Influence of humic acid imposed changes of ferrihydrite aggregation on microbial Fe(III) reduction Katja Amstaetter a,1 , Thomas Borch b , Andreas Kappler a,⇑ a Geomicrobiology, Center for Applied Geosciences, University of Tu ¨ bingen, Sigwartstrasse 10, D-72076 Tu ¨ bingen, Germany b Department of Soil and Crop Sciences and Department of Chemistry, Colorado State University, Fort Collins, CO 80523, USA Received 27 July 2011; accepted in revised form 1 February 2012; available online 21 February 2012 Abstract Microbial reduction of Fe(III) minerals at neutral pH is faced by the problem of electron transfer from the cells to the solid-phase electron acceptor and is thought to require either direct cell-mineral contact, the presence of Fe(III)-chelators or the presence of electron shuttles, e.g. dissolved or solid-phase humic substances (HS). In this study we investigated to which extent the ratio of Pahokee Peat Humic Acids (HA) to ferrihydrite in the presence and absence of phosphate influences rates of Fe(III) reduction by Shewanella oneidensis MR-1 and the identity of the minerals formed. We found that phosphate gen- erally decreased reduction rates by sorption to the ferrihydrite and surface site blocking. In the presence of low ferrihydrite concentrations (5 mM), the addition of HA helped to overcome this inhibiting effect by functioning as electron shuttle between cells and the ferrihydrite. In contrast, at high ferrihydrite concentrations (30 mM), the addition of HA did not lead to an increase but rather to a decrease in reduction rates. Confocal laser scanning microscopy images and ferrihydrite sedi- mentation behaviour suggest that the extent of ferrihydrite surface coating by HA influences the aggregation of the ferrihy- drite particles and thereby their accessibility for Fe(III)-reducing bacteria. We further conclude that in presence of dissolved HA, iron reduction is stimulated through electron shuttling while in the presence of only sorbed HA, no stimulation by elec- tron shuttling takes place. In presence of phosphate the stimulation effect did not occur until a minimum concentration of 10 mg/l of dissolved HA was reached followed by increasing Fe(III) reduction rates up to dissolved HA concentrations of approximately 240 mg/l above which the electron shuttling effect ceased. Not only Fe(III) reduction rates but also the mineral products changed in the presence of HA. Sequential extraction, XRD and 57 Fe-Mo ¨ ssbauer spectroscopy showed that crystal- linity and grain size of the magnetite produced by Fe(III) reduction in the presence of HA is lower than the magnetite pro- duced in the absence of HA. In summary, this study shows that both the concentration of HA and Fe(III) minerals strongly influence microbial Fe(III) reduction rates and the mineralogy of the reduction products. Thus, deviations in iron (hydr)oxide reactivity with changes in aggregation state, such as HA induced ferrihydrite aggregation, need to be considered within nat- ural environments. Ó 2012 Elsevier Ltd. All rights reserved. 1. INTRODUCTION Fe(II) and Fe(III) minerals such as siderite (Fe II CO 3 ), vivianite ðFe II 3 ðPO 4 Þ 2  8H 2 OÞ, magnetite ðFe II Fe III 2 O 4 Þ, ferrihydrite (Fe III (OH) 3 ), goethite (a-Fe III OOH), lepidocro- cite (c-Fe III OOH) or hematite ðc  Fe III 2 O 3 Þ are subject to a large number of chemical and biological redox transforma- tions in the environment. The biologically driven iron redox cycle includes Fe(III)-reducing as well as Fe(II)-oxidizing bacteria that inhabit a broad range of environments includ- ing, oxic, anoxic, microoxic, pH-neutral or acidic, ambient- temperature and hydrothermal conditions (Lovley et al., 2004; Kappler and Straub, 2005; Weber et al., 2006; Konhauser et al., 2011). Members of the genera Shewanella 0016-7037/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.gca.2012.02.003 ⇑ Corresponding author. Fax: +49 7071 295059. E-mail address: andreas.kappler@uni-tuebingen.de (A. Kappler). 1 Present address: CDM Consult GmbH, Neue Bergstrasse 13, D-64665 Alsbach, Germany. www.elsevier.com/locate/gca Available online at www.sciencedirect.com Geochimica et Cosmochimica Acta 85 (2012) 326–341