This work was co-funded by BRGM, a postdoctoral grant from the Conseil G&#233;n&#233;ral du Loiret and the Carnot Institute. We also gratefully acknowledge the financial support provided to the PIVOTS project by the R&#233;gion Centre - Val de Loire.

1. Experimental Preparation
<ol>
	<li>Acid-wash all materials (glass, polytetrafluoroethylene (PFTE)) in contact with samples (5 days in 20% nitric acid (HNO3) v/v) followed by 5 days in hydrochloric acid (HCl) 10% v/v). Rinse several times with ultra-pure water and dry under a laminar flow hood prior to use.</li>
	<li>Use polyethylene gloves (or similar) and a fume hood for all steps involving chemicals.</li>
</ol>
2. Prepare Hg and As Spiked Amorphous Iron Oxides
<ol>
	<li>Prepare...

<ol>
	<li>Oremland, R. S., Stolz, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Ecology+of+Arsenic.">The Ecology of Arsenic.</a> Science. 300 (5621), 939 (2003).</li><li>Silver, S., Phung, L. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genes+and+enzymes+involved+in+bacterial+oxidation+and+reduction+of+inorganic+arsenic.">Genes and enzymes involved in bacterial oxidation and reduction of inorganic arsenic.</a> Appl Environ Microbiol. 71 (2), 599-608 (2005).</li><li>Compeau, G. C., Bartha, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sulfate-Reducing+Bacteria:+Principal+Methylators+of+Mercury+in+Anoxic+Estuarine+Sediment.">Sulfate-Reducing Bacteria: Principal Methylators of Mercury in Anoxic Estuarine Sediment.</a> Appl. Environ. Microbiol. 50, (1985).</li><li>Fleming, E. J., Mack, E. E., Green, P. G., Nelson, D. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mercury+Methylation+from+Unexpected+Sources:+Molybdate-Inhibited+Freshwater+Sediments+and+an+Iron-Reducing+Bacterium.">Mercury Methylation from Unexpected Sources: Molybdate-Inhibited Freshwater Sediments and an Iron-Reducing Bacterium.</a> Appl. Environ. Microbiol. 72 (1), 457-464 (2006).</li><li>Barkay, T., Miller, S., Summers, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bacterial+mercury+resistance+from+atoms+to+ecosystems.">Bacterial mercury resistance from atoms to ecosystems.</a> FEMS Microbiol Rev. 27 (2-3), 355-384 (2003).</li><li>Dixit, S., Hering, J. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+arsenic(V)+and+arsenic(III)+sorption+onto+iron+oxide+minerals:+Implications+for+arsenic+mobility.">Comparison of arsenic(V) and arsenic(III) sorption onto iron oxide minerals: Implications for arsenic mobility.</a> Environ. Sci. Technol. 37, (2003).</li><li>Andersson, H. A., Nriagu, J. O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The Biochemistry of Mercury in the Environnment. , 79-112 (1979).</li><li>Khwaja, A., Bloom, P. R., Brezonik, P. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Binding+Constants+of+Divalent+Mercury+in+Soil+Humic+Acids+and+Soil+Organic.">Binding Constants of Divalent Mercury in Soil Humic Acids and Soil Organic.</a> Environ. Sci. Technol. 40, (2006).</li><li>Neculita, C. M., Zagury, G. J., Deschenes, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mercury+Speciation+in+Highly+Contaminated+Soils+from+Chlor-Alkali+Plants+Using+Chemical+Extractions.">Mercury Speciation in Highly Contaminated Soils from Chlor-Alkali Plants Using Chemical Extractions.</a> J Environ Qual. 34 (1), (2005).</li><li>Schuster, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+behaviour+of+mercury+in+the+soil+with+special+emphasis+on+complexation+and+adsorption+processes+-+a+review+of+the+literature.">The behaviour of mercury in the soil with special emphasis on complexation and adsorption processes - a review of the literature.</a> Water Air Soil pollut. 56 (56), 667-680 (1991).</li><li>Wallschl&#228;ger, D., Desai, M. V. M., Spengler, M., Windm&#246;ller, C. C., Wilken, R. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+humic+substances+dominate+mercury+geochemistry+in+contaminated+floodplain+soils+and+sediments.">How humic substances dominate mercury geochemistry in contaminated floodplain soils and sediments.</a> J. Environ. Qual. 27 (5), (1998).</li><li>Lovley, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dissimilatory+Fe(III)+and+Mn(IV)+reduction.">Dissimilatory Fe(III) and Mn(IV) reduction.</a> Microbiol. Mol. Biol. Rev. 55 (2), 259-287 (1991).</li><li>Lovley, D. R., Kashefi, K., Vargas, M., Tor, J. M., Blunt-Harris, E. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reduction+of+humic+substances+and+Fe(III)+by+hyperthermophilic+microorganisms.">Reduction of humic substances and Fe(III) by hyperthermophilic microorganisms.</a> Chem. Geol. 169 (3-4), 289-298 (2000).</li><li>Hansel, C. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Structural+constraints+of+ferric+(hydr)oxides+on+dissimilatory+iron+reduction+and+the+fate+of+Fe(II).">Structural constraints of ferric (hydr)oxides on dissimilatory iron reduction and the fate of Fe(II).</a> Geochimica Cosmochimica Acta. 68, 3217-3229 (2004).</li><li>Thamdrup, B., Fossing, H., J&#248;rgensen, B. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Manganese,+iron+and+sulfur+cycling+in+a+coastal+marine+sediment+Aarhus+bay,+Denmark.">Manganese, iron and sulfur cycling in a coastal marine sediment Aarhus bay, Denmark.</a> Geochim.Cosmochim. Acta. 58 (23), 5115-5129 (1994).</li><li>Planer-Friedrich, B., London, J., McCleskey, R. B., Nordstrom, D. K., Wallschl&#228;ger, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thioarsenates+in+Geothermal+Waters+of+Yellowstone+National+Park:+Determination,+Preservation,+and+Geochemical.">Thioarsenates in Geothermal Waters of Yellowstone National Park: Determination, Preservation, and Geochemical.</a> Environ. Sci. Technol. 41 (15), 5245-5251 (2007).</li><li>Burnol, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decoupling+of+arsenic+and+iron+release+from+ferrihydrite+suspension+under+reducing+conditions:+a+biogeochemical+model.">Decoupling of arsenic and iron release from ferrihydrite suspension under reducing conditions: a biogeochemical model.</a> Geochem. Trans. 8 (1), 12 (2007).</li><li>Kocar, B. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Integrated+biogeochemical+and+hydrologic+processes+driving+arsenic+release+from+shallow+sediments+to+groundwaters+of+the+Mekong+delta.">Integrated biogeochemical and hydrologic processes driving arsenic release from shallow sediments to groundwaters of the Mekong delta.</a> Appl. Geochem. 23 (11), (2008).</li><li>Harris-Hellal, J., Grimaldi, M., Garnier-Zarli, E., Bousserrhine, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mercury+mobilization+by+chemical+and+microbial+iron+oxide+reduction+in+soils+of+French+Guyana.">Mercury mobilization by chemical and microbial iron oxide reduction in soils of French Guyana.</a> Biogeochem. 103 (1), (2011).</li><li>Islam, F. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+metal-reducing+bacteria+in+arsenic+release+from+Bengal+delta+sediments.">Role of metal-reducing bacteria in arsenic release from Bengal delta sediments.</a> Nature. 430, (2004).</li><li>Schultz-Zunkel, C., Rinklebe, J., Bork, H. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trace+element+release+patterns+from+three+floodplain+soils+under+simulated+oxidized-reduced+cycles.">Trace element release patterns from three floodplain soils under simulated oxidized-reduced cycles.</a> Ecol. Eng. 83, 485-495 (2015).</li><li>Nickson, R. T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+of+arsenic+release+to+groundwater,+bangladesh+and+West+Bengal.">Mechanisms of arsenic release to groundwater, bangladesh and West Bengal.</a> App. Geochem. 15, 403-413 (2000).</li><li>Varsanyi, I., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Arsenic,+iron+and+organic+matter+in+sediments+and+groundwater+in+the+Pannonian+basin,+Hungary.">Arsenic, iron and organic matter in sediments and groundwater in the Pannonian basin, Hungary.</a> App. Geochem. 21, 949-963 (2006).</li><li>Hellal, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mercury+mobilization+and+speciation+linked+to+bacterial+iron+oxide+and+sulfate+reduction:+A+column+study+to+mimic+reactive+transfer+in+an+anoxic+aquifer.">Mercury mobilization and speciation linked to bacterial iron oxide and sulfate reduction: A column study to mimic reactive transfer in an anoxic aquifer.</a> J. Contam. Hydrol. 180, 56-68 (2015).</li><li>Battaglia-Brunet, F., Dictor, M. C., Garrido, F., Crouzet, C., Morin, D., Dekeyser, K., Clarens, M., Baranger, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+arsenic(III)-oxidizing+bacterial+population:+selection,+characterization,+and+performance+in+reactors.">An arsenic(III)-oxidizing bacterial population: selection, characterization, and performance in reactors.</a> J Appl. Microbiol. 93 (2002), 656-667 (2002).</li><li>Salvato, N., Pirola, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+mercury+traces+by+means+of+solid+sample+atomic+absorption+spectrometry.">Analysis of mercury traces by means of solid sample atomic absorption spectrometry.</a> Microchim Acta. 123 (1), 63-71 (1996).</li><li>Huguet, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Caract&#233;risation biog&#233;ochimique et potentiel de m&#233;thylation du mercure de biofilms en milieu tropical (retenue de Petit Saut et estuaire du Sinnamary, Guyane Fran&#231;aise). . , (2009).</li><li>Mamindy-Pajany, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Arsenic+in+Marina+Sediments+from+the+Mediterranean+Coast:+Speciation+in+the+Solid+Phase+and+Occurrence+of+Thioarsenates.">Arsenic in Marina Sediments from the Mediterranean Coast: Speciation in the Solid Phase and Occurrence of Thioarsenates.</a> Soil Sed. Contam. 22, 984-1002 (2013).</li><li>dos Santos Afonso, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reductive+dissolution+of+iron(III)+(hydro)oxides+by+hydrogen+sulfide.">Reductive dissolution of iron(III) (hydro)oxides by hydrogen sulfide.</a> Langmuir. 8, 1671-1675 (1992).</li><li>Postma, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Redox+zonation:+equilibrium+constraints+on+the+Fe(III)/SO4-reduction+interface.">Redox zonation: equilibrium constraints on the Fe(III)/SO4-reduction interface.</a> Geochem Cosmochim. 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The experimental column setup proved to be a convenient laboratory device to study anaerobic biogeochemical processes in continuous conditions. Continuous column systems allow working in conditions closer to those of real aquifers than slurry batch systems or microcosms. Continuous systems can simulate the movement of groundwater through aquifer sediments.
The most critical step within the protocol is preparing the TE-iron (oxy)hydroxides and the mixture with silica gel and sand, which needs t...

Understanding and predicting trace element (TE) mobility and biogeochemistry in the environment is essential in order to monitor, develop, and apply appropriate management decisions for polluted sites. This especially applies in the case of toxic TEs such as arsenic (As) and mercury (Hg). The fate and speciation of these TEs in soil or aquifers are closely related to physico-chemical conditions, such as Eh and pH, but also to microbial activities that can play either a direct role on speciation or an indirect role on mobility.
Indeed, some bacteria can directly oxidize As(III) to As(V) or reduce As(V) to As(III). This affects As toxicity, since As(III) is the most toxic form of As, and mobility, since As(III) is more mobile than As(V), which can readily adsorb to iron (oxy)hydroxides or organic matter1,2. Likewise, bacteria are strongly involved in mercury cycling, either through its methylation, mainly by sulfate and iron reducing bacteria3,4, forming the neurotoxin monomethyl mercury (readily bioaccumulated in the food chain), or through its reduction to volatile elementary Hg (Hg&#176;)5.
Both As and Hg fates are also strongly linked to soil or aquifer composition, since compounds such as organic matter or iron (oxy)hydroxides can influence their sequestration and bioavailability. As(V) adsorbs well to iron (oxy)hydroxides6, whereas Hg has a very high affinity for organic matter (OM; mainly for thiol groups) but also for colloidal iron or manganese (oxy)hydroxides in OM depleted environments7,8,9,10,11.
Bacterial activities can then influence the fate of TEs adsorbed to (oxy)hydroxides or organic matter through the reduction of iron (oxy)hydroxides or the mineralization of organic matter. Direct iron reduction by bacteria is the dominant pathway for iron reduction in sulfur depleted zones12,13, Fe(III) being used as a terminal electron acceptor, whereas indirectly, Fe(III) can be reduced to Fe(II) by sulfide formed by a bacterial sulfate reduction14. Moreover, the presence of sulfate can also modify Hg and As speciation through the formation of complexes such as thio-arsenates15 with As or metacinnabar with Hg.
Thus, a better understanding of the impact of iron and sulfate cycling on the fate of TE, such as Hg and As, could help us to better manage contaminated sites and maintain soil and water quality. Data could also contribute to reinforcing existing metal-mobility models. Microbial Fe(III)-reduction16,17,18 can cause the desorption of TE. Theoretically, the indirect reduction of iron (oxy)hydroxides by sulfide produced by the microbial reduction of sulfate could also impact TE mobility. However, the extent and kinetics of these reactions are generally studied in batch homogenous systems or batch microcosms16,18,19,20. The drawback of batch experiments is the lack of dissociation of the occurring phenomena; indeed, activity is based on and limited by the resources present in the batch and only gives a final result of the shifts in speciation and adsorption. Using a column approach enables the renewal of inflowing media and the monitoring of the fate of TE over time and space. These conditions are more realistic when compared to an aquifer, where real phenomena are closely linked to continuous percolation conditions. Moreover, heterogeneous iron (oxy)hydroxide occurrence in aquifer sediments is common21,23, and the spatial changes in the mineralogical and chemical composition of the solid phases certainly drives microbial activities.
To elucidate the influence of these heterogeneities on geo-microbial phenomena and the fate of iron-associated TE, we developed a laboratory, a continuously-fed column representing a simplified model aquifer. The column is filled to create an iron-depleted zone at the column entrance and an iron-rich zone at the top. Regular sampling ports enable us to study each zone individually as well as interface-associated phenomena. An example of the application of this experimental device for the study of Hg fate and speciation is already available24. Here we give a detailed description of the experimental setup and a second example of its application focused on the behavior of As in contaminated aquifers.

<table border="0" cellpadding="0" cellspacing="0" width="991">
	<tbody>
		<tr height="120">
			<td height="120" style="height:120px;width:276px;">Glass columns&#160;</td>
			<td style="width:171px;">Beaucaverre, France</td>
			<td style="width:127px;">Specific request</td>
			<td style="width:417px;">columns were composed of 3 separate pieces, the main column core with the cooling jacket and the 5 sampling ports (size GL14 with olive) and a top and bottom piece that fits to the main column body and is held in place with a silicone joint and screw (RIN F 40x38 &#38; SVL 42). note: this design was discussed directly with the company. We recommend to find a local glazier.</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Septa PTFE/silicone diam 20mm&#160;</td>
			<td>Sigma-Aldrich&#160;</td>
			<td>508608</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">PTFE tubing ID 3mm</td>
			<td style="width:171px;">VWR</td>
			<td>228-0745</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Peristaltic pump</td>
			<td style="width:171px;">Dominique Dutsher SAS</td>
			<td style="width:127px;">66493</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Peristaltic pump tubing LMT 55</td>
			<td style="width:171px;">VWR</td>
			<td>224-2250</td>
			<td>Tygon&#174; LMT 55&#160;</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Fontainbleau sand&#160; D50=209 &#181;m</td>
			<td>SIBELCO,&#160; France</td>
			<td style="width:127px;"></td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:276px;">N2 for bubbling&#160;</td>
			<td style="width:171px;">Air Liquide, France</td>
			<td style="width:127px;"></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Gamma iradiation</td>
			<td>Ionisos, Dagneux, France</td>
			<td style="width:127px;"></td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Automatic Mercury Analyser (AMA 254, )</td>
			<td style="width:171px;">Courtage Analyses, France</td>
			<td style="width:127px;"></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Varian SpectrAA 300 Zeeman</td>
			<td style="width:171px;">Agilent&#160;</td>
			<td style="width:127px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:276px;">Name</td>
			<td style="width:171px;">Company</td>
			<td style="width:127px;">Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Chemicals</td>
			<td style="width:171px;"></td>
			<td style="width:127px;"></td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:276px;">HNO3 Supra pur</td>
			<td style="width:171px;">VWR</td>
			<td style="width:127px;">1.00441.1000</td>
			<td>Producer: Merck</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">HCL 30% Supra pur</td>
			<td style="width:171px;">VWR</td>
			<td>1.00318.1000</td>
			<td>Producer: Merck</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:276px;">Hg(NO3)2</td>
			<td style="width:171px;">Merck</td>
			<td>516953</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:276px;">As2O3</td>
			<td style="width:171px;">Merck</td>
			<td style="width:127px;">202673</td>
			<td></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:276px;">FeCl3-6H2O</td>
			<td>Merck</td>
			<td>207926</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">silica gel</td>
			<td>Sigma-Aldrich&#160;</td>
			<td>336815-500G</td>
			<td></td>
		</tr>
	</tbody>
</table>

experimental column setup for studying anaerobic biogeochemical interactions between iron (oxy)hydroxides, trace elements, and bacteria

Fate and speciation of trace elements (TEs), such as arsenic (As) and mercury (Hg), in aquifers are closely related to physio-chemical conditions, such as redox potential (Eh) and pH, but also to microbial activities that can play a direct or indirect role on speciation and/or mobility. Indeed, some bacteria can directly oxidize As(III) to As(V) or reduce As(V) to As(III). Likewise, bacteria are strongly involved in Hg cycling, either through its methylation, forming the neurotoxin monomethyl mercury, or through its reduction to elemental Hg&#176;. The fates of both As and Hg are also strongly linked to soil or aquifer composition; indeed, As and Hg can bind to organic compounds or (oxy)hydroxides, which will influence their mobility. In turn, bacterial activities such as iron (oxy)hydroxide reduction or organic matter mineralization can indirectly influence As and Hg sequestration. The presence of sulfate/sulfide can also strongly impact these particular elements through the formation of complexes such as thio-arsenates with As or metacinnabar with Hg.
Consequently, many important questions have been raised on the fate and speciation of As and Hg in the environment and how to limit their toxicity. However, due to their reactivity towards aquifer components, it is difficult to clearly dissociate the biogeochemical processes that occur and their different impacts on the fate of these TE.
To do so, we developed an original, experimental, column setup that mimics an aquifer with As- or Hg-iron-oxide rich areas versus iron depleted areas, enabling a better understanding of TE biogeochemistry in anoxic conditions. The following protocol gives step by step instructions for the column set-up either for As or Hg, as well as an example with As under iron and sulfate reducing conditions.

Example 1. Impact of iron reduction of As mobility and speciation 
The As column was directly inoculated with groundwater from a site presenting an As concentration higher than the drinking standards (Bracieux, Loire et Cher, France). Groundwater was sampled in sterile bottles, and stored at 5 &#176;C until use. The column was fed from the bottom with this water containing the natural endogenous microbial commun...

Watch this Scientific Journal Video about Experimental Column Setup for Studying Anaerobic Biogeochemical Interactions Between Iron OxyHydroxides, Trace Elements, and Bacteria at JoVE.com

Experimental Column Setup for Studying Anaerobic Biogeochemical Interactions Between Iron OxyHydroxides, Trace Elements, and Bacteria

Fate and speciation of arsenic and mercury in aquifers are closely related to physio-chemical conditions and microbial activity. Here, we present an original experimental column setup that mimics an aquifer and enables a better understanding of trace element biogeochemistry in anoxic conditions. Two examples are presented, combining geochemical and microbiological approaches.

Experimental Column Setup for Studying Anaerobic Biogeochemical Interactions Between Iron (Oxy)Hydroxides, Trace Elements, and Bacteria

Fate and speciation of trace elements (TEs), such as arsenic (As) and mercury (Hg), in aquifers are closely related to physio-chemical conditions, such as ...

The overall goal of this experimental setup is to study the fate of trace metals under sulfate and/or iron-reducing conditions induced by microbial activities. This method can help answer key questions in the biogeochemistry field such as interactions between key biogeochemical cycles like mercury and arsenic. The main advantage of this technique is that due to the continuous water flow, the dynamics of the biogeochemical reactions and lability of trace elements are dissociated and accessible through the sampling ports.And so, this method can provide insight into the fate of mercury and arsenic as demonstrated here ending to iron oxides. It can also be applied to other trace elements or carrier substrates. Demonstrating the procedure will Hafida Tris, a technician from our laboratory.To begin this procedure, add 500 milliliters of ultrapure water to a glass reactor equipped with a magnetic stir bar. Add 50 grams of iron(III)chloride hexahydrate while stirring. Manually add approximately 50 milliliters of a 10-molar solution of sodium hydroxide to begin precipitation of ferrihydrite.Next, adjust the pH to six and continue stirring for one hour to stabilize. For arsenic-spiked hydroxides and oxyhydroxides, prepare 100 milliliters of arsenic trioxide at a concentration of 10 grams per liter. Then, add 70 milliliters of this solution to the iron oxide solution.Continue stirring for three hours. After this, centrifuge at 2, 000 G for 20 minutes. Remove the supernatant and resuspend the pellet with 50 milliliters of ultrapure water.Repeat the centrifugation ane resuspension two additional times. Recover and store the humid hydroxides and oxyhydroxides as outlined in the text protocol. Keep four grams of silica gel in 40 milliliters of 7%potassium hydroxide on a hot plate stirring with a magnetic stir bar until dissolved.Add 60 milliliters of ultrapure water. Cool the solution on ice to about 20 degrees Celsius to avoid the gel setting too quickly. Titrate the silica gel mixture with phosphoric acid 20%to pH 7.5.Then, quickly mix the liquid silica gel with 320 grams of sterile sand previously mixed with 18.3 grams of either mercury-spiked or arsenic-spiked iron oxides before it solidifies. Using a spatula, break up the gelified mixture. To begin setting up the glass columns, connect the water jackets to a water-cooling system.Set the system to maintain an average temperature of 20 degrees Celsius. Cut lengths of PTFE tubing with a three-millimeter inner diameter making sure there is sufficient length for the column inlet and outlet. Then, add a layer of damp rock wool to the column from the top.Add 320 grams of sterile sand. Add the final layer consisting of the previously mixed sterile sand and spiked hydrated amorphous iron oxides fixed in the silica gel matrix. Next, connect the inflow to peristaltic tubing.Connect the other end of the peristaltic tubing to the water medium supply. Connect an ascendent flow of ultrapure sterile water continuously bubbled by nitrogen gas at low velocity of approximately two milliliters per hour. After this, cover the column with aluminum foil to block out the light.After two weeks, a black-colored precipitate is observed at the interface between the two layers of sand. This black zone progressively invades the top iron hydroxide and oxyhydroxide-enriched zones with the entire upper layer becoming black by the end of the experiment. After 35 days of continuous running, the measured levels of sulfate decrease followed by a transient increase in the total dissolved iron.From day 60 onward, the total dissolved arsenic is seen to rise significantly. On day 54, the physical and chemical parameters are profiled. While the pH remains close to pH seven along the column, the redox potential changes significantly with values nearing negative 200 millivolts in the iron-depleted zone where sulfate reduction is occurring and nearing negative millivolts at the top where bacterial and chemical iron reduction is occurring.Dissolved sulfate concentrations reach to approximately 20 milligrams per liter in the bottom layers and decrease to under one milligram per liter in the iron-rich zone. Sulfate also decreases sharply at the interface between iron-deprived and iron-rich zones indicating a peak of sulfate reduction activity. Meanwhile, arsenic is detected in the upper zone while thioarsenate species are seen closer to the interface and thiosulfate is in the bottom iron-deprived layer.These concentration profiles indicate a peak of sulfate reduction activity at the interface between iron-deprived and iron-rich layers. Following this procedure, other methods like scanning electron microscopy, Raman spectroscopy or activity assays can be performed in order to answer additional questions such as which new minerals are formed or which bacterial communities have developed. Don't forget that working with mercury and arsenic can be extremely hazardous and precautions such gloves and fume hoods should be used while performing this procedure.

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7.7K Views.  BRGM. The overall goal of this experimental setup is to study the fate of trace metals under sulfate and/or iron-reducing conditions induced by microbial activities. This method can help answer key questions in the biogeochemistry field such as interactions between key biogeochemical cycles like mercury and arsenic. The main advantage of this technique is that due to the continuous water flow, the dynamics of the biogeochemical reactions and lability of trace elements are dissociated and accessible...