Hexavalent Chromium Generation within Naturally Structured Soils

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Hexavalent Chromium Generation within Naturally Structured Soils and Sediments Debra M. Hausladen, and Scott Fendorf Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.6b04039 • Publication Date (Web): 13 Jan 2017 Downloaded from http://pubs.acs.org on January 14, 2017

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Environmental Science & Technology

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EST Article

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Hexavalent Chromium Generation within Naturally Structured Soils and Sediments

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Debra M. Hausladen and Scott Fendorf*

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Earth System Science Dept.

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Stanford University, Stanford, CA 94305. USA

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*corresponding author. Email: [email protected]; Phone: (650) 723-5238; Fax: (650) 725-2199

ACS Paragon Plus Environment


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ABSTRACT

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Chromium(VI) produced from the oxidation of indigenous Cr(III) minerals is increasingly being

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recognized as a threat to groundwater quality. A critical determinant of Cr(VI) generation within

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soils and sediments is the necessary interaction of two low solubility phases - Cr(III) silicates or

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(hydr)oxides and Mn(III/IV) oxides - that lead to its production. Here we investigate the

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potential for Cr(III) oxidation by Mn oxides within fixed solid matrices common to soils and

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sediment. Artificial aggregates were constructed from Cr(OH)3- and Cr0.25Fe0.75(OH)3-coated

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quartz grains and mixed either with synthetic birnessite or inoculated with the Mn(II)-oxidizing

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bacteria Leptothrix cholodnii. In aggregates simulating low organic carbon environments, we

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observe Cr(VI) concentrations within advecting solutes at levels more than twenty-times the

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California drinking water standard. Chromium(VI) production is highly dependent on Cr-mineral

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solubility; increasing Fe-substitution (x=0 to x=0.75) decreases the solubility of the solid and

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concomitantly decreases total Cr(VI) generation by 37%. In environments with high organic

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carbon, reducing conditions within aggregate cores (microbially) generate sufficient Fe(II) to

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suppress Cr(VI) efflux. Our results illustrate Cr(VI) generation from reaction with Mn oxides

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within structured media simulating soils and sediments and provide insight into how fluctuating

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hydrologic and redox conditions impact coupled processes controlling Cr and Mn cycling.

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INTRODUCTION Anthropogenic chromium is a well-known pollutant that often results from industrial

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processes including leather tanning, metal plating, stainless steel production, and chrome

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pigment manufacturing.1,2 Geogenic Cr(III) is widespread3–9, however, and may represent an

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important source of Cr(VI) if an oxidation pathway exists capable of producing appreciable


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Cr(VI) despite the low solubility of Cr(III)-bearing minerals10. Inhalation, ingestion, and dermal

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exposure to Cr(VI) can result in severe adverse health effects to humans, inclusive of respiratory

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and non-respiratory cancers.11 As Cr(VI) structurally parallels phosphate and sulfate, chromate

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anions are actively transported to cells throughout the body and organs.12 In contrast to Cr(VI),

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Cr(III) is an essential nutrient for humans that is thought to help with glucose transport into

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cells.13 Further differentiating Cr(III) and Cr(VI) are the solubilities of their mineral phases and

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propensity for transport within surface and subsurface environments. Chromium(III) forms low

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solubility hydroxide precipitates10 and strong mineral complexes14. The more toxic Cr(VI)

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species resides as the chromate (HCrO4-) anion that binds less extensively to soil and sediment

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minerals, and thus has both greater dissolved concentrations and propensity for transport within

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water systems.15–17

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Owing to the potential for oxidation, naturally occurring Cr(III) residing within geologic

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strata poses a widespread threat to water quality and human health. Chromium is the tenth most

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abundant element in Earth’s mantle14, and Cr-bearing minerals cover ca. 1% of Earth’s land

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surface, principally found in serpentenized and ultramafic rocks that are concentrated around

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convergent plate boundaries3,18. Weathering of the primary minerals within soil and sediments

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commonly results in Cr(III) hydroxide precipitates, often coprecipitated with Fe(III)- and Al(III)-

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hydroxides, and may coincide with Fe and Mn oxides.19,20 While concentrations in unaltered

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bedrock may reach 2 g/kg, Cr(III) can become further enriched during weathering; Berger and

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Frei reported Cr enrichment within lateritic soil profiles in Madagascar with concentrations

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reaching 60 g/kg20, and Oze et al. found up to 10 g/Kg soil in a Californian serpentine soils21.

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Natural occurrence of hexavalent chromium has been reported in groundwater in pristine

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aquifers far from anthropogenic sources.5,7,22–25 High Cr(VI) levels are often present in aquifers



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surrounded by ophiolites and other ultramaphic rocks and have been reported along the western

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coast of North America4,5,7,8,24–29, southern Africa30, South America31, and Europe 6,9,32–34.

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Aqueous Cr(VI) has been speculatively linked to geogenic Cr(III) being oxidized by Mn

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oxides.35–37 Field-scale studies have showed Mn concentrations to be a good predictor of an

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aquifer’s capacity to form and solubilize Cr(VI)29,35, while microscale XRF and XANES

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spectroscopic approaches have revealed close spatial associations between Mn oxides and

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hexavalent chromium36. In most natural systems, Mn oxides are the only known compound

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capable of oxidizing Cr(III) to Cr(VI) at pH < 9.38,35,39–41 Contrasting Cr(III) oxidation, reduction of Cr(VI) in soils and sediments is common under

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oxygen-limiting (anaerobic) conditions. Iron(II), which is nearly ubiquitous in anaerobic soils

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and sediments, is a facile reductant of Cr(VI) that results in Cr(III)-Fe(III) hydroxides of limited

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solubility.42–44 The solubility of these phases decreases with increasing Fe substitution.10,45 In

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addition to Fe(II), sulfides and organic matter are also potential reductants of Cr(VI).45–47 Finally,

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a range of microorganisms (i.e., Pseudomonas, Desulfovibrio, Shewanella, Bacillus species) are

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capable of enzymatic Cr(VI) reduction under both aerobic and anaerobic conditions (ref 48, and

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references therein).

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Biogeochemical Constraints Imposed by Soil/Sediment Structure Having physical structure that results in a range of pore sizes, soils and sediments are

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often viewed as dual-pore domains, with inter-aggregate channels governed by advective flow

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and intra-aggregate flow dominated by diffusion.49,50 Diffusion and advection control

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biogeochemical networks and influence the spatial distribution and extent of redox processes.51–

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Due to high oxygen demand, microbial respiration limits O2 in all but the exterior few


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millimeters of the diffusive, intra-aggregate zones55,49,53, leading to anaerobic conditions even

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within seemingly well-oxygenated environments. The redox zones resulting from soil

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architecture control the cycling and fluxes of Mn, Fe, and Cr throughout soil and sediments.

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Under aerobic conditions, Fe and Mn precipitate as minerals of limited solubility. As redox

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conditions shift to anaerobic conditions, often due to increased inputs of organic matter,

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reductive dissolution mobilizes Fe(III) and Mn(III/IV) phases. Thus, within anaerobic aggregate

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interiors, conditions conducive to Cr(VI) reduction may prevail, including production of Fe(II)

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via dissimilatory iron reduction; outward diffusion of Fe(II) therefore has the potential to reduce

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Cr(VI).

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In order for Cr(III) to be oxidized by Mn oxides, one of the two phases must dissolve and

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migrate to the other solid.18 Cr(III) oxidation rates are proportional to the dissolved concentration

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of Cr(III) predicted from estimated mineral solubility.18 Therefore, within the structured media of

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soils and sediments, diffusion rates of Cr(III) phases dictate that the two solids must therefore be

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in close proximity (except under acidic conditions). The Cr-oxidizing capacity of Mn oxides is

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substantiated for well-mixed systems where transport limitations are minimized or

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eliminated.18,56–58 However, there are no studies on Cr(III) oxidation within structured,

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physically-rigid conditions found in soils and sediments, despite evidence that Cr(VI) genesis

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likely occurs within such environments.8,24 Understanding Cr cycling within these structured

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systems is vital for predicting the potential oxidation (or reduction) of chromium. The variation

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in geochemical conditions across the aggregated structure of soils and sediments leads to

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conditions potentially conducive to Cr(III) oxidation in the exterior region along flow paths

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while Cr(VI) reduction may proceed distal from oxygen supply, provided sufficient organic

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carbon is present.



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Here, we seek to understand the propensity for Cr(VI) formation by reaction of Cr(III)

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minerals with Mn oxides in fixed structured media characteristic of soils and sediments. Further,

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we investigate the propensity of anaerobic microsites to limit Cr(VI) concentration by promoting

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reduction.

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MATERIALS AND METHODS To assess physical constraints of Cr(VI) genesis by Mn oxides, synthetic aggregates of

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Cr(OH)3- and Cr0.25Fe0.75(OH)3-coated quartz grains were constructed and placed in flow-

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through reactors with synthetic groundwater medium. To mirror diffusion constraints within

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natural systems, we use synthetically constructed architecture to investigate whether low

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solubility Cr(III) and Mn(III/IV) minerals can interact within a fixed matrix to release Cr(VI) to

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advecting porewater at concentrations consistent with those observed in groundwater of Cr-

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bearing sediments. Previous studies illustrate that mass transfer within these synthetic aggregates

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is diffusion controlled, and that flow within the reaction cell does not result in advection within

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the aggregates.51–54 Variation in solubility between Cr(OH)3 and Cr0.25Fe0.75(OH)3 allowed us to

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compare the extent of mineral dissolution on Cr(III) oxidation within fixed media characteristic

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of soils and sediments. Most Mn oxides in the environment are layer-type MnO2 (e.g., birnessite)

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formed by biologically catalyzed reactions with oxygen.59–61 In this study, we investigate the

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oxidizing capacity of both synthetic birnessite and biogenic Mn oxides generated in situ by

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Leptothrix cholodnii, a well studied beta-proteobacteria capable of enzymatic oxidation of

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Mn(II) and Fe(II).62–64 Leptothrix sp. are most commonly found at aerobic/anaerobic interfaces

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where Fe and Mn are cycled between soluble and insoluble forms.65


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Mineral Synthesis. Chromium hydroxide was synthesized by titrating 20 mM CrCl3 to

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pH 6 with 0.1M NaOH and maintaining at this pH for 24 h. Cr0.25Fe0.75(OH)3 was synthesized by

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titrating stoichiometric concentrations of FeCl3 and CrCl3 solutions with 0.1M NaOH to pH 7

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and maintaining the pH value for 3 d, similar to the procedure described in Hansel et al.66.

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During titration, the pH was kept below 7.5. Suspensions were stored at 4° C and not allowed to

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age for more than 48 h. Cr(OH)3 and Cr0.25Fe0.75(OH)3 gels were centrifuged, rinsed three times

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with doubly deionized (DDI) water, and mixed with ground Iota quartz sand (Unimin

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Corporation, Spruce Pine, NC) (Cr(OH)3: 0.22mmol Cr g-1 sand; Cr0.25Fe0.75(OH)3: 0.05mmol Cr

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and 0.16mmol Fe g-1 sand; 106-125 μm quartz grain size). Sand grains were coated in 100 g

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batches and stirred over a 3-d period until completely dry before being rinsed repeatedly with

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DDI water until rinse water was free of particulates; the sand was then left to dry for an

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additional 3-d. Birnessite was synthesized by reducing KMnO4 based on McKenzie67, as outlined

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by Ying et al.5454 Minerals were confirmed with X-ray diffraction analysis on a rotating sample

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using a Rigaku Miniflex 600 diffractometer with Cu-Kα radiation fitted with a 1D silicon strip

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detector, and Cr(III):Fe(III) ratios were confirmed with X-ray fluorescence spectrometry

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(Spectro Xepos HE XRF Spectrometer) (Supporting Information, Figure S3).

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Microbial Inoculum. To investigate the effect of different functional microbial

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communities on Cr(VI) generation, two functionally diverse organisms were introduced to

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Cr0.25Fe0.75(OH)3-aggregates: Leptothrix cholodnii, an obligate aerobic heterotrophic bacterium

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capable of Mn(II) oxidation and known for its ability to precipitate Mn oxides68, and Shewanella

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sp. strain ANA-3, a facultative anaerobe that couples lactate oxidation with the reduction of a

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wide-variety of terminal electron acceptors (TEAs), including Fe(III) and Mn(III/IV) oxides69.

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Shewanella sp. ANA-3 was grown aerobically in autoclaved tryptic soy broth (30 g L-1 DDI



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water) at 25°C until late log phase from frozen seed culture (stored in 20% glycerol at -80°C).

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Leptothrix cholodnii was grown aerobically in liquid mineral salts-vitamin-pyruvate (MSVP)

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medium at 25°C containing the following ingredients (g/L): (NH4)2SO4 0.24; MgSO4*7H2O

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0.06; CaCl2*2H2O 0.06; KH2PO4 0.02; Na2HPO4 0.03; HEPES 2.383; FeSO4 0.002; C3H3NaO3

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1; and 1mL filter-sterilized Wolfe’s vitamin solution.70 All cells were harvested, washed by

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centrifuging liquid cultures (5000 x g; 15 min; 25°C), and re-suspended in 50 mL of sterile

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30mM HEPES- and 10mM sodium bicarbonate-buffered basal salts medium (BSM (g/L): KCl

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0.2; MgCl 0.05; NaCl 0.46; CaCl2*2H2O 0.06; KH2PO4 0.007) at pH 7.1 three times.

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Aggregate Synthesis and Biological Treatments. Three different biotic aggregate

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treatments were investigated: (1) Cr0.25Fe0.75(OH)3-coated sand inoculated with ~8x108 cells L.

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cholodnii g-1 sand, (2) Cr0.25Fe0.75(OH)3-coated sand mixed with birnessite (1:2.5 Mn:Cr molar

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ratio) and inoculated with ~8x108 cells S. sp. ANA3 g-1 sand, and finally (3) Cr0.25Fe0.75(OH)3-

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coated sand inoculated with ~8x108 cells L. cholodnii and ~8x108 cells S. sp. ANA3 g-1 sand.

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Abiotic aggregates were composed of either Cr(OH)3- or Cr0.25Fe0.75(OH)3-coated sand mixed

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with birnessite (1:10 Mn:Cr and 1:2.5 Mn:Cr molar ratio, respectively). In order to promote

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particle aggregation, all aggregates were made with 1% agarose (0.1g UltraPure agarose

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dissolved in 10 mL DDI water) and mixed thoroughly so all mineral phases and/or bacterial cells

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were homogenously distributed.52–54 The agarose sand mixture was then poured into sterile

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molds and formed into 30 x 15mm (height x diameter) cylinders.

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Flow-through reactor setup. Aggregates were placed in flow-through reactors with a

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volume of 77 mL (38x51mm; height x diameter) with 0.2 µm filters at inflow (bottom) and

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outflow (top) (Supporting Information, Figure S1). Styrene-butadiene rubber (ø = 17mm) capped

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circular planes of aggregates (3M™ Scotch-Weld™ Instant Adhesive CA5) in order to preclude


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vertical flow through aggregate cylinders. Sterile 5mm glass beads (80 (±2) g) were added to

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stabilize aggregate position. Two abiotic and three biotic treatments were investigated

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(Supporting Information, Table S1). Synthetic groundwater medium was pumped into reactor

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cells and advected around aggregates before being collected and analyzed. For all reactors, a

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synthetic groundwater media was used as the advecting solution consisting of: (in mg/L)

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CaCl2•2H2O 60; MgCl2•7H2O 50; KCl 200; NaCl 460; KH2PO4 7; NH4Cl 0.95. After

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autoclaving the groundwater media, the following filter sterilized solutions were added for a final

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concentration of: 30 mM HEPES, 10 mM NaHCO3, 0.23 mM MnCl2, and pH adjusted to 7.6

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with 6 M NaOH. A dual-buffered system was necessary to stabilize pH in the presence of

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microbial activity and was also used in abiotic systems for consistency. For the influent solution

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concentrations, rhodochrosite is oversaturated (SI = 2.1). However, to ensure precipitation of

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Mn(II) solids did not change Mn concentrations within influent solution prior to entering the

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reactor, Mn concentrations in the influent media were analyzed over the course of the

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experiment; concentrations were constant at 0.21 (±0.007) and 0.23 (±0.007) mM for the biotic

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and abiotic systems, respectively. For biotic reactors, 1 mL Wolfe’s trace mineral and vitamin

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solutions were added along with dissolved organic carbon as electron donor. Pyruvate and lactate

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were added for a final concentration of 3 mM each to represent dissolved organic carbon (DOC)

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levels of carbon-rich environments. Based on previous studies we expect little impact on Mn

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oxide dissolution at these DOC concentrations.71 The surrounding solute of all reactors was

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continuously sparged with filtered air. The solute flow eluted at a rate of 0.8 mL h-1 for biotic

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reactors and 0.6 mL h-1 for abiotic reactors. After 22 days, an acidified groundwater medium

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with sodium acetate buffer (30 mM, pH 5) consisting of: (in mg/L) CaCl2•2H2O 60; NaCl 460;



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KCl 200; NaCl 460; KH2PO4 7; NH4Cl 0.95, was pulsed through abiotic aggregates for 4 days

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before resuming initial synthetic groundwater composition (pH 8).

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Aqueous and solid phase analysis. Total Mn, Fe, and Cr concentrations were measured

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from filtered effluent using inductively coupled plasma mass spectrometry (Thermo Scientific

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XSERIES 2 ICP-MS). Quality control standards were analyzed every 15 samples to ensure a

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≤5% deviation from the standard curve was maintained. Unacidified filtrate was measured

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immediately for aqueous Cr(VI) concentrations using the diphenyl carbazide (DPC)

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spectrophotometric method.35 Aggregates for solid phase analysis were removed from reactors

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after aqueous Cr(VI) production had subsided (15 and 11 days for abiotic and biotic treatments,

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respectively). Additionally, separate abiotic aggregates (Cr(OH)3 (n=3); Cr0.25Fe0.75(OH)3 (n=2))

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were run for 50 days before being harvested and analyzed. Before removal from the flow cell,

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submerged aggregates were rotated 90° (after removing glass beads) and dissolved oxygen (DO)

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concentrations were measured in the middle of the aggregate (from exterior to interior) using a

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microsensor with a tip diameter of 10 μm (OX-10, Unisense). The tip, mounted on a motor-

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driven micromanipulator stage (MMS, Unisense) positioned via a motor controller (MC-232,

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Unisense), and connected to a picoammeter (Microsensor Multimeter, Unisense), was slowly

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lowered from aggregate exterior to interior. Linear calibrations were performed before each

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measurement in 0.1M sodium ascorbate in 0.1M NaOH (0% O2 saturation) and air bubbled water

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(100% O2 saturation). Each cylindrical aggregate was sectioned into three concentric semi-

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circles labeled as ‘E’ for exterior (0-2 mm), ‘M’ for midsection (2-4.5 mm), and ‘I’ for interior

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(4.5-7.5 mm) for three 4 mm slices representative of the bottom (2-6 mm), middle (18-22 mm),

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and top (24-29 mm) of the aggregate; a similar approach was used in previous studies to examine


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spatial variation in dissimilatory Fe(III) reduction52, As(V) reduction72, and Mn(IV)-Fe(III)

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reduction54.

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X-ray absorption spectroscopic (XAS) analysis was performed on dry sediments to

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determine Cr, Mn, and Fe speciation and quantify Fe phases. Acid digestion with 6 M HCl was

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used to quantify solid phase Cr, Mn, and Fe concentrations. All solids from Shewanella-

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inoculated aggregates were processed anoxically within a glove-bag atmosphere of

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95%N2:5%H2.

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Micro-X-ray fluorescence (μ-XRF) analysis of radial slices of each aggregate was carried out

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on beamline 2-3 and 10-2 at the Stanford Synchrotron Radiation Lightsource (SSRL) to map the

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spatial distribution of Cr, Mn, and Fe from the interior to exterior of the aggregate. Manually

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sliced 1 mm samples placed between Kapton tape were measured on BL10-2. Samples mapped

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on BL2-3 were dried, embedded in EPOTEK301-2FL epoxy, and then sectioned to 30 µm

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thickness and mounted on a quartz slide. The beam was calibrated by setting the position of the

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pre-edge absorption peak of Na2CrO4 at 5993eV. Cr maps were taken at three energies (5993,

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6003, 6010 eV) at 2-4 μm steps for high-resolution mapping. A Fe-beta window was used to

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subtract interfering intensities between Cr, Fe, and Mn from spectra collected with a vortex

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detector.

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For bulk XAS measurements, samples were dried, mixed with BN and pressed into

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pellets. Spectra were then collected on beamlines 11-2 and 4-1 at SSRL. A double-crystal, Si

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(220) LN-cooled monochromator was used for energy selection. Cr K-edge XANES spectra

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were collected with energy steps of 0.3 eV from 5969 to 6019 eV. Coarser steps were taken

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outside of this region for normalization purposes. Solid-phase iron was investigated using the

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extended portion (EXAFS) of the Fe K-edge spectrum; scans were obtained from 6882 to 7922



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eV, which is equivalent to a maximum k=15.2 Å-1. The XANES portion of the Fe spectra was

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taken with a resolution of 0.3 eV, with the monochromator detuned 20% to minimize higher-

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order harmonics, and spectra collected with 30-element Ge detector. Spectra were background

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subtracted, normalized, converted to k-space (Å-1) and k3 weighted. The x(k)k3 spectrum was

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Fourier-transformed over 0 to 10 Å-1. Then peaks were individually isolated and

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backtransformed. Qualitative analysis was performed by comparing unknowns to reference

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compounds.

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RESULTS Aqueous Cr(VI) effluent concentrations Synthetic soil aggregates (abiotic or bacterially inoculated) were placed within reactors

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and flow was initiated. Within reactors having abiotic aggregates composed of Cr(OH)3, or

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Cr0.25Fe0.75(OH)3, and birnessite, Cr(VI) was detected in advecting solutes within 5 h (Figure 1).

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Concentrations increased for the first 44 h of reaction, reaching a maximum concentration of ca.

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4 μM Cr(VI); concentrations then decreased steadily. Despite differences in expected solubility,

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abiotic aggregates having birnessite and Cr(OH)3 or Cr0.25Fe0.75(OH)3 generate similar amounts

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of Cr(VI) within the first 2 d of reaction; thereafter, the concentrations of Cr(VI) produced via

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oxidation of Cr(III) diverge, with the greater solubility of Cr(OH)3 leading to higher Cr(VI)

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concentrations (Figure 1). The cumulative mass of Cr(VI) eluted from the Cr0.25Fe0.75(OH)3

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reactor was 0.45 (± 0.05) μmole while the cumulative mass for the Cr(OH)3-containing reactor

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was almost double, 0.72 (± 0.02) μmole, prior to an acid injection (Figure 1).

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After an initial peak in Cr(VI) concentration at ca. 2 to 4 d, the Cr(VI) production

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steadily decreased. To test whether the decline in Cr(VI) production resulted from a passivating


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surface layer on the Mn oxides, potentially MnCO3, an acidified synthetic groundwater (pH 5)

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pulse was introduced after 23 days (Figure 1, grey bar). During the period of acidified influent,

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effluent Cr(VI) concentrations continue to decrease as the pH drops. Once effluent pH values

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decrease to that of influent pH (pH = 5), effluent Mn(II) concentrations spike, resulting in a

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concentration of 0.48 (± 0.14) mM for Cr(OH)3 and 0.37 (± 0.19) mM for Cr0.25Fe0.75(OH)3

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aggregates (Figure 1a). A secondary pulse of Cr(VI) then occurs subsequent to the Mn(II) pulse

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as the reactor pH returns to the initial pH (pH = 8).

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Within soils and subsurface sediments, Mn oxides are likely formed through biologically

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mediated processes. Using the Mn oxidizing bacterium L. cholodnii (hereafter referred to

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generally as Leptothrix), we tested for the production of Cr(VI) within the Cr(OH)3- and

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Cr0.25Fe0.75(OH)3-aggregate reactors having influent Mn(II) concentrations of 0.2 mM. Cr(VI)

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production within the Leptothrix-inoculated Cr0.25Fe0.75(OH)3 –aggregate quickly (41.4 h)

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reaches a peak concentration of 1.1 μM (Figure 2). Despite releasing only 11% of the cumulative

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Cr(VI) mass eluted from the abiotic birnessite-aggregate, peak Cr(VI) concentrations from

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microbial-inoculated aggregates reach over 26% those of the abiotic treatment (Figures 1 and 2).

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Owing to soil/sediment architecture, redox heterogeneity commonly prevails, leading to

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aggregates having anaerobic interiors and aerobic exteriors. To test for the influence of anaerobic

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zones proximal to aerobic regions on the production and efflux of Cr(VI), we utilized aggregates

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having either synthetic birnessite with the metal reducing bacterium Shewanella sp. ANA3

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(hereafter referred to more generally as Shewanella) or co-cultures of Leptothrix and Shewanella.

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The presence of metal reducing bacteria suppresses Cr(VI) elution within both abiogenic and

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biogenic Mn-oxides. When Shewanella is added to synthetic birnessite, only 18.5 nmoles of

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cumulative Cr(VI) is released (only 4.7% of Cr(VI) release from synthetic birnessite without iron



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reduction) (Figure 2b). In Leptothrix and Shewanella co-inoculated Cr0.25Fe0.75(OH)3-aggregates,

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Cr(VI) concentrations remain below the California drinking water standard of 10 µg/L

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throughout the experiment and cumulative release of Cr(VI) is just over half of the

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birnessite/Shewanella aggregate at 11.3 nmoles (Figure 2b).

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Oxygen Profiles Localized geochemical gradients created by diffusion-limited transport within soil

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aggregates restrict oxygen supply and can induce anaerobic conditions. Within oxygenated

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regions proximal to advective flow paths, obligate aerobic communities of Mn-oxidizing bacteria

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and fungi are able to colonize. Radial heterogeneity in Mn oxides and secondary Fe-minerals

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correlate with this geometric redox zonation resulting from oxygen gradients and associated

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metabolic activity (Figure 3; dissolved oxygen profiles are given in Supporting Information,

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Figure S4). In abiotic aggregates, with no microbial oxygen consumption, oxygen levels remain

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near saturation throughout the profile (Figure 3a; Figure S4). Oxygen penetration into aggregates

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decreases when Shewanella (a facultative anaerobe capable of Fe(III) and Mn(III/IV) reduction)

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is present (Figure 3c,d). The extent of oxygenation is shallowest when only Shewanella is

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present, suggesting that in the absence of competing obligate aerobes, Shewanella may consume

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oxygen more efficiently thereby limiting an aggregate’s oxic zone. In these aggregates, dissolved

306

oxygen (DO) levels drop below 200 μM just 1.5 mm below the exterior aggregate surface

307

(Figure 3d) and drop below detection limit (0.3 μM) 2.3 mm from the surface. With the addition

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of Leptothrix (aerobic, Mn-oxidizing bacteria) and Shewanella, DO levels greater than 200 μM

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are maintained only in the outer 3.9 mm of the aggregate (Figure 3c) and DO levels drop to

Hexavalent Chromium Generation within Naturally Structured Soils

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