Bacterial DGGE fingerprints of biofilms on electrodes of membraneless microbial fuel cells Pierangela Cristiani a, * , Andrea Franzetti b , Isabella Gandolfi b , Edoardo Guerrini c , Giuseppina Bestetti b a RSE e Ricerca sul Sistema Energetico SpA, Environment and Sustainable Development Department, v. Rubattino 54, 20134 Milano, Italy b Dept. of Environmental Sciences (DISAT), University of Milano-Bicocca, Milan, Italy c Dept. of Chemistry, University of Milan, Milan, Italy article info Article history: Received 23 April 2012 Received in revised form 10 May 2012 Accepted 10 May 2012 Available online 13 November 2012 Keywords: Sulphate-reducing bacteria, SRB Sprirochetes Biocathode Microbial fuel cells Sulphur cycle DGGE fingerprint abstract Bacteria communities on electrodes of membraneless single chamber microbial fuel cells were sampled and analysed after one and three months of operation, using the fingerprinting molecular technique DGGE (Denaturing Gradient Gel Electrophoresis). The materials of the anodes were carbon brush, carbon cloth and stainless steel mesh (AISI 304). The cathodes were made of graphite, with or without Platinum as catalyst. The microbial fuel cells were inoculated with raw wastewater coming from a municipal plant of the city of Milan and fed with sodium acetate 3 g L 1 . DGGE profiles enabled to calculate the Jaccard similarity indexes for the bacterial communities. The excision and sequencing of selected bands permitted the characterization of the different communities and bacteria groups. The cluster analysis (based on band presence/absence and similarity data) showed that after one month the microbial populations of anodic biofilms diverged in relation to the material and the geometry of the anode. However, at the end of the three months of experimentation, the anodic communities did not signifi- cantly differed from those found at the cathodes. The results suggest a central role of Sulphate-reducing Bacteria (SRB) alone or in synergy with the other microorganisms found (Spirochetes and photosynthetic purple non-sulphur bacteria e PNS), at the cathode as well as at the anode. The mechanism of cathodic reaction activation by bacteria metabolism is put forward. Ó 2012 Elsevier Ltd. All rights reserved. 1. Introduction Microbial fuel cells (MFCs) are attracting considerable attention as innovative systems for energy production from renewable biomass and biomass-derived wastes. Potential applications include niche devices such as sediment-powered remote data collection. In an MFC, a microbial biofilm oxidises organic matter and transfers electrons from reduced compounds to an anode as the electron acceptor. The electrons then pass through a circuit and combine with a terminal electron acceptor (oxygen) and protons, giving water at the cathode. The closest analogues to anode for microbial metabolism in natural environments are Fe 3þ oxides, as both electrodes and Fe 3þ oxides are insoluble, extracellular electron acceptors. The investigation of how metal reducing bacteria use the oxides as electron acceptors led to the discovery of many mechanisms of extracellular electron transfer (EET). Bacteria can either produce extracellular mediators (shuttles) or directly transfer electrons by membrane cytochromes (Lovley, 2006). Further investigations revealed also a third potential mechanism which involves pili-like appendages able to transfer electric current, called nanowires. The ecology of bacteria and their electrogenic abilities have been extensively studied and recently reviewed (Lovley, 2006; Logan, 2009). The production of biosynthesized mediators have been described in Shewanella species, although direct contact and nanowire production have also been demonstrated (Gorby et al., 2006). Other organisms, such as Geothrix spp. and Pseudomonas spp. also produce electron shuttles. Conversely, Geobacter species seem to have evolved a specific strategy of direct contact to Fe 3þ oxides which allows these microorganisms to win the competition with other bacteria in open environment in which the production of exogenous mediation is a disadvantage (Lovley, 2006). In subsequent investigations several phylogenetically diverse bacteria were reported to be able to generate electricity in MFCs without the provision of an exogenous mediator. These include the well-known dissimilatory iron-reducing Geobacter spp. (Bond et al., 2002; Bond and Lovley, 2003), Shewanella spp. (Kim et al., 1999; Gorby et al., 2006) and other microorganisms such as Aeromonas hydrophila * Corresponding author. E-mail address: pierangela.cristiani@rse-web.it (P. Cristiani). Contents lists available at SciVerse ScienceDirect International Biodeterioration & Biodegradation journal homepage: www.elsevier.com/locate/ibiod 0964-8305/$ e see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.ibiod.2012.05.040 International Biodeterioration & Biodegradation 84 (2013) 211e219