B12A-01 INVITED
Thermodynamic Modeling of the Adsorption of Metals onto Microbial Cell Walls: Current Challenges
Adsorption of metals onto biological surfaces can influence the speciation, bioavailability, and mobility of metals in a range of geologic systems. In this talk, I will summarize research that improves our understanding of metal adsorption reactions involving microbial cell walls (including bacterial, fungal, algal, and plant root cells). The review covers two general types of investigations: 1) those that aim to improve our molecular-scale understanding of metal adsorption onto cell walls, thereby improving the accuracy of thermodynamic adsorption models; and 2) those that aim to improve our ability to apply thermodynamic models of metal-microbial adsorption to complex realistic settings. The first type of research involves a range of experimental approaches, and the most common approaches will be described. The second type of research results from the need to balance the flexibility that comes with chemical sophistication in geochemical models with the practical impossibility of modeling the complexity of real systems at a molecular scale. Hybrid approaches that incorporate some degree of molecular-scale insight with testable simplifying assumptions are described and offer some hope for extrapolating our newfound molecular-scale understanding of metal-microbial adsorption reactions to estimate mass transport in microbe-bearing systems. The objective of this review is to describe both progress and remaining challenges, in order to promote research that ultimately will yield accurate and flexible models of the effects of microbial adsorption on metal distributions and speciation in natural geologic systems.
B12A-02 INVITED
Probing for the Activities of Arsenic and Selenium Metabolizing Microbes
Microbial activities can directly impact the mobility and toxicity of arsenic and selenium in the environment. Arsenic is cycled through oxidation/reduction and methylation/demethylation reactions as part of resistance and respiratory processes. The requirement for selenium is primarily for incorporation into selenocysteine and its function in selenoenzymes. Selenium oxyanions can also serve as an electron acceptor in anaerobic respiration. Both culture and culture-independent methods have been developed to detect the presence and activity of organisms capable of arsenic and selenium transformations. Enrichment media have been successful at cultivating arsenate respiring bacteria from a variety of environments, however, both electron donor and the concentration of arsenic can exert strong selective pressure. Thus, the organisms in the enrichment culture may not be the dominant organisms in the environment. Culture-independent methods, including immunological approaches (e.g., polyclonal antibodies to ArrA) and PCR-based technologies, have also had mixed success. PCR-primers designed to amplify portions of genes involved in resistance (e.g., arsC, acr3), respiration (e.g., arrA), and oxidation (e.g., aoxB) have been useful in several environments. Applications include T-RFLP, rt-PCR, and DGGE analyses. Nevertheless, these primers do not work with certain organisms suggesting the existence of additional enzymes and pathways. Although the biosynthetic pathway (and the proteins involved) for selenocysteine has been described in detail, much less is known about selenium methylation, assimilation and respiration. Only one respiratory selenate reductase has been characterized and its close sequence identity with chlorate and perchlorate reductases has complicated efforts to design a functional probe. Thus many aspects of the biogeochemical cycle of selenium remains to be explored.
B12A-03
Zinc Isotopes in the Soil-Plant Interface
From the geosphere/biosphere system as a whole down to the organism and cellular level, isotopes have the potential capability to uniquely understand the fluxes of inorganic elements. Zinc is of particular interest as it is one of the trace elements essential for living organisms and most usefully its chemistry is simplified by the possession of only one oxidised state (II). The use of multicollector ICP-MS, together with complete sample digestion and anion exchange chromatography, has allowed the measurement of Zn isotopes to be made precisely (below ±0.1‰ (2 S.D., n=4 typically)) and accurately in the geological and biological matrices studied. Zinc deficiency is the most widespread micronutrient disorder in rice (\it Oryza \it sativa) and differences between genotypes render some genotypes more susceptible to deficiency than others. Hence rice was chosen as a model species in our uptake and fractionation studies. A previous hydroponic study in our laboratory showed Zn uptake by tomato, lettuce and rice all produced an enrichment of the light Zn isotopes in plant shoots. A study of vegetation in a watershed, however, revealed a more complex picture, and plant shoots and roots were generally enriched in heavy isotopes relative to the litter and superficial soils. In the results presented here, rice grown under field conditions showed only heavy or insignificant fractionations relative to the soil matrix (in contrast to the hydroponic study). A genotype tolerant to Zn soil deficiency (line 46) and a genotype intolerant to deficiency (IR74) were grown in both zinc fertilised and unfertilised (Zn deficient) plots as part of a larger study. On the zinc fertilised plots, shoot samples of both genotypes showed a negligible difference in δ66ZnIMP-Zn compared to the growth soil. On unfertilised plots (soil δ66ZnIMP- Zn = 0.14 ± 0.10 ‰ (2 S.E., n=3)), however, line 46 rice showed preferential heavy uptake (δ66ZnIMP-Zn = 0.35 ± 0.04 ‰ (2 S.E., n=4)) compared to IR74 (δ66ZnIMP-Zn = 0.22 ± 0.07 ‰ (2 S.E., n=4)). The most likely explanation for a heavy signature is that of an equilibrium reaction induced in the zinc deficient soil environment. It has been hypothesised by plant scientists that under Zn limiting conditions, plants may secrete complexing agents that bind Zn2+ outside the plant and that it is these complexes that are then taken up across the cell membrane. It has been difficult to prove such a hypothesis due to the inherent problems of conducting experiments under normal Zn deficient conditions; however, new isotope studies such as these promise to provide additional insight. http://www3.imperial.ac.uk/earthscienceandengineering/people/phdstudents/a-b/arnoldt
B12A-04
Sorption of Yttrium and the Rare Earth Elements on Non-Living Macroalgal Tissue
We have investigated sorption of yttrium and the rare earth elements (YREEs) on tissue of the green macroalga Ulva lactuca, commonly known as sea lettuce. Due to its nearly worldwide distribution in coastal waters, very simple morphology, and prodigious capacity for trace metal uptake from seawater, members of the Ulva genus serve as a basic but representative model of marine organic substrates in this type of study. In order to exclude active biological uptake effects, allowing us to focus on passive chemical mechanisms, we performed our initial experiments with sea lettuce Certified Reference Material consisting of a dehydrated, powdered tissue homogenate. A small quantity of this powder was suspended in NaCl solutions containing all YREEs, except Pm, at pH 3 and T = 25°C. The extent of YREE sorption was determined as a function of pH at constant temperature by titrating the solution with dilute NaOH and measuring the YREE concentrations of 0.2-μm filtered aliquots with an ICP-MS at regular time intervals after each pH adjustment. In NaCl solutions with an ionic strength approaching that of seawater, distribution coefficients, which quantify the proportion of sorbed and dissolved metal concentrations, are a highly linear function of pH in the range 3-8. The slope of the line suggests a sorption mechanism that involves ion exchange with both H+ and Na+ on surface functional groups. The shape of solution YREE patterns indicates that these functional groups are probably carboxylates at low and intermediate pH, but that other groups may contribute at high pH. The identification of carboxylate functional groups appears to be confirmed by preliminary results from EXAFS spectroscopic analyses of individual REE sorbed on the surface of Ulva lactuca tissue under similar conditions, conducted at the ANL Advanced Photon Source. In dilute NaCl solutions the distribution coefficient is largely independent of pH. We believe that prolonged exposure of the tissue to a low ionic strength solution may modify the chemical structure of the cell wall and make it permeable to organic ligands that otherwise sequester the YREEs in the cell interior. Chemical extraction of filtered solutions from the low ionic strength experiment with silica-bonded C18, showing that a substantial fraction of dissolved YREEs is distinctly hydrophobic, seems to support this hypothesis. Additional experiments to clarify these observations, including acid-base titrations of the Ulva lactuca tissue to assess the number of different functional groups and their surface densities, are currently ongoing.
B12A-05
Macroscopic and Spectroscopic Analysis of Lanthanide Adsorption to Bacterial Cells
The adsorptive behaviour of lanthanides is increasingly becoming a major focus of research, partly because lanthanides are good analogues for understanding the behaviour of the more problematic actinides, and also because when studied as a suite, their fractionation patterns make them important indicators of geochemical processes, including possible biosignature applications. We have combined macroscopic adsorption modelling, spectroscopic analysis and linear free energy relationships to explore mechanisms by which lanthanides interact with bacterial cell surfaces. Macroscopic adsorption modelling suggests that all lanthanides preferentially adsorb to phosphate sites below neutral pH values. However, carboxyl complexation also explains most of the adsorption density below pH ~5. Furthermore, analysis of the recovered stability constants using linear free energy relationships showed that most of the adsorption density could be predicted using stability constants estimated using fulvic acid complexation. Nevertheless, EXAFS analysis confirmed mixed-mode co-ordination, showing lanthanide co- ordination to phosphate sites at low pH and low metal to biomass ratios, with secondary involvement of carboxyl sites at pH values greater than 5. This study therefore demonstrates the importance of combining direct spectroscopic measurement with modelling in determining metal adsorption sites.
B12A-06
Can Environmental Microbes Mobilize and Oxidize Arsenic from Shale into Groundwater?
Elevated levels of arsenic are found in New Jersey well water in the Newark Basin where 15% of the wells tested exceed 10 μg/L, to a maximum of 215 μg/L. The source may be from the weathering of pyrite (FeS2) found in the black shale, which can contain up to 4% arsenic by weight. We hypothesize that microorganisms found in the environment can oxidize sulfide in the pyrite to release the bound arsenic, and in addition, that microbes can oxidize As(III) to As(V) to further enhance the mobilization of the arsenic released from the shale. To examine this, cultures were established with weathered black shale from an outcrop of the Newark Basin's Lockatong formation. A chemoautotrophic As(III)-oxidizer, strain WAO, was isolated, physiologically and phylogenetically characterized, and based on 16S rDNA sequence analysis it is most closely related to the genus Bosea. In the presence of the mineral arsenopyrite (FeAsS) strain WAO releases arsenic and sulfur with oxidation of stoichiometric amounts to arsenate and sulfate. Strain WAO also displays preferential colonization of the pyrite surface on sections of arsenic-bearing black shale from the Lockatong formation. These observations suggest that microbial mobilization can be a mechanism for arsenic release into groundwater in the Newark Basin and elsewhere as well.
B12A-07
Influence of Arsenic on the Reduction of Lepidocrocite and Hydroxycarbonate Green Rust 1 Into Ferrous-Carbonate Hydroxide by Shewanella putrefaciens
Shewanella putrefaciens, an iron-respiring bacterium, is capable of reducing As(V) to As(III) when HAs(V)O42- is the sole electron acceptor, or when it is adsorbed on the surface of lepidocrocite (γ-FeOOH), a common well-crystallized hydromorphic soil mineral. Cultures in which lepidocrocite was used as the sole electron acceptor (i.e., pure lepidocrocite and As(III)-adsorbed lepidocrocite) led to the formation of biogenic hydroxycarbonate green rust 1 (GR1) prior to precipitation of ferrous-carbonate hydroxide (FCH). However, the presence of As(III) slowed down the biotransformation of hydroxycarbonate GR1 into FCH, leading to the co-occurrence of both phases after 22-month of aging. In contrast, when the electron acceptor was As(V)- bearing lepidocrocite, XRD analysis revealed FCH to be the dominant reaction product; no green rust formation was observed in this case. Arsenic K-edge XANES spectroscopy indicated that all As(V) (K-edge(max) = 11875.0eV) was reduced to As(III) (K-edge(max) = 11871.3eV), suggesting the presence of As(III) either on the surface of FCH, and/or in another ferrous-containing solid phase. Mössbauer analysis of the 22-month aged biogenic sample obtained from the bioreduction of As(III)-adsorbed lepidocrocite revealed the presence of FCH (80%) and GR (20%). In contrast, Mössbauer spectra of white biogenic solid phases resulting from the bioreduction of As(V)-adsorbed lepidocrocite and of pure lepidocrocite were characterized by ferrous iron doublets. These spectra indicate that the white biogenic sample without arsenic contains 100% Fe(II) assigned to FCH, while the white biogenic As(III)-Fe(II)-containing reaction products consists of 87% FCH and of 13% of another ferrous phase, probably an As(III)-Fe(II)-containing compound. These results show for the first time bacterial reduction of stoichiometric hydroxycarbonate green rust 1.
B12A-08
Effect of Wetland Plants on the Reduction of Hexavalent Chromium in Sediments
The effect of wetland plants (Typha latifolia and Carex lurida) on the reduction of Cr(VI) to Cr(III) at relatively low initial Cr(VI) concentrations (< 5 mg/L) was studied. Experiments were carried out using tubular microcosms operated in a greenhouse under plug flow conditions. Cr(VI) reduction was enhanced significantly by the presence of plants. This is explained by a decrease of the redox potential promoted by the root exudates released by the roots of the plants. Using these root exudates as their carbon source, and in the absence of nitrates, sulfate-reducing bacteria are able to reduce sulfate, building up the concentration of acid volatile sulfide (AVS), which in turn reduce Cr(VI) to Cr(III). Moreover, evapotranspiration induced by plants also contributes to enhance the removal of Cr(VI) by concentrating the species in the sediment porewater and therefore affecting the reaction kinetics. The effect of diurnal cycles on the Cr(VI) reduction rate has also been also studied. The diurnal cycle influences both root exudates released by plants as well as evapotranspiration. Higher levels of dissolved organic carbon in the pore water and evapotranspiration, occurring from around noon to 3 pm, correspond to a higher sulphate reduction rate and a lower redox potential in the sediments. This promotes a higher Cr(VI) removal during the day hours. Finally the Cr(VI) reduction dynamics were fitted to a kinetic equation that depends on the AVS and Cr(VI) concentration.