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Title: NanoSIMS imaging reveals metabolic stratification within current-producing biofilms

Abstract

Metal-reducing bacteria direct electrons to their outer surfaces, where insoluble metal oxides or electrodes act as terminal electron acceptors, generating electrical current from anaerobic respiration. Geobacter sulfurreducens is a commonly enriched electricity-producing organism, forming thick conductive biofilms that magnify total activity by supporting respiration of cells not in direct contact with electrodes. Hypotheses explaining why these biofilms fail to produce higher current densities suggest inhibition by formation of pH, nutrient, or redox potential gradients; but these explanations are often contradictory, and a lack of direct measurements of cellular growth within biofilms prevents discrimination between these models. To address this fundamental question, we measured the anabolic activity of G. sulfurreducens biofilms using stable isotope probing coupled to nanoscale secondary ion mass spectrometry (nanoSIMS). Our results demonstrate that the most active cells are at the anode surface, and that this activity decreases with distance, reaching a minimum 10 µm from the electrode. Cells nearest the electrode continue to grow at their maximum rate in weeks-old biofilms 80-µm-thick, indicating nutrient or buffer diffusion into the biofilm is not rate-limiting. This pattern, where highest activity occurs at the electrode and declines with each cell layer, is present in thin biofilms (<5 µm) and fullymore » grown biofilms (>20 µm), and at different anode redox potentials. These results suggest a growth penalty is associated with respiring insoluble electron acceptors at micron distances, which has important implications for improving microbial electrochemical devices as well as our understanding of syntrophic associations harnessing the phenomenon of microbial conductivity.« less

Authors:
ORCiD logo; ORCiD logo; ORCiD logo; ORCiD logo; ORCiD logo
Publication Date:
Research Org.:
California Institute of Technology (CalTech), Pasadena, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER)
OSTI Identifier:
1565965
Alternate Identifier(s):
OSTI ID: 1625052
Grant/Contract Number:  
DE - SC0016469; SC0016469; NNA13AA92A; 542393; DEB 1542513; T32 GM007616; N000141612194
Resource Type:
Published Article
Journal Name:
Proceedings of the National Academy of Sciences of the United States of America
Additional Journal Information:
Journal Name: Proceedings of the National Academy of Sciences of the United States of America Journal Volume: 116 Journal Issue: 41; Journal ID: ISSN 0027-8424
Publisher:
Proceedings of the National Academy of Sciences
Country of Publication:
United States
Language:
English
Subject:
Science & Technology - Other Topics; Geobacter sulfurreducens; nanoSIMS; extracellular electron transfer; electrode reduction; stable isotope tracers

Citation Formats

Chadwick, Grayson L., Jiménez Otero, Fernanda, Gralnick, Jeffrey A., Bond, Daniel R., and Orphan, Victoria J. NanoSIMS imaging reveals metabolic stratification within current-producing biofilms. United States: N. p., 2019. Web. doi:10.1073/pnas.1912498116.
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Chadwick, Grayson L., Jiménez Otero, Fernanda, Gralnick, Jeffrey A., Bond, Daniel R., & Orphan, Victoria J. NanoSIMS imaging reveals metabolic stratification within current-producing biofilms. United States. doi:10.1073/pnas.1912498116.
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Chadwick, Grayson L., Jiménez Otero, Fernanda, Gralnick, Jeffrey A., Bond, Daniel R., and Orphan, Victoria J. Mon . "NanoSIMS imaging reveals metabolic stratification within current-producing biofilms". United States. doi:10.1073/pnas.1912498116.
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@article{osti_1565965,
title = {NanoSIMS imaging reveals metabolic stratification within current-producing biofilms},
author = {Chadwick, Grayson L. and Jiménez Otero, Fernanda and Gralnick, Jeffrey A. and Bond, Daniel R. and Orphan, Victoria J.},
abstractNote = {Metal-reducing bacteria direct electrons to their outer surfaces, where insoluble metal oxides or electrodes act as terminal electron acceptors, generating electrical current from anaerobic respiration. Geobacter sulfurreducens is a commonly enriched electricity-producing organism, forming thick conductive biofilms that magnify total activity by supporting respiration of cells not in direct contact with electrodes. Hypotheses explaining why these biofilms fail to produce higher current densities suggest inhibition by formation of pH, nutrient, or redox potential gradients; but these explanations are often contradictory, and a lack of direct measurements of cellular growth within biofilms prevents discrimination between these models. To address this fundamental question, we measured the anabolic activity of G. sulfurreducens biofilms using stable isotope probing coupled to nanoscale secondary ion mass spectrometry (nanoSIMS). Our results demonstrate that the most active cells are at the anode surface, and that this activity decreases with distance, reaching a minimum 10 µm from the electrode. Cells nearest the electrode continue to grow at their maximum rate in weeks-old biofilms 80-µm-thick, indicating nutrient or buffer diffusion into the biofilm is not rate-limiting. This pattern, where highest activity occurs at the electrode and declines with each cell layer, is present in thin biofilms (<5 µm) and fully grown biofilms (>20 µm), and at different anode redox potentials. These results suggest a growth penalty is associated with respiring insoluble electron acceptors at micron distances, which has important implications for improving microbial electrochemical devices as well as our understanding of syntrophic associations harnessing the phenomenon of microbial conductivity.},
doi = {10.1073/pnas.1912498116},
journal = {Proceedings of the National Academy of Sciences of the United States of America},
number = 41,
volume = 116,
place = {United States},
year = {2019},
month = {9}
}
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DOI: 10.1073/pnas.1912498116
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Works referenced in this record:

Proving Geobacter biofilm connectivity with confocal Raman microscopy
journal, May 2017

Look@NanoSIMS - a tool for the analysis of nanoSIMS data in environmental microbiology: Look@NanoSIMS
journal, January 2012

Long-range electron transport in Geobacter sulfurreducens biofilms is redox gradient-driven
journal, September 2012

  • Snider, R. M.; Strycharz-Glaven, S. M.; Tsoi, S. D.
  • Proceedings of the National Academy of Sciences, Vol. 109, Issue 38
  • DOI: 10.1073/pnas.1209829109

Reduction of low potential electron acceptors requires the CbcL inner membrane cytochrome of Geobacter sulfurreducens
journal, February 2016

Role of metal-reducing bacteria in arsenic release from Bengal delta sediments
journal, July 2004

  • Islam, Farhana S.; Gault, Andrew G.; Boothman, Christopher
  • Nature, Vol. 430, Issue 6995
  • DOI: 10.1038/nature02638

Biofilm and Nanowire Production Leads to Increased Current in Geobacter sulfurreducens Fuel Cells
journal, August 2006

  • Reguera, G.; Nevin, K. P.; Nicoll, J. S.
  • Applied and Environmental Microbiology, Vol. 72, Issue 11
  • DOI: 10.1128/AEM.01444-06

Temporal-Spatial Changes in Viabilities and Electrochemical Properties of Anode Biofilms
journal, April 2015

  • Sun, Dan; Cheng, Shaoan; Wang, Aijie
  • Environmental Science & Technology, Vol. 49, Issue 8
  • DOI: 10.1021/acs.est.5b00175

Influence of hydrodynamics and nutrients on biofilm structure
journal, December 1998

Arsenic Mobilization through Microbially Mediated Deflocculation of Ferrihydrite
journal, May 2005

  • Tadanier, Christopher J.; Schreiber, Madeline E.; Roller, Jonathan W.
  • Environmental Science & Technology, Vol. 39, Issue 9
  • DOI: 10.1021/es048206d

Heavy water and 15 N labelling with NanoSIMS analysis reveals growth rate-dependent metabolic heterogeneity in chemostats : NanoSIMS analysis of metabolic diversity in chemostats
journal, March 2015

  • Kopf, Sebastian H.; McGlynn, Shawn E.; Green-Saxena, Abigail
  • Environmental Microbiology, Vol. 17, Issue 7
  • DOI: 10.1111/1462-2920.12752

Bioelectrochemical Systems: An Outlook for Practical Applications
journal, June 2012

  • Sleutels, Tom H. J. A.; Ter Heijne, Annemiek; Buisman, Cees J. N.
  • ChemSusChem, Vol. 5, Issue 6
  • DOI: 10.1002/cssc.201100732

Structure and mechanism of a Hypr GGDEF enzyme that activates cGAMP signaling to control extracellular metal respiration
journal, April 2019

Geobacter Dominates the Inner Layers of a Stratified Biofilm on a Fluidized Anode During Brewery Wastewater Treatment
journal, March 2018

  • Tejedor-Sanz, Sara; Fernández-Labrador, Patricia; Hart, Steven
  • Frontiers in Microbiology, Vol. 9
  • DOI: 10.3389/fmicb.2018.00378

Voltammetry and Growth Physiology of Geobacter sulfurreducens Biofilms as a Function of Growth Stage and Imposed Electrode Potential
journal, March 2010

  • Marsili, Enrico; Sun, Jian; Bond, Daniel R.
  • Electroanalysis, Vol. 22, Issue 7-8
  • DOI: 10.1002/elan.200800007

Real-Time Spatial Gene Expression Analysis within Current-Producing Biofilms
journal, May 2012

  • Franks, Ashley E.; Glaven, Richard H.; Lovley, Derek R.
  • ChemSusChem, Vol. 5, Issue 6
  • DOI: 10.1002/cssc.201100714

Microbial Communities Associated with Electrodes Harvesting Electricity from a Variety of Aquatic Sediments
journal, June 2004

Effects of biofilm structures on oxygen distribution and mass transport
journal, May 1994

  • de Beer, Dirk; Stoodley, Paul; Roe, Frank
  • Biotechnology and Bioengineering, Vol. 43, Issue 11
  • DOI: 10.1002/bit.260431118

Electrode-Reducing Microorganisms That Harvest Energy from Marine Sediments
journal, January 2002

Redox potential as a master variable controlling pathways of metal reduction by Geobacter sulfurreducens
journal, January 2017

  • Levar, Caleb E.; Hoffman, Colleen L.; Dunshee, Aubrey J.
  • The ISME Journal, Vol. 11, Issue 3
  • DOI: 10.1038/ismej.2016.146

Trace incorporation of heavy water reveals slow and heterogeneous pathogen growth rates in cystic fibrosis sputum
journal, December 2015

  • Kopf, Sebastian H.; Sessions, Alex L.; Cowley, Elise S.
  • Proceedings of the National Academy of Sciences, Vol. 113, Issue 2
  • DOI: 10.1073/pnas.1512057112

Single cell activity reveals direct electron transfer in methanotrophic consortia
journal, September 2015

  • McGlynn, Shawn E.; Chadwick, Grayson L.; Kempes, Christopher P.
  • Nature, Vol. 526, Issue 7574
  • DOI: 10.1038/nature15512

Conduction-based modeling of the biofilm anode of a microbial fuel cell
journal, December 2007

  • Kato Marcus, Andrew; Torres, César I.; Rittmann, Bruce E.
  • Biotechnology and Bioengineering, Vol. 98, Issue 6
  • DOI: 10.1002/bit.21533

Biofilms as Complex Differentiated Communities
journal, October 2002

Visible spectroelectrochemical characterization of Geobacter sulfurreducens biofilms on optically transparent indium tin oxide electrode
journal, December 2011

Evaluation of energy-conversion efficiencies in microbial fuel cells (MFCs) utilizing fermentable and non-fermentable substrates
journal, March 2008

Mass transport to streambed biofilms
journal, June 1988

Direct Exchange of Electrons Within Aggregates of an Evolved Syntrophic Coculture of Anaerobic Bacteria
journal, December 2010

  • Summers, Z. M.; Fogarty, H. E.; Leang, C.
  • Science, Vol. 330, Issue 6009, p. 1413-1415
  • DOI: 10.1126/science.1196526

Propidium ion enters viable cells with high membrane potential during live-dead staining
journal, November 2017

Limitations for Current Production in Geobacter sulfurreducens Biofilms
journal, February 2013

  • Bonanni, P. Sebastian; Bradley, Dan F.; Schrott, Germán D.
  • ChemSusChem, Vol. 6, Issue 4
  • DOI: 10.1002/cssc.201200671

Current density reversibly alters metabolic spatial structure of exoelectrogenic anode biofilms
journal, July 2017

Comparison of anode bacterial communities and performance in microbial fuel cells with different electron donors
journal, September 2007

Microtoming coupled to microarray analysis to evaluate the spatial metabolic status of Geobacter sulfurreducens biofilms
journal, December 2009

  • Franks, Ashley E.; Nevin, Kelly P.; Glaven, Richard H.
  • The ISME Journal, Vol. 4, Issue 4
  • DOI: 10.1038/ismej.2009.137

Natural acidophilic biofilm communities reflect distinct organismal and functional organization
journal, October 2008

  • Wilmes, Paul; Remis, Jonathan P.; Hwang, Mona
  • The ISME Journal, Vol. 3, Issue 2
  • DOI: 10.1038/ismej.2008.90

Micro-scale observations of the structure of aerobic microbial granules used for the treatment of nutrient-rich industrial wastewater
journal, February 2008

  • Lemaire, Romain; Webb, Richard I.; Yuan, Zhiguo
  • The ISME Journal, Vol. 2, Issue 5
  • DOI: 10.1038/ismej.2008.12

Spatiometabolic Stratification of Shewanella oneidensis Biofilms
journal, August 2006

  • Teal, T. K.; Lies, D. P.; Wold, B. J.
  • Applied and Environmental Microbiology, Vol. 72, Issue 11
  • DOI: 10.1128/AEM.01163-06

Pseudomonas aeruginosa Displays Multiple Phenotypes during Development as a Biofilm
journal, February 2002

pH, redox potential and local biofilm potential microenvironments within Geobacter sulfurreducens biofilms and their roles in electron transfer
journal, May 2012

  • Babauta, Jerome T.; Nguyen, Hung Duc; Harrington, Timothy D.
  • Biotechnology and Bioengineering, Vol. 109, Issue 10
  • DOI: 10.1002/bit.24538

Desmids and Biofilms of Freshwater Wetlands: Development and Microarchitecture
journal, April 2007

Identification of an Extracellular Polysaccharide Network Essential for Cytochrome Anchoring and Biofilm Formation in Geobacter sulfurreducens
journal, December 2010

  • Rollefson, J. B.; Stephen, C. S.; Tien, M.
  • Journal of Bacteriology, Vol. 193, Issue 5
  • DOI: 10.1128/JB.01092-10

Proton transport inside the biofilm limits electrical current generation by anode-respiring bacteria
journal, August 2008

  • Torres, César I.; Kato Marcus, Andrew; Rittmann, Bruce E.
  • Biotechnology and Bioengineering, Vol. 100, Issue 5
  • DOI: 10.1002/bit.21821

Mechanistic stratification in electroactive biofilms of Geobacter sulfurreducens mediated by pilus nanowires
journal, August 2016

  • Steidl, Rebecca J.; Lampa-Pastirk, Sanela; Reguera, Gemma
  • Nature Communications, Vol. 7, Issue 1
  • DOI: 10.1038/ncomms12217

Physiological Stratification in Electricity-Producing Biofilms of Geobacter sulfurreducens
journal, December 2013

  • Schrott, Germán David; Ordoñez, María Victoria; Robuschi, Luciana
  • ChemSusChem, Vol. 7, Issue 2
  • DOI: 10.1002/cssc.201300605

Microbial Biofilm Voltammetry: Direct Electrochemical Characterization of Catalytic Electrode-Attached Biofilms
journal, October 2008

  • Marsili, E.; Rollefson, J. B.; Baron, D. B.
  • Applied and Environmental Microbiology, Vol. 74, Issue 23
  • DOI: 10.1128/AEM.00177-08

Abundance of the Multiheme c-Type Cytochrome OmcB Increases in Outer Biofilm Layers of Electrode-Grown Geobacter sulfurreducens
journal, August 2014

Harnessing microbially generated power on the seafloor
journal, July 2002

  • Tender, Leonard M.; Reimers, Clare E.; Stecher, Hilmar A.
  • Nature Biotechnology, Vol. 20, Issue 8
  • DOI: 10.1038/nbt716

Spatially Resolved Confocal Resonant Raman Microscopic Analysis of Anode-Grown Geobacter sulfurreducens Biofilms
journal, January 2014

  • Lebedev, Nikolai; Strycharz-Glaven, Sarah M.; Tender, Leonard M.
  • ChemPhysChem, Vol. 15, Issue 2
  • DOI: 10.1002/cphc.201300984

Structure of Microbial Nanowires Reveals Stacked Hemes that Transport Electrons over Micrometers
journal, April 2019

The effect of biofilm thickness on electrochemical activity of Geobacter sulfurreducens
journal, October 2016

An Inner Membrane Cytochrome Required Only for Reduction of High Redox Potential Extracellular Electron Acceptors
journal, November 2014

Metabolic spatial variability in electrode-respiring Geobacter sulfurreducens biofilms
journal, January 2013

  • Renslow, R. S.; Babauta, J. T.; Dohnalkova, A. C.
  • Energy & Environmental Science, Vol. 6, Issue 6
  • DOI: 10.1039/c3ee40203g

Intercellular wiring enables electron transfer between methanotrophic archaea and bacteria
journal, October 2015

  • Wegener, Gunter; Krukenberg, Viola; Riedel, Dietmar
  • Nature, Vol. 526, Issue 7574
  • DOI: 10.1038/nature15733

Biofilms: an emergent form of bacterial life
journal, August 2016

  • Flemming, Hans-Curt; Wingender, Jost; Szewzyk, Ulrich
  • Nature Reviews Microbiology, Vol. 14, Issue 9
  • DOI: 10.1038/nrmicro.2016.94

Linking Spectral and Electrochemical Analysis to Monitor c-type Cytochrome Redox Status in Living Geobacter sulfurreducens Biofilms
journal, June 2011

Contributions of microbial biofilms to ecosystem processes in stream mesocosms
journal, November 2003

  • Battin, Tom J.; Kaplan, Louis A.; Denis Newbold, J.
  • Nature, Vol. 426, Issue 6965
  • DOI: 10.1038/nature02152

Spectroscopic Slicing to Reveal Internal Redox Gradients in Electricity-Producing Biofilms
journal, November 2012

  • Robuschi, Luciana; Tomba, J. Pablo; Schrott, Germán D.
  • Angewandte Chemie International Edition, Vol. 52, Issue 3
  • DOI: 10.1002/anie.201205440
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