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Operational temperature regulates anodic biofilm growth and the development of electrogenic activity


The operational temperature of microbial fuel cell reactors influences biofilm development, and this has an impact on anodic biocatalytic activity. In this study, we compared three microbial fuel cell (MFC) reactors acclimated at 10°C, 20°C and 35°C to investigate the effect on biomass development, methanogenesis and electrogenic activity over time. The start-up time was inversely influenced by temperature, but the amount of biomass accumulation increased with increased temperatures, the 10°C, 20°C and 35°C acclimated biofilms resulted in 0.57, 0.82 and 5.43 g biomass (volatile suspended solids) per litre respectively at 56 weeks of operation. Biofilm build-up on the 35°C anode was further demonstrated by scanning electron microscopy, which showed large aggregations of biomass accumulating on the anode when compared to 10°C and 20°C biofilms. Biomass accumulation had a direct impact on biocatalytic performance, with the maximum power at 35°C after 60 weeks of operation being 2.14 W m−3 and power densities for the 10°C and 20°C reactors being and 4.29 W m−3. Methanogenic activity was also shown to be higher at 35°C, with a rate of 10.1 mmol CH4 biofilm per gram of volatile suspended solid (VSS) per day, compared to 0.28 mmol CH4 per gram of VSS per day produced at 20°C. These results demonstrate that higher MFC operating temperatures could be detrimental to the biocatalytic performance of electrochemically active bacteria in anodic biofilms due to biomass accumulation with enhanced development of non-electrogenic communities (e.g. methanogens and fermenters), meaning that, over time, psychro- or mesophilic operation can have beneficial effects for the development of electrogenically active populations in the reactor.

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  1. Aelterman P, Freguia S, Keller J, Verstraete W, Rabaey K (2008) The anode potential regulates bacterial activity in microbial fuel cells. Appl Microbiol Biotechnol 78(3):409–418. doi:

  2. Ahn Y, Logan BE (2010) Effectiveness of domestic wastewater treatment using microbial fuel cells at ambient and mesophilic temperatures. Bioresour Technol 101:469–475

  3. Amato P, Christner BC (2009) Energy metabolism response to low-temperature and frozen conditions in Psychrobacter cryohalolentis. Appl Environ Microbiol 75(3):711–718. doi:

  4. Bard AJ, Faulkner LR (2001) Electrochemical methods: fundamentals and applications, 2nd edn. John Wiley, New York

  5. Boe-Hansen R, Albrechtsen HJ, Arvin E, Jorgensen C (2002) Bulk water phase and biofilm growth in drinking water at low nutrient conditions. Water Res 36(18):4477–4486

  6. Cheng S, Xing D, Logan BE (2010) Electricity generation of single-chamber microbial fuel cells at low temperatures. Biosens Bioelectron 26(5):1913–1917

  7. Elferink SJWHO, Rinia HA, Bruins ME, Vos WMD, Stams AJM (1997) Detection and quantification of Desulforhabdus amnigenus in anaerobic granular sludge by dot blot hybridization and PCR amplification. J Appl Microbiol 83:102–110

  8. Flemming H-C, Szewzyk U, Griebe T (2000) Biofilms: investigative methods & applications. Technomic Pub. Co, Lancaster, Pa

  9. Franks AE, Nevin KP, Jia H, Izallalen M, Woodard TL, Lovley DR (2009) Novel strategy for three-dimensional real-time imaging of microbial fuel cell communities: monitoring the inhibitory effects of proton accumulation within the anode biofilm. Energy & Environmental Science 2(1):113–119

  10. Freguia S, Rabaey K, Yuan ZG, Keller J (2007) Electron and carbon balances in microbial fuel cells reveal temporary bacterial storage behavior during electricity generation. Environ Sci Technol 41(8):2915–2921. doi:

  11. Gorby YA, Yanina S, McLean JS, Rosso KM, Moyles D, Dohnalkova A, Beveridge TJ, Chang IS, Kim BH, Kim KS, Culley DE, Reed SB, Romine MF, Saffarini DA, Hill EA, Shi L, Elias DA, Kennedy DW, Pinchuk G, Watanabe K, Si I, Logan B, Nealson KH, Fredrickson JK (2006) Electrically conductive bacterial nanowires produced by Shewanella oneidensis strain MR-1 and other microorganisms. Proceedings of the National Academy of Sciences 103(30):11358–11363

  12. Hall-Stoodley L, Costerton JW, Stoodley P (2004) Bacterial biofilms: from the Natural environment to infectious diseases. Nat Rev Microbiol 2:95–108

  13. Jadhav GS, Ghangrekar MM (2009) Performance of microbial fuel cell subjected to variation in pH, temperature, external load and substrate concentration. Bioresour Technol 100:717–723

  14. Kim J, Lee C, Shin S, Hwang S (2008) Correlation of microbial mass with ATP and DNA concentrations in acidogenesis of whey permeate. Biodegradation 19(2):187–195

  15. Kim JR, Premier GC, Hawkes FR, Dinsdale RM, Guwy AJ (2009) Development of a tubular microbial fuel cell (MFC) employing a membrane electrode assembly cathode. J Power Sources 187:393–399

  16. Kim JR, Premier GC, Hawkes FR, Rodríguez J, Dinsdale RM, Guwy AJ (2010) Modular tubular microbial fuel cells for energy recovery during sucrose wastewater treatment at low organic loading rate. Bioresour Technol 101(4):1190–1198

  17. Kim JR, Rodriguez J, Hawkes FR, Dinsdale RM, Guwy AJ, Premier GC (2011) Increasing power recovery and organic removal efficiency using extended longitudinal tubular microbial fuel cell (MFC) reactors. Energy & Environmental Science 4(2):459–465

  18. Knoblauch C, Jorgensen BB, Harder J (1999) Community size and metabolic rates of psychrophilic sulfate-reducing bacteria in arctic marine sediments. Appl Environ Microbiol 65(9):4230–4233

  19. Lee H-S, Parameswaran P, Kato-Marcus A, César I, Torres, Rittmann BE (2008) Evaluation of energy-conversion efficiencies in microbial fuel cells (MFCs) utilizing fermentable and non-fermentable substrates. Water Res 42:1501–1510

  20. Lies DP, Hernandez ME, Kappler A, Mielke RE, Gralnick JA, Newman DK (2005) Shewanella oneidensis MR-1 uses overlapping pathways for iron reduction at a distance and by direct contact under conditions relevant for biofilms. Appl Environ Microbiol 71(8):4414–4426

  21. Liu Y, Harnisch F, Fricke K, Schroder U, Climent V, Feliu JM (2010) The study of electrochemically active microbial biofilms on different carbon-based anode materials in microbial fuel cells. Biosens Bioelectron 25(9):2167–2171. doi:

  22. Logan BE (2008) Microbial fuel cells. Wiley-Interscience, Hoboken

  23. Logan BE, Hamelers B, Rozendal R, Schroeder U, Keller J, Freguia S, Aelterman P, Verstraete W, Rabaey K (2006) Microbial fuel cells: methodology and technology. Environ Sci Technol 40(17):5181–5192

  24. Low EW, Chase HA (1999) The effect of maintenance energy requirements on biomass production during wastewater treatment. Water Res 33(3):847–853

  25. Marstorp H, Guan X, Gong P (2000) Relationship between dsDNA, chloroform labile C and ergosterol in soils of different organic matter contents and pH. Soil Biology and Biochemistry 32(6):879–882

  26. Marstorp H, Witter E (1999) Extractable dsDNA and product formation as measures of microbial growth in soil upon substrate addition. Soil Biology & Biochemistry 31(10):1443–1453

  27. McCoy WF, Olson BH (1985) Fluorometric determination of the DNA concentration in municipal drinking water. Appl Environ Microbiol 49(4):811–817

  28. Michie IS, Kim JR, Dinsdale RM, Guwy AJ, Premier GC (2011) The influence of psychrophilic and mesophilic start-up temperature on microbial fuel cell system performance. Energy Environ Sci 4: 1011–1019. doi:

  29. Moon H, Chang IS, Kim BH (2006) Continuous electricity production from artificial wastewater using a mediator-less microbial fuel cell. Bioresour Technol 97:621–627

  30. Oh ST, Kim JR, Premier GC, Lee TH, Kim C, Sloan WT (2010) Sustainable wastewater treatment: How might microbial fuel cells contribute. Biotechnol Adv 28(6):871–881. doi:

  31. Pant D, Singh A, Van Bogaert G, Gallego YA, Diels L, Vanbroekhoven K (2011) An introduction to the life cycle assessment (LCA) of bioelectrochemical systems (BES) for sustainable energy and product generation: relevance and key aspects. Renewable & Sustainable Energy Reviews 15(2):1305–1313. doi:

  32. Pant D, Van Bogaert G, Diels L, Vanbroekhoven K (2010) A review of the substrates used in microbial fuel cells (MFCs) for sustainable energy production. Bioresour Technol 101(6):1533–1543. doi:

  33. Patil SA, Harnisch F, Kapadnis B, Schroder U (2010) Electroactive mixed culture biofilms in microbial bioelectrochemical systems: the role of temperature for biofilm formation and performance. Biosens Bioelectron 26(2):803–808. doi:

  34. Pham HT, Boon N, Aelterman P, Clauwaert P, De Schamphelaire L, Van Oostveldt P, Verbeken K, Rabaey K, Verstraete W (2008) High shear enrichment improves the performance of the anodophilic microbial consortium in a microbial fuel cell. Microb Biotechnol 1(6):487–496. doi:

  35. Picioreanu C, Head IM, Katuri KP, van Loosdrecht MCM, Scott K (2007) A computational model for biofilm-based microbial fuel cells. Water Res 41(13):2921–2940. doi:

  36. Premier GC, Kim JR, Michie I, Dinsdale RM, Guwy AJ (2011) Automatic control of load increases power and efficiency in a microbial fuel cell. J Power Sources 196(4):2013–2019

  37. Ramasamy RP, Ren Z, Mench MM, Regan JM (2008) Impact of initial biofilm growth on the anode impedance of microbial fuel cells. Biotechnol Bioeng 101(1)

  38. Reguera G, Nevin KP, Nicoll JS, Covalla SF, Woodard TL, Lovley DR (2006) Biofilm and nanowire production leads to increased current in Geobacter sulfurreducens fuel cells. Appl Environ Microbiol 72(11):7345–7348. doi:

  39. Ren ZY, Ramasamy RP, Cloud-Owen SR, Yan HJ, Mench MM, Regan JM (2011) Time-course correlation of biofilm properties and electrochemical performance in single-chamber microbial fuel cells. Bioresour Technol 102(1):416–421. doi:

  40. Stams AJM, Bok FAMD, Plugge CM, Eekert MHAV, Dolfing J, Schraa G (2006) Exocellular electron transfer in anaerobic microbial communities. Environ Microbiol 8(3):371–382

  41. Stenstrom J, Stenberg B, Johansson M (1998) Kinetics of substrate-induced respiration (SIR): theory. Ambio 27(1):35–39

  42. Summers ZM, Fogarty HE, Leang C, Franks AE, Malvankar NS, Lovley DR (2010) Direct exchange of electrons within aggregates of an evolved syntrophic coculture of anaerobic bacteria. Science 330(6009):1413–1415. doi:

  43. Torres CI, Marcus AK, Lee H-S, Parameswaran P, Krajmalnik-Brown R, Rittmann BE (2009) A kinetic perspective on extracellular electron transfer by anode-respiring bacteria. FEMS Microbiol Rev 34(1):3–17

  44. Whitchurch CB, Tolker-Nielsen T, Ragas PC, Mattick JS (2002) Extracellular DNA required for bacterial biofilm formation. Science 295(5559):1487. doi:

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This research was funded by the RCUK Energy Programme, SUPERGEN Biological Fuel Cell project (EP/D047943/1 and EP/H019480/1). The Energy Programme is an RCUK cross-council initiative led by EPSRC and contributed to by ESRC, NERC, BBSRC and STFC.

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Correspondence to Iain S. Michie.

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Michie, I.S., Kim, J.R., Dinsdale, R.M. et al. Operational temperature regulates anodic biofilm growth and the development of electrogenic activity. Appl Microbiol Biotechnol 92, 419–430 (2011).

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  • Microbial fuel cell (MFC)
  • Tubular reactor
  • Biofilm
  • Temperature
  • Electricity generation