Name: self-standing electrochemical set-up to enrich anode-respiring bacteria on-site
Uploaded: 2018-07-24T13:00:00
Description: Watch this Scientific Journal Video about Self-standing Electrochemical Set-up to Enrich Anode-respiring Bacteria On-site at JoVE.com

We would like to acknowledge Roger Raiche and David McCrory for allowing us access to the Cedars and consulting on locations for long term incubation. We also thank the Cedars field crew during the 2013-2014 season: Shino Suzuki, Shunichi Ishii, Greg Wanger, Grayson Chadwick, Bonita Lam and Matthew Schechter. Additional thanks to Shino Suzuki and Gijs Kuenen for insightful research and culturing support. This work was funded through a Grant-in-Aid for Young Scientists A and B from the Japan Society for Promotion of Science (JSPS) KAKENHI Grant Number 17H04969 and 26810085, respectively, and the Japan Agency for Medical Research and Development (17gm6010002h0002). US funding provided by the US Office of Global Naval Research (N62909-17-1-2038), and the Center for Dark Energy Biosphere Investigations (C-DEBI) (OCE0939564) and the NASA Astrobiology Institute - Life Underground (NAI-LU) (NNA13AA92A). Part of this work was conducted as part of a Japan Society for the Promotion of Sciences: Short-term postdoctoral fellowship for Annette Rowe (PE15019) at the University of Tokyo in the lab of Kazuhito Hashimoto.

1. Construction of a Two-electrode System for Environmental Incubation
<ol>
	<li>Preparation of the anode material and treatment of Carbon felt electrode (Figure 2).
	<ol>
		<li>Cut the carbon felt to equal dimensions depending on desired biomass enrichment. Soak each electrode in 90% ethanol for 30 min, then rinse at least 8 times with deionized water, sonicating for 1 min after each rinse.</li>
		<li>Wash the electrodes twice in 1 M HCl, stirring for a minimum of 12 h fo...

<ol>
	<li>Nealson, K. H., Saffarini, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Iron+and+manganese+in+anaerobic+respiration:+environmental+significance,+physiology,+and+regulation.">Iron and manganese in anaerobic respiration: environmental significance, physiology, and regulation.</a> Annual Reviews of Microbiology. 48, 311-343 (1994).</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=Bug+juice:+harvesting+electricity+with+microorganisms.">Bug juice: harvesting electricity with microorganisms.</a> Nature Reviews Microbiology. 4 (7), 497-508 (2006).</li><li>Rabaey, K., Rozendal, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microbial+electrosynthesis+-+revisiting+the+electrical+route+for+microbial+production.">Microbial electrosynthesis - revisiting the electrical route for microbial production.</a> Nature Reviews Microbiology. 8 (10), 706-716 (2010).</li><li>Lovley, D. R., Coates, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioremediation+of+metal+contamination.">Bioremediation of metal contamination.</a> Current Opinion in Biotechnology. 8 (3), 285-289 (1997).</li><li>Dinh, H. 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=Iron+corrosion+by+novel+anaerobic+microorganisms.">Iron corrosion by novel anaerobic microorganisms.</a> Nature. 427 (6977), 829-832 (2004).</li><li>Myers, C. R., Nealson, K. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bacterial+manganese+reduction+and+growth+with+manganese+oxide+as+the+sole+electron+acceptor.">Bacterial manganese reduction and growth with manganese oxide as the sole electron acceptor.</a> Science. 240 (4857), 1319-1321 (1988).</li><li>Lovley, D. R., Phillips, E. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+mode+of+microbial+energy+metabolism:+organic+carbon+oxidation+coupled+to+dissimilatory+reduction+of+iron+or+manganese.">Novel mode of microbial energy metabolism: organic carbon oxidation coupled to dissimilatory reduction of iron or manganese.</a> Applied and Environmental Microbiology. 54 (6), 1472-1480 (1988).</li><li>Arnold, R. G., DiChristina, T. J., Hoffmann, M. R. <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+Fe(III)+oxides+by+Pseudomonas+sp+200.">Reductive dissolution of Fe(III) oxides by Pseudomonas sp 200.</a> Biotechnology and Bioengineering. 32 (9), 1081-1096 (1988).</li><li>Rowe, A. R., 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=In+situ+electrochemical+enrichment+and+isolation+of+a+magnetite-reducing+bacterium+from+a+high+pH+serpentinizing+spring.">In situ electrochemical enrichment and isolation of a magnetite-reducing bacterium from a high pH serpentinizing spring.</a> Environmentakl Microbiology. 19 (6), 2272-2285 (2017).</li><li>Suzuki, 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=Microbial+diversity+in+The+Cedars,+an+ultrabasic,+ultrareducing,+and+low+salinity+serpentinizing+ecosystem.">Microbial diversity in The Cedars, an ultrabasic, ultrareducing, and low salinity serpentinizing ecosystem.</a> Proceedings of the National Academy of Science U S A. 110 (38), 15336-15341 (2013).</li><li>Suzuki, 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=Physiological+and+genomic+features+of+highly+alkaliphilic+hydrogen-utilizing+Betaproteobacteria+from+a+continental+serpentinizing+site.">Physiological and genomic features of highly alkaliphilic hydrogen-utilizing Betaproteobacteria from a continental serpentinizing site.</a> Nature Communications. 5, 3900 (2014).</li><li>McCollom, T. 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=Temperature+trends+for+reaction+rates,+hydrogen+generation,+and+partitioning+of+iron+during+experimental+serpentinization+of+olivine.">Temperature trends for reaction rates, hydrogen generation, and partitioning of iron during experimental serpentinization of olivine.</a> Geochimica et Cosmochimica Acta. 181, 175-200 (2016).</li><li>Morrill, P. L., 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=Geochemistry+and+geobiology+of+a+present-day+serpentinization+site+in+California:+The+Cedars.">Geochemistry and geobiology of a present-day serpentinization site in California: The Cedars.</a> Geochimica et Cosmochimica Acta. 109, 222-240 (2013).</li><li>Okamoto, A., Nakamura, R., Hashimoto, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In-vivo+identification+of+direct+electron+transfer+from+Shewanella+oneidensis+MR-1+to+electrodes+via+outer-membrane+OmcA-MtrCAB+protein+complexes.">In-vivo identification of direct electron transfer from Shewanella oneidensis MR-1 to electrodes via outer-membrane OmcA-MtrCAB protein complexes.</a> Electrochimica Acta. 56 (16), 5526-5531 (2011).</li><li>Okamoto, A., Hashimoto, K., Nakamura, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Spectroelectrochemical Investigation on Biological Electron Transfer Associated with Anode Performance in Microbial Fuel Cells. , 207-222 (2012).</li><li>Deng, X., Nakamura, R., Hashimoto, K., Okamoto, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electron+from+an+Extracellular+Electrode+by+Desulfovibrio+ferrophilus+Strain+IS5+Without+Using+Hydrogen+as+an+Electron+Carrier.">Electron from an Extracellular Electrode by Desulfovibrio ferrophilus Strain IS5 Without Using Hydrogen as an Electron Carrier.</a> Electrochemistry. 83 (7), 529-531 (2015).</li></ol>

In the described study, we show the enrichment of a microbial consortium, linked with in situ current production. The observed patterns in current support microbial activity in this system over short and long time scales. The critical step for constructing a functional two-electrode (fuel cell type) system is identifying and utilizing a location with a stable water-level and oxygen concentration in the environment. The cathode is exposed to oxygen at the air water interface, while the anode is kept under anaerob...

Several mineral-reducing microbes have been shown to utilize solid-phase minerals as terminal electron acceptors, by extracellular electron transport (EET) processes that conduct electrons to the exterior of the cell via redox enzymes1. EET is critical, not only for microbe-mineral processes but also applied energy and environmental technologies, such as microbial fuel cells2, electrode synthesis3, and bioremediation4. New EET-capable bacteria are highly sought after, and have been extensively studied from a fundamental or applied perspective5. However, we only have limited insight into the&#160;ecological or biogeochemical significance of these bacteria. The majority of EET-capable microbes have been isolated following enrichment from aqua, sediment, or anaerobic digesters using solid electron acceptors such as MnO2, Fe2O3 or poised electrodes in laboratory6,7,8. However, these methods often produce similar consortia and potentially miss more sensitive taxa that may dominate low energy or low biomass systems, biasing the ability of these microbes to adapt to the lab or axenic culture environment9. Usually for low biomass environments, large quantities of water from a site are filtered to concentrate bacterial cells. However, EET-capable bacteria often exhibit anaerobic metabolisms and therefore oxygen exposure may further inhibit or prevent their cultivation. Alternative on-site methodologies to concentrate cells without exposing them to oxygen could facilitate the isolation of EET-capable bacteria. Here, we report setup details for an on-site electrochemical technique to enrich EET-capable microbe over a long period of time without the need for an external power source.
Using our electrocultivation experiments from a highly alkaline spring in Northern California, the Cedars10, we describe this on-site electrochemical technique. The geochemistry of the springs at The Cedars are impacted by serpentinization in the subsurface. The springs are highly reductive, with oxygen concentrations below the limit of detection under the air water interface highlighting the potential for microbial energy production via EET in this functionally anoxic environment11. However, there is no evidence to support EET-capable microbes from the Cedars (in either 16S rRNA or Metagenomic analysis). Even though this environment has been characterized as electron acceptor limited, the potential for using insoluble minerals as terminal electron acceptors, including minerals such as the iron baring minerals that result from serpentinization (i.e., magnetite), has not been extensively investigated12. We, therefore, deployed our electrochemical system at Campsite Spring, a high pH spring at the Cedars, to enrich for EET-capable microbes (Figure1)13.

<table border="0" cellpadding="0" cellspacing="0" width="688">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:215px;">Carbon felt sheet</td>
			<td style="width:147px;">n/a</td>
			<td style="width:115px;">n/a</td>
			<td style="width:212px;">Used for anode and cathode</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Titanium wire</td>
			<td>The Nilaco Cooporation</td>
			<td>TI-451485</td>
			<td>Used to construct fuel cell system</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Graphite epoxy</td>
			<td style="width:147px;">Electrolytica lnc.</td>
			<td style="width:115px;">n/a</td>
			<td style="width:212px;">Used to connect the 
			electrodes and Ti wire</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:215px;">Drying oven</td>
			<td style="width:147px;">Yamato</td>
			<td style="width:115px;">DY300</td>
			<td style="width:212px;">bake the electrode to 
			solidify conductive graphite epoxy</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Digital multi meter</td>
			<td style="width:147px;">Fluke</td>
			<td style="width:115px;">616-1454</td>
			<td style="width:212px;">to check the ohmic value 
			of resistance</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:215px;">Dissolved oxygen probe</td>
			<td style="width:147px;">Sper Science</td>
			<td style="width:115px;">#&#160; 850045</td>
			<td style="width:212px;">to check the oxygen 
			concentration in the environments</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Resistor</td>
			<td style="width:147px;">Sodial</td>
			<td style="width:115px;"></td>
			<td style="width:212px;">Used to construct fuel cell 
			system</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Conducting wire</td>
			<td style="width:147px;">Pico</td>
			<td style="width:115px;">81141s</td>
			<td style="width:212px;">Used to construct fuel cell 
			system</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:215px;">Voltmeter and Data logger</td>
			<td style="width:147px;">T&#38;D corporation</td>
			<td style="width:115px;">VR-71</td>
			<td>Used for data recording</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:215px;">Hydrogen Hexachloroplatinate(IV) Hexahydrate</td>
			<td style="width:147px;">wako</td>
			<td style="width:115px;">18497-13-7</td>
			<td>Used for electropolation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:215px;">Citric acid</td>
			<td style="width:147px;">Wako</td>
			<td style="width:115px;">038-06925</td>
			<td>Used for electropolation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:215px;">Sulfuric acid</td>
			<td style="width:147px;">Wako</td>
			<td style="width:115px;">192-04696</td>
			<td>Used for electropolation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:215px;">HCl</td>
			<td style="width:147px;">Wako</td>
			<td style="width:115px;">083-01095</td>
			<td>Used for electrode washing</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Glass cylinder</td>
			<td style="width:147px;">N/A</td>
			<td style="width:115px;">N/A</td>
			<td style="width:212px;">Custom-made, used as the electrochemical reactor</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:215px;">PTFE cover and base</td>
			<td style="width:147px;">N/A</td>
			<td style="width:115px;">N/A</td>
			<td style="width:212px;">Custom-made, used as a cover and a foundation of the electrochemical reactor</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:215px;">Buthyl rubber</td>
			<td style="width:147px;">N/A</td>
			<td style="width:115px;">N/A</td>
			<td style="width:212px;">Custom-made, inserted between each component of electrochemical reactor</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Septa</td>
			<td style="width:147px;">GL Science</td>
			<td style="width:115px;">3007-16101</td>
			<td style="width:212px;">Used as an injection port of electrochemical reactor</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Indium tin-doped oxide (ITO) electrode</td>
			<td style="width:147px;">GEOMATEC</td>
			<td style="width:115px;">No.0001</td>
			<td style="width:212px;">Used as a working electrode, 5&#937;/sq</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Ag/AgCl KCl saturated electrode</td>
			<td style="width:147px;">HOKUTO DENKO</td>
			<td style="width:115px;">HX-R5</td>
			<td style="width:212px;">Used as a reference electrode, &#934;0.30mm</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Platinum wire</td>
			<td style="width:147px;">The Nilaco Cooporation</td>
			<td style="width:115px;">PT-351325</td>
			<td style="width:212px;">Used as a counter electrode</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">NaHCO3</td>
			<td style="width:147px;">Wako</td>
			<td style="width:115px;">191-01305</td>
			<td style="width:212px;">Used for The Cedars Media (CMS)</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:215px;">CaCO3</td>
			<td style="width:147px;">Wako</td>
			<td style="width:115px;">030-00385</td>
			<td style="width:212px;">Used for CMS</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:215px;">NH4Cl</td>
			<td style="width:147px;">Wako</td>
			<td style="width:115px;">011-03015</td>
			<td style="width:212px;">Used for CMS</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:215px;">MgCl2&#160;&#8226; 6H2O</td>
			<td style="width:147px;">Wako</td>
			<td style="width:115px;">135-00165</td>
			<td style="width:212px;">Used for CMS</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:215px;">NaOH&#160;</td>
			<td style="width:147px;">Wako</td>
			<td style="width:115px;">198-13765</td>
			<td style="width:212px;">Used for CMS</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:215px;">Na2SO4</td>
			<td style="width:147px;">Wako</td>
			<td style="width:115px;">194-03355</td>
			<td style="width:212px;">Used for CMS</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:215px;">K2HPO4</td>
			<td style="width:147px;">Wako</td>
			<td style="width:115px;">164-04295</td>
			<td style="width:212px;">Used for CMS</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:215px;">CABS</td>
			<td style="width:147px;">SANTA CRUZ</td>
			<td style="width:115px;">SC-285279</td>
			<td style="width:212px;">Used for CMS</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Incubator</td>
			<td style="width:147px;">TOKYO RIKAKIKAI CO. LTD.</td>
			<td style="width:115px;">LTI-601SD</td>
			<td style="width:212px;">Used for precultivation</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:215px;">Autoclave machine</td>
			<td style="width:147px;">TOMY SEIKO CO. LTD.</td>
			<td style="width:115px;">LSX-500</td>
			<td style="width:212px;">Used for sterilization of the electrochemical reactor and the medium</td>
		</tr>
		<tr height="105">
			<td height="105" style="height:105px;width:215px;">Clean bench</td>
			<td style="width:147px;">SANYO</td>
			<td style="width:115px;">MCV-91BNF</td>
			<td style="width:212px;">Used to prevent the contamination of the electrochemical reactor and the medium with other microbes</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Centrifuge separator</td>
			<td style="width:147px;">Eppendorf</td>
			<td style="width:115px;">5430R</td>
			<td style="width:212px;">Rotational speed upto 6000&#215;g is required</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:215px;">Nitrogen gas generator</td>
			<td style="width:147px;">Puequ CO. LTD.</td>
			<td style="width:115px;">PNTN-2</td>
			<td style="width:212px;">Nitrogen gas cylinder can also be used instead of gas generator</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">UV-vis spectrometer</td>
			<td style="width:147px;">SHIMADZU</td>
			<td style="width:115px;">UV-1800</td>
			<td style="width:212px;">Used for optimization of cell density</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:215px;">Potentiostat</td>
			<td style="width:147px;">BioLogic</td>
			<td style="width:115px;">VMP3</td>
			<td style="width:212px;">Used for biofilm formation and kinetic isotope effect experiments</td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:215px;">Thermal water circulator</td>
			<td style="width:147px;">AS ONE</td>
			<td style="width:115px;">TR-1A</td>
			<td style="width:212px;">Used for maintanance of temperature of electrochemcial reactor</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Faraday cage</td>
			<td style="width:147px;">HOKUTO DENKO</td>
			<td style="width:115px;">HS-201S</td>
			<td style="width:212px;">Used for electrochemical experiments</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:215px;">Anaerobic Chamber</td>
			<td style="width:147px;">COY</td>
			<td style="width:115px;">TypeB (Vinyl)</td>
			<td style="width:212px;">TO conduct experiments 
			under anaerobic condition</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Ultraclean DNA Extraction kit</td>
			<td style="width:147px;">MoBio</td>
			<td style="width:115px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

self-standing electrochemical set-up to enrich anode-respiring bacteria on-site

Anaerobic respiration coupled with electron transport to insoluble minerals (referred to as extracellular electron transport [EET]) is thought to be critical for microbial energy production and persistence in many subsurface environments, especially those lacking soluble terminal electron acceptors. While EET-capable microbes have been successfully isolated from various environments, the diversity of bacteria capable of EET is still poorly understood, especially in difficult-to-sample, low energy or extreme environments, such as many subsurface ecosystems. Here, we describe an on-site electrochemical system to enrich EET-capable bacteria using an anode as a respiratory terminal electron acceptor. This anode is connected&#160;to a cathode capable of&#160;catalyzing abiotic oxygen reduction. Comparing this approach with electrocultivation methods that use a potentiostat for poising the electrode potential, the two-electrode system does not require an external power source. We present an example of our on-site enrichment utilized in an alkaline pond at the Cedars, a terrestrial serpentinization site in Northern California. Prior attempts to cultivate mineral reducing bacteria were unsuccessful, which is likely due to the low-biomass nature of this site and/or the low relative abundance of metal reducing microbes. Prior to implementing our two-electrode enrichment, we measured the vertical profile of dissolved oxygen concentration. This allowed us to place the carbon felt anode and platinum-electroplated carbon felt cathode at depths that would support anaerobic and aerobic processes, respectively. Following on-site incubation, we further enriched the anodic electrode in the laboratory and confirmed&#160;a distinct microbial community compared to the surface-attached or biofilm communities normally observed at the Cedars. This enrichment subsequently led to the isolation of the first electrogenic microbe from the Cedars. This method of on-site microbial enrichment has the potential to greatly enhance the isolation of EET-capable bacteria from low biomass or difficult to sample habitats.

Current production was successfully measured for approximately 3 months using a voltage data logger as shown in Figure 3. This time was chosen as it was the longest stable incubation period for the spring, due to strong fall rains affecting the spring. A shorter period could be sufficient, though a longer period could provide stronger enrichment of biomass. We confirmed the connection of the two-electrode system after electrochemical incubation and observed n...

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Self-standing Electrochemical Set-up to Enrich Anode-respiring Bacteria On-site

On-site microbial enrichment or in situ cultivation techniques can facilitate the isolation of difficult-to-culture microbial taxa, especially from low-biomass or geochemically extreme environments. Here, we describe an electrochemical set-up without using an external power source to enrich microbial strains that are capable of extracellular electron transport (EET).

Anaerobic respiration coupled with electron transport to insoluble minerals (referred to as extracellular electron transport [EET]) is thought to be ...

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