitable tissue culture or animal models. Here we show, for the first time, that NoV can infect and replicate in an organoid, three-dimensional (3-D) model of human small intestinal epithelium (INT-407). Cellular differentiation for this model was achieved by growing the cells in 3-D on porous collagen I-coated microcarrier beads under conditions of physiological fluid shear in rotating wall vessel bioreactors. Microscopy, PCR, and fluorescent in-situ hybridization were employed to provide evidence of NoV infection. CPE and norovirus RNA was detected at each of the five cell passages for both genogroup I and II viruses. Our results demonstrate that the highly differentiated 3-D cell culture model can support the natural growth of human noroviruses, whereas previous attempts using differentiated monolayer cultures failed.
2007. "Cell Culture Assay for Human Noroviruses [response]." Emerging Infectious Diseases 13(7):1117-1118. Abstract We appreciate the comments provided by Leung et al., in response to our recently published article “In Vitro Cell Culture Infectivity Assay for Human Noroviruses” by Straub et al. (1). The specific aim of our project was to develop an in vitro cell culture infectivity assay for human noroviruses (hNoV) to enhance risk assessments when they are detected in water supplies. Reverse transcription (RT) qualitative or quantitative PCR are the primary assays for waterborne NoV monitoring. However, these assays cannot distinguish between infectious vs. non-infectious virions. When hNoV is detected in water supplies, information provided by our infectivity assay will significantly improve risk assessment models and protect human health, regardless of whether we are propagating NoV. Indeed, in vitro cell culture infectivity assays for the waterborne pathogen Cryptosporidium parvum that supplement approved fluorescent microscopy assays, do not result in amplification of the environmentally resistant hard-walled oocysts (2). However, identification of life cycle stages in cell culture provides evidence of infectious oocysts in a water supply. Nonetheless, Leung et al.’s assertion regarding the suitability of our method for the in vitro propagation of high titers of NoV is valid for the medical research community. In this case, well-characterized challenge pools of virus would be useful for developing and testing diagnostics, therapeutics, and vaccines. As further validation of our published findings, we have now optimized RT quantitative PCR to assess the level of viral production in cell culture, where we are indeed finding significant increases in viral titer. The magnitude and time course of these increases is dependent on both virus strain and multiplicity of infection. We are currently preparing a manuscript that will discuss these findings in greater detail, and the implications this may have for creating viral challenge pools
2007. "The effect of U(VI) bioreduction kinetics on subsequent reoxidation of biogenic U(IV)." Geochimica et Cosmochimica Acta 71(19):4644-4654. doi:10.1016/j.gca.2007.07.021 Abstract Microbially mediated in situ reduction of soluble U(VI) to insoluble U(IV) (as UO2) has been proposed as a means of preventing the migration of that radionuclide with groundwater, but preventing the oxidative resolubilization of U has proven difficult. We hypothesized that relatively slow rates of U(VI) bioreduction would yield larger UO2 precipitates that would be more resistant to oxidation than those produced by rapid U(VI) bioreduction. We manipulated U(VI) bioreduction rates by varying the density of Shewanella putrefaciens CN32 added to U(VI) containing solutions with lactate as an electron donor. Characterization of biogenic UO2 particles by extended X-ray absorption fine-structure spectroscopy and transmission electron microscopy revealed that UO2 nanoparticles formed by relatively slow rates of U(VI) reduction were larger and more highly aggregated than those formed by relatively rapid U(VI) reduction. UO2 particles formed at various rates were incubated under a variety of abiotically and biologically oxidizing conditions. In all cases, UO2 that was formed by relatively slow U(VI) reduction was oxidized at a slower rate and to a lesser extent than UO2 formed by relatively rapid U(VI) bioreduction, suggesting that the stability of UO2 in situ may be enhanced by stimulation of relatively slow rates of U(VI) reduction.
2007. "Fe-Solid Phase Transformations Under Highly Basic Conditions." Applied Geochemistry 22(9):2054-2064. doi:10.1016/j.apgeochem.2007.04.023 Abstract Hyperalkaline and saline radioactive waste fluids with elevated temperatures from S-SX high-level waste tank farm at Hanford, WA accidentally leaked into sediments beneath the tanks, initiating a series of geochemical processes and reactions whose significance and extent was unknown. Among the most important processes was the dissolution of soil minerals and precipitation of stable secondary phases. The objective of this investigation was to study the release of Fe into the aqueous phase upon dissolution of Fe-bearing soil minerals, and the subsequent formation of Fe rich precipitates. Batch reactors were used to conduct experiments at 50 0C using solutions similar in composition to the waste fluids. Results clearly showed that, similarly to Si and Al, Fe was released from the dissolution of soil minerals (most likely phyllosilicates such as biotite, smectite, and chlorite). The extent of Fe release increased with base concentration and decreased with Al concentration in the contacting solution. The maximum apparent rate of Fe release (0.566 × 10-13 mol m-2 s-1) was measured in the treatment with no Al and a concentration of 4.32 mol L-1 NaOH in the contacting solution. Results from electron microscopy indicated that while Si and Al precipitated together to form feldspathoids in the groups of cancrinite and/or sodalite, Fe precipitation followed a different pathway leading to the formation of hematite and goethite. The newly formed Fe oxy-hydroxides may increase the sorption capacity of the sediments, promote surface mediated reactions such as precipitation and heterogeneous redox reactions, and affect the phase distribution of contaminant and radionuclides.
2007. "Submicron and Nanoscale Inorganic Particles Exploit the Actin Machinery to be Propelled Along Microvillilike Structures into Alveolar Cells." ACS Nano 1(5):463-475. doi:10.1021/nn700149r Abstract The growing commerce in micro- and nanotechnology is expected to increase our exposure to submicron and nanoscale particles. One of the main targets of this exposure are the cells that line the respiratory tract, among them are the alveolar type II epithelial cells that have microvilli at their exposed apical surface. Here we show a pathway by which positively charged inorganic submicron and nanoscale particles take advantage of the actin turnover machinery within filopodia and microvilli-like structures to guide and advance their way into these cells. Our observations bring a new view of how submicron and nanoscale inorganic matter can be assimilated into the cellular environment and take advantage of its machinery. While the pathway that we describe can be exploited for a targeted drug delivery, it also points to properties of submicron or nanoscale particles that should be avoided in order to reduce particle internalization and possible toxicity.
2007. "Uranium immobilization by sulfate-reducing biofilms grown on hematite, dolomite, and calcite." Environmental Science & Technology 41(24):8349-8354. doi:10.1021/es071335k Abstract Biofilms of sulfate-reducing bacteria Desulfovibrio desulfuricans G20 wereused to reduce dissolved U(VI)and subsequently immobilize U(IV) in the presence of uranium-complexing carbonates. The biofilms were grown in three identically operated fixed bed reactors, filled with three types of minerals: one noncarbonate-bearing mineral(hematite) and two carbonate-bearing minerals (calcite and dolomite). The source of carbonates in the reactors filled with calcite and dolomite were the minerals, while in the reactor filled with hematite it was a 10 mM carbonate buffer, pH 7.2, which we added to the growth medium. Our five-month study demonstrated that the sulfate-reducing biofilms grown in all reactors were able to immobilize/reduce uranium efficiently, despite the presence of uranium-complexing carbonates.
2007. "Apoferritin-Templated Synthesis of Encoded Metallic Phosphate Nanoparticle Tags." Analytical Chemistry 79(15):5614-5619. doi:10.1021/ac070086f Abstract Encoded metallic-phosphate nanoparticle tags, with distinct encoding patterns, have been prepared using an apoferritin template. A center-cavity structure as well as the disassociation and reconstructive characteristics of apoferritin at different pH environments provide a facile route for preparing such encoded nanoparticle tags. Encapsulation and diffusion approaches have been investigated during the preparation. The encapsulation approach, which is based on the dissociation and reconstruction of apoferritin at different pHs, exhibits an effective route to prepare such encoded metallic-phosphate nanoparticle tags. The compositionally encoded nanoparticle tag leads to a high coding capacity with a large number of distinguishable voltammetric signals, reflecting the predetermined composition of the metal mixture solution (and hence the nanoparticle composition). Releasing the metal components from the nanoparticle tags at pH 4.6 acetate buffer avoids harsh dissolution conditions, such as strong acids. Such a synthesis of encoded nanoparticle tags, including single-component and compositionally encoded nanoparticle tags, is substantially simple, fast, and convenient compared to that of encoded metal nanowires and semiconductor nanoparticle (CdS, PbS, and ZnS) incorporated polystyrene beads. The encoded metallic-phosphate nanoparticle tags thus show great promise for bioanalytical or product-tracking/identification/protection applications.
2007. "Biogenic Production of Photosensitive Arsenic-Sulfide Nanotubes by Shwanella sp. strain HN-41." Proceedings of the National Academy of Sciences of the United States of America 104(51):20410-20415. Abstract This paper describes the novel production of extensive filamentous, arsenic-sulfide (As-S) nanotubes (20 - 100 nm dia. X ∼30 μm length) resulting from the dissimilatoryreduction of thiosulfate and arsenate by the bacterium Shewanella sp. HN-41. While there have been several reports of bacterial-produced nanowires composed entirely of biological macromolecules, here we report for the first time the biogenic formation ofphotosensitive and electroconductive nanotubes comprised of crystalline As-S and bacterial extracellular polymeric substances (EPS). To our knowledge, there have been no previous reports of the synthesis of such a material either by chemical or biological means. The biogenic As-S nanotubes reported here represent a significant advancement as building blocks for the production of nanodevices because of their high aspect ratios and unique size dependent properties. We characterized the structural evolution of the bacterial As-S nanotubes formed by strain HN-41 using XAFS spectra as well as XRD analyses, as a function of time and extent of bacterial reduction. In addition, we characterized the electrical and photoconductive properties of the As-S nanotubes. Upon aging, the As-S nanotubes behaved as metals and semiconductors in terms of their electrical and photoconductive properties, respectively. These results indicate that the dissimilatory bacterium Shewanella may be an excellent biological tool to bioengineer As-S nanotubes, which may provide useful materials for novel nano- and optoelectronic devices.
2007. "Biogenic formation of photoactive arsenic-sulfide nanotubes by Shewanella sp. strain HN-41 ." Proceedings of the National Academy of Sciences of the United States of America 104(51):20410-20415. doi:10.1073/pnas.0707595104 Abstract Microorganisms facilitate the formation of a wide range of minerals that have unique physical and chemical properties as well as morphologies that are not produced by abiotic processes. Here, we report the production of an extensive extracellular network of filamentous, arsenic-sulfide (As-S) nanotubes (20–100 nm in diameter by 30 µm in length) by the dissimilatory metal-reducing bacterium Shewanella sp. HN-41. The As-S nanotubes, formed via the reduction of As(V) and S2O, were initially amorphous As2S3 but evolved with increasing incubation time toward polycrystalline phases of the chalcogenide minerals realgar (AsS) and duranusite (As4S). Upon maturation, the As-S nanotubes behaved as metals and semiconductors in terms of their electrical and photoconductive properties, respectively. The As-S nanotubes produced by Shewanella may provide useful materials for novel nano- and opto-electronic devices.
2007. "Biogenic mineral production by a novel arsenic-metabolizing thermophilic bacterium from the Alvord Basin, OR." Applied and Environmental Microbiology 73(18):5928-5936. doi:10.1128/AEM.00371-07 Abstract The Alvord Basin in southeast Oregon, USA contains a variety of hydrothermal features, which have never been microbiologically characterized. Murky Pot (61°C, pH 7.1) was selected for this study. Sampling of Murky Pot led to the isolation of a novel arsenic-metabolizing organism (YeAs), which produces an arsenic sulfide mineral known as beta-realgar, a mineral that has not previously been observed as a product of bacterial arsenic metabolism. Our goal was to characterize and identify YeAs based on its phylogenetic, physiological, and morphological characteristics. 16S rRNA gene analysis revealed that YeAs has 98.9% sequence similarity to that of Thermobrachium celere. YeAs was grown on a freshwater medium and could utilize a variety of organic substrates, particularly carbohydrates and organic acids. Optimum growth of the organism was seen at 55ºC, but showed growth at a range of 37° to 75°C. No growth was observed when YeAs was grown under aerobic conditions. Microscopic examination revealed Gram-indeterminate, non-spore forming, rod shaped cells. Electron microscopy and elemental analysis revealed significant arsenic sulfide mineralization of cell walls, and extracellular particulate deposition of arsenic sulfide minerals. YeAs showed no detectable respiratory arsenate reductase; however, the organism did display significant detoxification arsenate reductase activity. The phylogenetic, physiological, and morphological characteristics of YeAs demonstrate that it is an anaerobic, moderately thermophilic, arsenic-reducing bacterium. This organism and its associated metabolism could have major implications in the search for innovative methods for arsenic waste management and in the search for novel biogenic signatures.
2007. "Crosslinked Enzyme Aggregates in Hierarchically-Ordered Mesoporous Silica: A Simple and Effective Method for Enzyme Stabilization." Biotechnology and Bioenegineering 96(2):210-218. doi:10.1002/bit.21107 Abstract alpha-chymotrypsin (CT) and lipase (LP) were immobilized in hierarchically-ordered mesocellular mesoporous silica (HMMS) in a simple but effective way for the enzyme stabilization, which was achieved by the enzyme adsorption followed by glutaraldehyde (GA) crosslinking. This resulted in the formation of nanometer scale crosslinked enzyme aggregates (CLEAs) entrapped in the mesocellular pores of HMMS (37 nm), which did not leach out of HMMS through narrow mesoporous channels (13 nm). CLEA of alpha-chymotrypsin (CLEA-CT) in HMMS showed a high enzyme loading capacity and significantly increased enzyme stability. No activity decrease of CLEA-CT was observed for two weeks under even rigorously shaking condition, while adsorbed CT in HMMS and free CT showed a rapid inactivation due to the enzyme leaching and presumably autolysis, respectively. With the CLEA-CT in HMMS, however, there was no tryptic digestion observed suggesting that the CLEA-CT is not susceptible to autolysis. Moreover, CLEA of lipase (CLEA-LP) in HMMS retained 30% specific activity of free lipase with greatly enhanced stability. This work demonstrates that HMMS can be efficiently employed as host materials for enzyme immobilization leading to highly enhanced stability of the immobilized enzymes with high enzyme loading and activity.
2007. "Reactive Ballistic Deposition of Porous TiO2 Films: Growth and Characterization." Journal of Physical Chemistry C 111(12):4765-4773. doi:10.1021/jp067641m Abstract Nanoporous, high-surface area films of TiO2 are synthesized by reactive ballistic deposition of titanium metal in an oxygen ambient. Auger electron spectroscopy (AES) is used to investigate the stoichiometric dependence of the films on growth conditions (surface temperature and partial pressure of oxygen). Scanning and transmission electron microscopy show that the films consist of arrays of separated filaments. The surface area and the distribution of binding site energies of the films are measured as functions of growth temperature, deposition angle, and annealing conditions using temperature programmed desorption (TPD) of N2. TiO2 films deposited at 50 K at 70º from substrate normal display the greatest specific surface area of ~100 m2/g. In addition, the films retain greater than 70% of their original surface area after annealing to 600 K. The combination of high surface area and thermal stability suggest that these films could serve as supports for applications in heterogeneous catalysis.
2006. "Proteomic Analysis of Salmonella enterica Serovar Typhimurium Isolated from RAW 264.7 Macrophages: identification of a novel protein that contributes to the replication of serovar Typhimurium inside macrophages." Journal of Biological Chemistry 281:29131-29140. doi:10.1074/jbc.M604640200 Abstract ABSTRACT: To evade host resistance mechanisms, Salmonella enterica serovar Typhimurium (STM), a facultative intracellular pathogen, must alter its proteome following macrophage infection. To identify new colonization and virulence factors that mediate STM pathogenesis, we have isolated STM cells from RAW 264.7 macrophages at various time-points following infection and used a liquid chromatography-mass spectrometry (LC-MS)-based proteomic approach to detect the changes in STM protein abundances. Because host resistance to STM infection is strongly modulated by the expression of a functional host resistant regulator, i.e., natural resistance associated macrophage protein 1 (Nramp1, also called Slc11a1), we have also examined the effects of Nramp1 activity on the changes of STM protein abundances. A total of 315 STM proteins have been identified from isolated STM cells, which are largely house-keeping proteins whose abundances remain relatively constant during the time-course of infection. However, 39 STM proteins are strongly induced after infection, suggesting their involvement in modulating colonization and infection. Of the 39 induced proteins, 6 proteins are specifically modulated by Nramp1 activity, including STM3117, as well as STM3118-3119 whose time-dependent abundance changes were confirmed using Western blot analysis. Deletion of the gene encoding STM3117 resulted in a dramatic reduction in the ability of STM to colonize wild-type RAW 264.7 macrophages, demonstrating a critical involvement of STM3117 in promoting the replication of STM inside macrophages. The predicted function common for STM3117-3119 is biosynthesis and modification of the peptidoglycan layer of STM cell wall, emphasizing their important roles in the colonization of macrophages by Salmonella.
2006. "Metal Reduction and Iron Biomineralization by a Psychrotolerant Fe(III)-Reducing Bacterium, Shewanella sp. Strain PV-4." Applied and Environmental Microbiology 72(5):3236-3244. Abstract A marine psychrotolerant, dissimilatory Fe(III)-reducing bacterium, Shewanella sp. strain PV-4, from the microbial mat at a hydrothermal vent of Loihi Seamount in the Pacific Ocean has been further characterized, with emphases on metal reduction and iron biomineralization. The strain is able to reduce metals such as Fe(III), Co(III), Cr(VI), Mn(IV), and U(VI) as electron acceptors while using lactate, formate, pyruvate, or hydrogen as an electron donor. Growth during iron reduction occurred over the pH range of 7.0 to 8.9, a sodium chloride range of 0.05 to 5%, and a temperature range of 0 to 37°C, with an optimum growth temperature of 18°C. Unlike mesophilic dissimilatory Fe(III)-reducing bacteria, which produce mostly superparamagnetic magnetite (<35 nm), this psychrotolerant bacterium produces well-formed single-domain magnetite (>35 nm) at temperatures from 18 to 37°C. The genome size of this strain is about 4.5 Mb. Strain PV-4 is sensitive to a variety of commonly used antibiotics except ampicillin and can acquire exogenous DNA (plasmid pCM157) through conjugation.
2006. "Metal Reduction and Iron Biomineralization by a Psychrotolerant Fe(III)-Reducing Bacterium, Shewanella sp. Strain PV-4." Applied and Environmental Microbiology 72(5):3236-3244. doi:10.1128/AEM.72.5.3236-3244.2006 Abstract A marine psychrotolerant, dissimilatory Fe(III)-reducing bacterium, Shewanella sp. strain PV-4, from the microbial mat at a hydrothermal vent of Loihi Seamount in the Pacific Ocean has been further characterized, with emphases on metal reduction and iron biomineralization. The strain is able to reduce metals such as Fe(III), Co(III), Cr(VI), Mn(IV), and U(VI) as electron acceptors while using lactate, formate, pyruvate, or hydrogen as an electron donor. Growth during iron reduction occurred over the pH range of 7.0 to 8.9, a sodium chloride range of 0.05 to 5%, and a temperature range of 0 to 37°C, with an optimum growth temperature of 18°C. Unlike mesophilic dissimilatory Fe(III)-reducing bacteria, which produce mostly superparamagnetic magnetite (<35 nm), this psychrotolerant bacterium produces well-formed single-domain magnetite (>35 nm) at temperatures from 18 to 37°C. The genome size of this strain is about 4.5 Mb. Strain PV-4 is sensitive to a variety of commonly used antibiotics except ampicillin and can acquire exogenous DNA (plasmid pCM157) through conjugation.
2006. "c-Type Cytochrome-Dependent Formation of U(IV) Nanoparticles by Shewanella oneidensis ." PloS Biology 4(8):1324-1333. Abstract Modern approaches for bioremediation of radionuclide contaminated environments are based on the ability of microorganisms to effectively catalyze changes in the oxidation states of metals that in turn influence their solubility. Although microbial metal reduction has been identified as an effective means for immobilizing highly-soluble uranium(VI) complexes in situ, the biomolecular mechanisms of U(VI) reduction are not well understood. Here, we show that c-type cytochromes of a dissimilatory metal reducing bacterium, Shewanella oneidensis MR-1 are essential for the reduction of U(VI) and formation of extracelluar UO2 nanoparticles. In particular, the outer membrane (OM) decaheme cytochrome MtrC, previously implicated in Mn(IV) and Fe(III) reduction, directly transferred electrons to U(VI). Additionally, deletions of mtrC and/or omcA significantly affected the in vivo U(VI) reduction rate relative to wild type MR-1. Similar to the wild type, the mutants accumulated UO2 nanoparticles extracellularly to high densities in association with an exopolymeric substance (EPS). In wild type cells, this UO2-EPS matrix exhibited glycocalyx-like properties, contained multiple elements of the OM, polysaccharide, and heme containing proteins. Using a novel combination of methods including synchrotron-based X-ray fluorescence microscopy and high resolution immune-electron microscopy, we demonstrate a close association of the extracellular UO2 nanoparticles with MtrC and OmcA. This is the first study to directly localize the OM-associated cytochromes with EPS, which contains biogenic UO2 nanoparticles. In the environment, such association of UO2 nanoparticles with biopolymers may exert a strong influence on subsequent behavior including susceptibility to oxidation by O2 or transport in soils and sediments.
2006. "Kinetics of Microbial Reduction of Solid Phase U(VI)." Environmental Science and Technology 40(20):6290-6296. Abstract Sodium boltwoodite (NaUO2SiO3OH ∙1.5H2O) was used to assess the kinetics of microbial reduction of solid phase U(VI) by a dissimilatory metal-reducing bacterium (DMRB), Shewanella oneidensis strain MR-1. The bioreduction kinetics was studied with Na-boltwoodite in suspension or within alginate beads. Concentrations of U(VI)tot and cell number were varied to evaluate the coupling of U(VI) dissolution, diffusion, and microbial activity. Batch experiments were performed in a non-growth medium with lactate as electron donor at pH 6.8 buffered with PIPES. Microscopic and spectroscopic analyses with transmission electron microscopy (TEM), energy dispersive spectroscopy (EDS), and laser-induced fluorescence spectroscopy (LIFS) collectively indicated that solid phase U(VI) was first dissolved and diffused out of grain interiors before it was reduced on bacterial surfaces and/or within the periplasm. The kinetics of solid phase U(VI) bioreduction was well described by a coupled model of bicarbonate-promoted dissolution of Na-boltwoodite, intraparticle uranyl diffusion, and Monod type bioreduction kinetics with respect to dissolved U(VI) concentration. The results demonstrated the intimate coupling of biological, chemical, and physical processes in microbial reduction of solid phase U(VI).
2006. "Single Enzyme Nanoparticles in Nanoporous Silica: A Heirachical Approach to Enzyme Stabilization and Immobilization." Enzyme and Microbial Technology 39:474-480. Abstract Single enzyme nanoparticles of alpha-chymotrypsin (SEN-CT), in which each CT molecule is surrounded by a thin polymeric organic/inorganic network, stabilized the CT activity in a shaking condition as well as in a non-shaking condition. Since SEN-CT is soluble in a buffer solution and less than 10 nm in size, SEN-CT could be immobilized in nanoporous silica with an average pore size of 29 nm. Free CT and SEN-CT were immobilized in nanoporous silica (NPS), and nanoporous silica that was first silanized with aminopropyltriethoxysilane (amino-NPS) to generate a positive surface charge. The SEN-CT adsorbed in amino-NPS was more stable than CT immobilized by either adsorption in NPS or covalent bonding to amino-NPS. In shaking conditions, nanoporous silica provided an additional stabilization by protecting SEN-CT from shear stresses. At 22oC with harsh shaking, free, NPS- adsorbed and NPS-covalently-attached CT showed half lives of 1, 62, and 80 h, respectively; whereas SEN-CT adsorbed in amino-NPS showed no activity loss within 12 days. The combination of SENs and nanoporous silica, which makes an active and stable immobilized enzyme system, represents a new structure for biocatalytic applications.
2006. "Electrically Conductive Bacterial Nanowires Produced by Shewanella Oneidensis Strain MR-1 and Other Microorganisms ." Proceedings of the National Academy of Sciences of the United States of America 103(30):11358-11363. Abstract Shewanella oneidensis MR-1 produced electrically conductive pilus-like appendages called bacterial nanowires in direct response to electron-acceptor limitation. Mutants deficient in genes for c-type decaheme cytochromes MtrC and OmcA, and those that lacked a functional Type II secretion pathway displayed nanowires that were poorly conductive. These mutants were also deficient in their ability to reduce hydrous ferric oxide and in their ability to generate current in a microbial fuel cell. Nanowires produced by the oxygenic phototrophic cyanobacterium Synechocystis PCC6803 and the thermophilic, fermentative bacterium Pelotomaculum thermopropionicum reveal that electrically conductive appendages are not exclusive to dissimilatory metal-reducing bacteria and may, in fact, represent a common bacterial strategy for efficient electron transfer and energy distribution.
2005. "Geophysical Imaging of Stimulated Microbial Biomineralization." Environmental Science and Technology 39(19):7592-7600. doi:10.1021/es0504035 Abstract Understanding how microorganisms influence the physical and chemical properties of the subsurface is hindered by our inability to observe microbial dynamics in real time and with high spatial resolution. Here, we investigate the use of noninvasive geophysical methods to monitor biomineralization at the laboratory scale during stimulated sulfate reduction under dynamic flow conditions. Alterations in sediment characteristics resulting from microbe-mediated sulfide mineral precipitation were concomitant with changes in complex resistivity and acoustic wave propagation signatures. The sequestration of zinc and iron in insoluble sulfides led to alterations in the ability of the pore fluid to conduct electrical charge and of the saturated sediments to dissipate acoustic energy. These changes resulted directly from the nucleation, growth, and development of nanoparticulate precipitates along grain surfaces and within the pore space. Scanning and transmission electron microscopy (SEM and TEM) confirmed the sulfides to be associated with cell surfaces, with precipitates ranging from aggregates of individual 3-5 nm nanocrystals to larger assemblages of up to 10-20 ím in diameter. Anomalies in the geophysical data reflected the distribution of mineral precipitates and biomass over space and time, with temporal variations in the signals corresponding to changes in the aggregation state of the nanocrystalline sulfides. These results suggest the potential for using geophysical techniques to image certain subsurface biogeochemical processes, such as those accompanying the bioremediation of metalcontaminated aquifers.
2005. "Reoxidation of Reduced Uranium with Iron(III) (Hydr)Oxides under Sulfate-Reducing Conditions." Environmental Science and Technology 39(7):2059-2066. Abstract In cultures of Desulfovibrio desulfuricansG20 the effects of iron(III) (hydr)oxides (hematite, goethite, and ferrihydrite) on microbial reduction and reoxidation of uranium (U) were evaluated under lactate-limited sulfate-reducing conditions. With lactate present, G20 reduced U(VI) in both 1,4-piperazinediethanesulfonate (PIPES) and bicarbonate buffer. Once lactate was depleted, however, microbially reduced U served as an electron donor to reduce Fe(III) present in iron(III) (hydr)oxides. With the same initial amount of Fe(III) (10 mmol/L) for each iron(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 to 20 mmol of Fe(III)/L, the rate and extent of U(IV) reoxidation increased. Subsequent addition of hematite [15 mmol of 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 X-ray absorption near-edge spectroscopy (XANES) 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. By the same techniques, iron sulfide precipitates were found to have a variable Fe and S stoichiometry and were not associated with cells.
2005. "Uranium removal by sulfate reducing biofilms in the presence of carbonates ." Water Science and Technology 52(7):49-55. Abstract Hexavalent uranium [U(VI)] was immobilized in biofilms composed of the sulfate reducing bacteria (SRB), Desulfovibrio desulfuricans G20. The biofilms were grown in two flat-plate, continuous-flow reactors using lactate as the electron donor and sulfate as the electron acceptor. The growth medium contained uranium U(VI) and the pH was maintained constant using bicarbonate buffer. The reactors were operated for 5 months, and during that time biofilm activity and uranium removal were evaluated. The efficiency of uranium removal strongly depended on the concentration of uranium in the influent, and was estimated to be 30.4% in the reactor supplied with 3 mg/L of U(VI) and 73.9% in the reactor supplied with 30 mg/L of U(VI). TEM and SAED analysis showed that uranium in both reactors accumulated mostly on microbial cell membranes and in the periplasmic space. The deposits had amorphous or poor nanocrystalline structures.
2005. "Simple Synthesis of Hierarchically Ordered Mesocellular Mesoporous Silica Materials Hosting Crosslinked Enzyme Aggregates." Small 1(7):744-753. Abstract : Hierarchically ordered mesocellular mesoporous silica materials (MMS) were synthesized using a single structure directing agent under neutral conditions for the first time. The mesocellular pores are synthesized without adding any pore expander, and the walls of cellular pores in MMS are composed of SBA-15 type mesopores. The small-angle X-ray scattering (SAXS) pattern of MMS revealed the presence of ordered pore structures with two different length scales. The current MMS possesses four different pore systems; complementary micro/mesopores, main 13 nm mesopores, 40 nm mesocellular spherical pores, and textural inter-particle macropores. Nanometer-scale enzyme reactors (NER) were developed in mesocellular mesoporous silica (MMS) via a ship-in-a-bottle approach, which employs adsorption of enzymes followed by cross-linking using glutaraldehyde (GA) treatment. The resulting NER show an impressive stability and activity with an extremely high loading of enzymes. For example, NER containing α-chymotrypsin (NER-CT) could hold 0.5 g CT in 1 g of silica, but the specific activity of NER-CT was 10.4 times higher than that of the adsorbed CT with a lower loading (0.07 g CT per 1 g of silica), which was further decreased by a continuous leaching of adsorbed CT. NER-CT showed excellent stability without any leaching, i.e. no activity decrease at all in a rigorously-shaking condition for two weeks (a half-life with 3.8 years), while the conventional adsorption method resulted in a half-life of 3.6 days in the same condition.
2005. "Simple Fabrication of a Highly Sensitive and Fast Glucose Biosensor using Enzyme Immobilized in Mesocellular Carbon Foam." Advanced Materials 17(23):2828-2833. Abstract We fabricated a highly sensitive and fast glucose biosensor by simply immobilizing glucose oxidase in mesocellular carbon foam. Due to its unique structure, the MSU-F-C enabled high enzyme loading without serious mass transfer limitation, resulting in high catalytic efficiency. As a result, the glucose biosensor fabricated with MSU-F-C/GOx showed a high sensitivity and fast response. Given these results and the inherent electrical conductivity, we anticipate that MSU-F-C will make a useful matrix for enzyme immobilization in various biocatalytic and electrobiocatalytic applications.
2005. "Ferrous Hydroxy Carbonate is a Stable Transformation Product of Biogenic Magnetite." American Mineralogist 90(2-3):510-515. Abstract A ~1:1 mixture of ferrihydrite and nanocrystalline akaganeite (β-FeOOH; 10-15 nm) was incubated with Shewanella putrefaciens (strain CN32) under anoxic conditions with lactate as an electron donor and anthraquinone-2,6-disulfonate (AQDS) as an electron shuttle. The incubation was carried out in a 1,4-piperazinediethanesulfonic acid (PIPES)-buffered medium, without PO₄³⁻ at circumneutral pH. Iron reduction was measured as a function of time (as determined by 0.5 N HCl extraction), and solids were characterized by X-ray diffraction (XRD), electron microscopy, and Mössbauer spectroscopy. The biogenic reduction of Fe(III) was rapid; with 60% of the total Fe (Feтот) reduced in one day. Only an additional 10% of Feтот was reduced over the next three years. A fine-grained (10 nm), cation-excess (CE) magnetite with a Fe(II)/Feтот ratio of 0.5-0.6 was the sole biogenic product after one day of incubation. The CE magnetite was unstable and partially transformed to micron-sized ferrous hydroxy carbonate [FHC; Fe2 (OH)2CO3⒮], a rosasite-type mineral, with time. Ferrous hydroxy carbonate dominated the mineral composition of the three year incubated sample. The Fe(II)/Feтот ratio of the residual CE magnetite after three years of incubation was lower than the day 1 sample and was close to that of stochiometric magnetite (0.33). To best of our knowledge, this is the first report of biogenic FHC, and only the third observation of this material in nature. Ferrous hydroxy carbonate appeared to form by slow reaction of microbially produced carbonate with Fe(II)-excess magnetite. The FHC may be an overlooked mineral phase that explains the infrequent occurrence of fine-grained, biogenic magnetite in anoxic sediments.
2005. "A Magnetically Separable, Highly Stable Enzyme System Based on Nanocomposites of Enzymes and Magnetic Nanoparticles Shipped in Hierarchically Ordered, Mesocellular, Mesoporous Silica." Small 1(12):1203-1207. doi:10.1022/smll.200500245 Abstract Enzymes are versatile nanoscale biocatalysts, and find increasing applications in many areas, including organic synthesis[1-3] and bioremediation.[4-5] However, the application of enzymes is often hampered by the short catalytic lifetime of enzymes and by the difficulty in recovery and recycling. To solve these problems, there have been a lot of efforts to develop effective enzyme immobilization techniques. Recent advances in nanotechnology provide more diverse materials and approaches for enzyme immobilization. For example, mesoporous materials offer potential advantages as a host of enzymes due to their well-controlled porosity and large surface area for the immobilization of enzymes.[6,7] On the other hand, it has been demonstrated that enzymes attached on magnetic iron oxide nanoparticles can be easily recovered using a magnet and recycled for iterative uses.[8] In this paper, we report the development of magnetically-separable and highly-stable enzyme system by the combined use of two different kinds of nanostructured materials: magnetic nanoparticles and mesoporous silica.
2004. "Biogeochemical Transformation of Fe Minerals in a Petroleum-Contaminated Aquifer." Geochimica et Cosmochimica Acta 68(8):1791-1805. Abstract Biogeochemical Transformation of Fe Minerals in a Petroleum-Contaminated Aquifer
2004. "Microscale Distribution of Cesium Sorbed to Biotite and Muscovite." Environmental Science and Technology 38(4):1017-1023. Abstract Individual 1 – 3 mm biotite and muscovite clasts from Hanford sediment were contacted with 0.001 – 0.08 M CsNO--3, then either left whole or sectioned perpendicular to their basal planes for examination by electron or X-ray microprobe. Cs+ was observed to preferentially sorb to mica edges, steps on mica surfaces, or to fractured regions. The observed localization conformed to hypothesized strong binding to frayed edge sites in preference to sites on basal planes. In section, Cs+ was found to penetrate the mica interior, forming discrete zones of concentration, particularly in muscovite. In biotite, Cs was more abundant, permeating the clasts at low abundance and forming also discrete zones of higher concentration. Concentrated Cs on both clast edges and within clast interiors corresponded to microscopic but relatively extensive zones where K was depleted. The localization of sorbed Cs in areas where K was depleted suggested that weathering reactions caused the formation of frayed edges sites within the micas. Cs+ accessed crystal interiors by diffusion along channels following crystal defects, cracks, or partings where pore fluids had previously migrated to form the interior alteration zones. At nm scale, areas with localized Cs were disordered, confirming that FES were developed in clast interiors. Cs+ that was adsorbed in this manner could be very resistant to desorption, as has been often observed.
2004. "Electron Microscopy Evaluation of the Role of Dissimilatory Metal-reducing Bacteria in Biomineralization Pathways." Microscopy and Microanalysis 10(suppl 2):1538-1539. Abstract The importance of microorganisms in the biogeochemical cycling of Fe is well-recognized [1]. Dissimilatory metal-reducing bacteria (DMRB), which are ubiquitous in soils and aquifers, couple the oxidation of organic matter or H2 with the reduction of various Fe(III) oxide phases to obtain energy for growth and function. They can also catalyze Fe(III) reduction under anaerobic conditions, utilizing crystalline and poorly crystalline iron oxides as a terminal electron acceptor. Microbially induced Fe mineral transformations were examined using the Shewanella putrefaciens, strain CN32 in an artificial groundwater medium in columns under advective flow conditions. Columns were filled with ferrihydrite-coated quartz sand inoculated with S. putrefaciens (initial cell density 10⁸ mL-¹). Lactate was added as an electron donor. Changes in microbial metabolism, aqueous chemistry, and solid phase distributions were monitored at time points until termination of the column experiment at 16 days [2]. Transmission (TEM) and scanning electron microscopy (SEM) was used for investigating mineral association with bacterial cells, crystal size, morphology, and spatial relationships. A special TEM sample preparation protocol developed in our laboratory was used for the accurate preservation of both the biological and mineral portion of the sample [3]. To eliminate the anaerobic sample exposure to oxygen, the whole embedding procedure, as well as the thin sectioning on an ultramicrotome was carried out in an anaerobic glove box (95% argon, 5% hydrogen). Ultra thin sections of the material were studied using a JEOL 2010 TEM operating at 200 kV coupled to an Oxford EDS system. Images were collected and analyzed using a Digital Micrograph (Gatan). Selected area diffraction patterns were evaluated by the Desktop Microscopist (Lacuna) software. Visual changes in the solid-phase within the column were evident: initial orange ferrihydrite started turning brown, and further darkened over the course of the experiment, as it was converted to predominantly goethite and magnetite [Fig. 1A, 1B]. In addition to spectroscopic methods, the presence of goethite and magnetite was further confirmed by TEM and SEM, and the spatial orientations and particle size of mineral particles were determined. Typical needle-like structures of goethite crystals were predominantly associated with the surface of ferrihydrite, but were also found coupled with microbial cell surfaces. In fact, some bacterial cells appeared completely encrusted in goethite, most likely, a result of electrostatic attraction between newly precipitated goethite and the microbial surface. Magnetite, on the other hand, was mainly associated with the ferrihydrite surface, and only rarely with the cell surface. Thus, the bacterial cell is only indirectly (by ferrous iron production) responsible for goethite and magnetite formation. Although intracellular precipitation of iron oxides in S. putrefaciens was recently reported [4], only extracellular precipitation was observed in this experimental setup. The ability of bacteria to shed the mineral deposits from their outer membranes in order to prevent their surfaces passivation caused by mineral sorption will be discussed. Bacteria appeared to primarily serve as an Fe(II) source for the system; secondary mineralization was confirmed as a function of initial Fe(II) concentrations. The mechanism of the enzymatic reduction is not completely understood, and the accountable protein functions are being intensively investigated by several molecular biology techniques. Current and future studies will include immunogold labeling at the electron microscopy level as a method for determining the localization of these proteins.
2004. "Uranium Immobilization by Sulfate-reducing Biofilms." Environmental Science and Technology 38(7):2067-2074 . Abstract Hexavalent uranium [U(VI)] was immobilized using biofilms of the sulfate-reducing bacterium (SRB) Desulfovibrio desulfuricans G20. The biofilms were grown in flat-plate continuous-flow reactors using lactate as the electron donor and sulfate as the electron acceptor. U(VI) was continuously fed into the reactor for 32 weeks at a concentration of 126 íM. During this time, the soluble U(VI) was removed (between 88 and 96% of feed) from solution and immobilized in the biofilms. The dynamics of U immobilization in the sulfate-reducing biofilms were quantified by estimating: (1) microbial activity in the SRB biofilm, defined as the hydrogen sulfide (H2S) production rate and estimated from the H2S concentration profiles measured using microelectrodes across the biofilms; (2) concentration of dissolved U in the solution; and (3) the mass of U precipitated in the biofilm. Results suggest that U was immobilized in the biofilms as a result of two processes: (1) enzymatically and (2) chemically, by reacting with microbially generated H2S. Visual inspection showed that the dissolved sulfide species reacted with U(VI) to produce a black precipitate. Synchrotron-based U L3-edge X-ray absorption near edge structure (XANES) spectroscopy analysis of U precipitated abiotically by sodium sulfide indicated that U(VI) had been reduced to U(IV). Selected-area electron diffraction pattern and crystallographic analysis of transmission electron microscope lattice-fringe images confirmed the structure of precipitated U as being that of uraninite.
2003. "Sorption versus Biomineralization of Pb(II) within Burkholderia cepacia Biofilms." Environmental Science and Technology 37((2)):300-307. Abstract X-ray spectroscopy measurements have been combined with macroscopic uptake data and transmission electron microscopy (TEM) results to show that Pb(II) uptake by Burkholderia cepacia is due to simultaneous sorption and biomineralization processes. X-ray microprobe mapping of B. cepacia biofilms formed on -Al2O3 surfaces shows that Pb(II) is distributed heterogeneously throughout the biofilms because of the formation of Pb "hot spots". EXAFS data and TEM observations show that the enhanced Pb accumulation is due to the formation of nanoscale crystals of pyromorphite (Pb5(PO4)3(OH)) adjacent to the outer-membrane of a fraction of the total population of B. cepacia cells. In contrast, B. cepacia cell suspensions or biofilms that were heat-killed or pretreated with X-rays do not form pyromorphite, which suggests that metabolic activity is required. Precipitation of pyromorphite occurs over several orders of magnitude in [H+] and [Pb] and accounts for approximately 90% of the total Pb uptake below pH 4.5 but only 45-60% at near-neutral pH because of the formation of additional Pb(II) adsorption complexes. Structural fits of Pb LIII EXAFS data collected for heat-treated cells at near-neutral pH suggest that Pb(II) forms inner-sphere adsorption complexes with carboxyl functional groups in the biofilms.
2003. "Transformation of 2-line Ferrihydrite to 6-line Ferrihydrite under Oxic and Anoxic Conditions." American Mineralogist 88(11-12):1903-1914 pt. 2. Abstract New Insights into the Transformation of 2-line Ferrihydrite to Crystalline Iron Oxides: The First Observation of 6-line Ferrihydrite during Solid-state Transformation
2003. "Secondary Mineralization Pathways Induced by Dissimilatory Iron Reduction of Ferrihydrite Under Advective Flow." Geochimica et Cosmochimica Acta 67(16):2977-2992. Abstract Secondary Mineralization Pathways Induced by Dissimilatory Iron Reduction of Ferrihydrite Under Advective Flow
2002. "Lithium-Assisted Self-Assembly of Aluminum Carbide Nanowires and Nanoribbons." Nano Letters 2(2):105-108. Abstract We report on the synthesis and self-assemlby of Al4C3 nanowires and nanoribbons using lithium as a catalyst. Large quantities of Al4C3 nanowires (diameters from 5 to 70 nm) and nanoribbons (5-70 nm thick and 20-5600 nm wide) tens of micrometers long were synthesized serendipitously in a solid-state reaction involving Al/C/Li at less than 780 degrees Celcius. High-resolution electron microscopy revealed that the nanowires all grew along the c-axis of hexagonal Al4C3, whereas the nanoribbons all grew within the basal plane. The facile synthesis of the Al4C3 nanowires and nanoribbons suggest similar nanostructures of other carbide and nitride materials may be made using the lithium-assisted self-assembly process.
2002. "Modeling the Inhibition of the Bacterial Reduction of U(VI) by beta-MnO2(S)(g)." Environmental Science and Technology 36(7):1452-1459. Abstract Modeling the Inhibition of the Bacterial Reduction of U(VI) by b-MnO2(s)
2002. "Influence of Mn oxides on the reduction of U(VI) by the metal-reducing bacterium Shewanella putrefaciens." Geochimica et Cosmochimica Acta 66(18):3247-3262. Abstract Dissimilatory metal-reducing bacteria (DMRB) enzymatically reduce Fe(III), Mn(III/IV), U(VI), and other polyvalent metals during anaerobic respiration. Previous investigations of the bacterial reduction of U(VI) in the presence of goethite (a-FeOOH) found that, in spite of potential competition as an electron acceptor, goethite had little impact on the bacterial reduction of U(VI) to insoluble U(IV). Mn(III/IV) oxides are also electron acceptors for DMRB but are stronger oxidants than Fe(III) oxides. Differences in the solubility of oxidized Mn and U challenges predictions of their biogeochemical behavior during redox cycling. The potential for Mn oxides to modify the biogeochemical behavior of U during reduction by a subsurface bacterium Shewanella putrefaciens CN32 was investigated using synthetic Mn(III/IV) oxides [pyrolusite (?-MnO2), bixbyite (Mn2O3) and K+-birnessite (K4Mn14O27?8H2O)]. In the absence of bacteria, pyrolusite and bixbyite oxidized biogenic uraninite (UO2(s)) to soluble U(VI) species, with bixbyite being the most rapid oxidant. The Mn(III/IV) oxides lowered the bioreduction rate of U(VI) relative to rates in their absence, or in the presence of gibbsite [Al(OH)3] added as a non-redox reactive surface. Evolved Mn(II) increased with increasing initial U(VI) concentration in the biotic experiments, indicating that valence cycling of U facilitated the reduction of Mn(III/IV). Despite an excess of the Mn oxide, 43-100% of the initial U was bioreduced after extended incubation. Analysis of thin sections of bacterial-Mn oxide suspensions revealed that the reduced U resided in the periplasmic space of the bacterial cells. In the absence of Mn(III/IV) oxides, UO2(s) accumulated as copius fine-grained particles external to the cell. These results indicate that the presence of Mn(III/IV) oxides may impede the biological reduction of U(VI) in subsoils and sediments?..
2002. "Structural and chemical characterization of aligned crystalline nanoporous MgO films grown via reactive ballistic deposition ." Journal of Physical Chemistry B 106(14):3526-3529 . doi: 10.1021/jp013801c Abstract Highly-porous (~ 90%), high-surface area (~ 1000 m2/g), thermally stable (1200 K) crystalline films of MgO are synthesized using a novel reactive ballistic deposition technique. The film consists of a tilted array of porous nanoscale crystalline filaments. Surprisingly, the individual filaments exhibit a high degree of crystallographic order with respect to each other. These films have chemical binding sites analogous to those on MgO(100). However the fraction of chemically active, high energy binding sites is greatly enhanced on the nanoporous film. This unique collection of properties makes these materials attractive candidates for chemical applications such as sensors and heterogeneous catalysts.
2002. "Structural and Chemical Characterization of Aligned Crystalline Nanoporous MgO Films Grown via Reactive Ballistic Deposition." Journal of Physical Chemistry B 106(14):3526-3529. Abstract Highly-porous (~90%), high-surface area (~1000 m2/g), thermally stable (1200K) crystalline films of MgO are synthesized using a novel reactive ballistic deposition techniques. The film consists of a tilted array of porous nanoscale crystalline filaments. Suprisingly, the individual filaments exhibit a high degree of crystallographic order with respect to each other. These films have chemical binding sites analogous to those on MgO (100). However, the fraction of chemically active, high energy binding sites is greatly enhanced on the nanoporous film. This unique collection of properties makes these materials attractive candidates for chemical applications such as sensors and heterogeneous catalysts.
2002. "Steam Reforming of Methanol Over Highly Active Pd/ZnO Catalyst." Catalysis Today 77 (1-2)(DEC 1 2002):79-88. Abstract "Pd/ZnO catalysts were investigated for steam reforming of nethanol. Unlike precious metal-based catalysts, Pd/ZnO catalysts not only exhibited high activity, but more importantly very low selectivity to CO for methanol steam reforming. Under the conditions examined, the decomposition activity is minimal. The novel function is attributed to the formation of highly structured Pd-Zn alloy at moderate temperatures under mild reducing environments."
2002. "Steam Reforming of Methanol over Highly Active Pd/ZnO Catalyst ." Catalysis Today 77(1-2):79-88. Abstract Pd/ZnO catalysts were investigated for steam reforming of methanol. Unlike precious metal based catalysts, Pd/ZnO catalysts not only exhibited high activity, but more importantly very low selectivity to CO for methanol steam reforming. Under the conditions examined, the decomposition activity is minimal. The novel function is attributed to the formation of highly structured Pd-Zn alloy at moderate temperatures under mild reducing environments. The current catalytic system was characterized by TPR, TEM, H₂ chemisorption, and XRD.
2001. "Alkaliphilus transvaalensis gen. nov., sp. nov., an extremely alkaliphilic bacterium isolated from a deep South African gold mine." International Journal of Systematic and Evolutionary Microbiology 51(part 4):1245-1256. Abstract A novel extreme alkaliphile was isolated from a mine water containment dam at 3.2 km bellow land surface in an ultra-deep gold mine near Carletonville, South Africa. The cells of this bacterium were straight to slightly curved rods, motile by flagella and formed endospores, Growth was observed over the temperature range 20-50 degreesC (optimum 40 degreesC; 45 min doubling time) and ph range 8.5-12.5 (optimum pH 10.0). The novel isolate, one of the most alkaliphilic micro-organisms yet described, was a strictly anaerobic chemo-organotroph capable of utilizing proteinaceous substrates such as yeast extract, peptone. tryptone and casein, Elemental sulfur, thiosulfate or fumarate, when included as accessory electron acceptors, improved growth. The G+C content of genomic DNA was 36.4 mol%, Phylogenetic analysis based on the 16S rDNA sequence indicated that the isolate is a member of cluster XI within the low G+C Cram-positive bacteria, but only distantly related to previously described members. On the basis of physiological and molecular properties, the isolate represents a novel species, for which the name Alkaliphilus transvaalensis gen. nov., sp. nov. is proposed (type strain SAGM1(T) = JCM 10712(T) = ATCC 700919(T), The mechanism of generation of the highly alkaline microbial habitat and the possible source of the alkaliphile are discussed.
2001. "Iron Sulfides and Sulfur Species Produced at (001) Hematite Surfaces in the Presence of Sulfate-Reducing Bacteria." Geochimica et Cosmochimica Acta 65(2):223-235. Abstract In the presence of sulfate-reducing bacteria (Desulfovibrio desulfuricans) hematite (a-Fe2O3) dissolution is affected and hydrogen sulfide, the product of sulfate reduction is released. As a consequence, ferrous ions are free to react with excess H2S to form insoluble iron sulfides. X-ray photoelectron spectra indicate binding energies consistent with the iron sulfides having a pyrrhotite structure (Fe2p3/2 708.4 eV; S2p3/2 161.5 eV). Other sulfur species identified at the surface include sulfate, sulfite and polysulfides. X-ray diffraction suggests an unidentifiable crystal structure at the hematite surface develops within 3 months, HRTEM confirms the presence of a hexagonal structure again suggesting the formation of pyrrhotite. The identification of pyrrhotite is inconsistent with previous reports in which mackinawite and greigite were products of biological sulfate reduction (Rickard 1969; Herbert et al 1998). The apparent differences in stoiciometries may be related to the availability of Fe2+(aq.) at the mineral surface through respiratory iron reduction by subsurface bacteria. The significance of pyrrhotite and polysulfide production in relation to the S- and Fe-cycles and to trace metal bioavailability is discussed.