Novel method for screening microbes for application in microbial fuel cell

doi:10.1016/j.biortech.2014.12.004

Bioresource Technology

Volume 179, March 2015, Pages 123-127

https://doi.org/10.1016/j.biortech.2014.12.004 Get rights and content

Highlights

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The correlation between growth and iron-reducing capacity was observed.
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Regression model (changes of bio-current) was obtained.
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Novel, fast method was developed for screening microbe for application in MFC.

Abstract

The ability to produce and to transport exo-electrons by microbes either to external acceptors or to electrodes are reported in our study. All investigated microorganisms (exception of Lactobacillus plantarum) exhibited strong iron-reducing capabilities in the absence of mediator meaning production and secretion of exo-electrons to the growth medium. L. plantarum, Saccharomyces cerevisiae and Escherichia coli need an electron shuttle molecule to reduce Fe³⁺ ion. Significant correlation was observed between growth and iron-reducing capacity, as well as between initial cell counts and iron-reducing capacity. Changes of bio-current generated in MFC and iron-reduction were experimentally monitored, and a mathematical model was established by regression analysis. Based on these results, a novel and rapid screening method was developed for the selection of microorganisms for potential application in MFC. The method is based on the measurement of absorbance of bacterial and yeast cultures at 460 nm, providing a robust and high sample throughput approach.

Introduction

Microbial fuel cell (MFC) is a promising technology for the production of electricity from numerous raw materials such as natural organic matter, complex organic waste or renewable biomass, and can be advantageously combined with applications in wastewater treatment (Oliveira et al., 2013, Wen et al., 2010). Organic compounds commonly used as electron donors are easily biodegradable, including simple carbohydrates (e.g., glucose and saccharose), starch, low-molecular weight organic acids (acetate, oxalate, fumarate), xylose and amino acids (Hassan et al., 2012). The main advantage of this technology is the application of microorganism(s) as a catalyst to convert organic materials directly into electricity, thus it produces bioenergy even from wastes. It is well known that electron transport chains play an important role in adenosine triphosphate (ATP) synthesis where a series of compounds are involved transferring electrons from an electron donor to an electron acceptor via redox reactions. Organic matters generally act as electron donors, whereas many electron acceptor compounds (both organic and inorganic) are known. If oxygen is available (i.e., in aerobic respiration), it is used as the terminal electron acceptor, because oxygen provides the greatest Gibbs free energy difference. Moreover, it produces the highest level of energy and microbes usually maximize their energy gain by selecting electron acceptor with the highest potential (Rabaey et al., 2007). In anaerobic environments, different electron acceptors are used, including nitrate, nitrite, ferric iron, sulfate, carbon dioxide, and small organic molecules such as fumarate. Many microbes, for examples iron-reducing bacteria (Geobacter sulfurreducens, Geobacter metallireducens, Geobacter toluenoxydans (Caccavo et al., 1994), Rhodoferax ferrireducens (Chaudhuri and Lovley, 2003), Shewanella putrefaciens (Kim et al., 2002), Shewanella japonica, Shewanella algae (Ivanova et al., 2001), are reported to use soluble or insoluble metals or metal-oxides as the electron acceptor, and they are able to transfer electrons out of the cell membrane (Luu and Ramsay, 2003). Only those microorganisms can be potentially used in MFC that are able to transfer extracellular electrons to electrodes. Electron transfer can occur either through membrane-associated components (Logan and Regan, 2006, Aelterman et al., 2006), or soluble electron shuttles generated by specific bacteria (Logan and Regan, 2006, Bond and Lovley, 2003) or highly conductive nanowires (Reguera et al., 2005).

The electron transfer mechanism of the iron-reducing bacteria is represented mostly by Geobacteria spp. It is usually direct and independent from the type of electron acceptors (Fe(III)-oxide, Fe(III)-citrate, etc.) or different kinds of electrodes (Stams et al., 2006, Feng et al., 2013). For most microbes, the transfer of electrons to electrodes is inefficient because of electrically non-conductive cell walls and impedance by the peptide chain adjoining the active redox center of proteins (Kim et al., 2002). Some special bacterial strains can produce electron mediators and some of them are capable to transfer electrons directly (Chaudhuri and Lovley, 2003, Park and Zeikus, 2000, Pizzariello et al., 2002), therefore such inefficiency can be avoided. Also, another method may be the addition of artificial electron shuttle molecules (mediator). Yeasts generally need a mediator such as methylene blue, methyl orange, resazurin, etc., for electron transport (Watanabe et al., 2009, Rahimnejad et al., 2011); accordingly, the application of yeasts in MFC is still very limited.

The operation efficiency of the MFC can be influenced by numerous factors such as microorganisms, design of MFC, type of proton exchange membrane (PEM), type of electrodes, etc. (Feng et al., 2013). Among these, the microbial factor is crucial, so the screening of the potential species is essential. Several studies demonstrated different methods to screen the electrochemically active bacteria (e.g., U-tube MFCs (Yu et al., 2012), micro-fabricated MFC arrays (Hou et al., 2009) or Tungsten-oxide nanocluster probe (Yuan et al., 2013). However, these methods take relatively long time (5–6 days) to provide quantitative information about the extracellular-electron production of the microbes. Another limiting factor of these methods is the requirement of expensive equipment and materials; therefore there is an increasing demand for novel and rapid screening methods both in research and in development of MFCs. The main aim of this study was to develop a simple, affordable and high sample throughput method for the screening of microorganism strains for MFC application.

Section snippets

Microorganisms

G. sulfurreducens DSMZ 12127, G. toluenoxydans DSMZ 19350, Shewanella algae DSMZ 9167, Shewanella woodyi DSMZ 12036 and Shewanella xiamenensis DSMZ 22215 were purchased from Deutsche Sammlung von Mikroorganizmen und Zellkulturen (DSMZ), Braunschweig, Germany. Escherichia coli ATCC 8739 and Lactobacillus plantarum 2142 strains were obtained from University of Perugia, Italy. The yeast strain was Saccharomyces cerevisiae WS-120 (Hefebank Weihenstephan, TUM, Freising, Germany).

Inoculum preparation

Geobacter strains

Results and discussion

The iron(III)-reduction capability of different microorganisms were tested both in the presence and in the absence of methylene blue as the mediator. The results are shown in Table 1.

With the exception of L. plantarum all investigated microorganisms exhibited strong iron(III)-reducing capabilities in the absence of mediator compounds, which indicates the production and secretion of exo-electrons into the growth medium. Generally, Geobacter species and Shewanella species are iron-reducing

Conclusions

This study has made significant step toward screening microorganisms for application in MFC. The capability of production of exo-electrons and their transportation either to electron acceptors or to electrodes can be detected by a very simple and rapid photometric method. The current density produced by certain microorganism (if the initial cell count is higher than 10⁶ CFU/mL) can be estimated by the model of current density = 46.77 ΔA/460 nm/ + 4.17. No doubt that this method is robust, takes a

Acknowledgements

This work is supported by National Development Agency (Project no. TÁMOP-4.2.1./B-09/1-KMR-2010-0005, TÁMOP-4.2.2/B-10/1-2010-0023 and TECH_09-A3-2009-0194), as well as Bolyai Research Grant from Hungarian Academic of Sciences. Authors would like to thank Dr. Mihály Dernovics Associate Professor at Department of Applied Chemistry, Corvinus University of Budapest for reading manuscript.

References (31)

C.H. Feng et al.
Bio-current as an indicator for biogenic Fe(II) generation driven by dissimilatory iron reducing bacteria
Biosens. Bioelectron.
(2013)
S.H.A. Hassan et al.
Power generation from cellulose using mixed and pure cultures of cellulose-degrading bacteria in a microbial fuel cell
Enzyme Microb. Technol.
(2012)
E. Herrero-Hernandez et al.
Electricity generation from wastewaters with starch as carbon source using a mediatorless microbial fuel cell
Biosens. Bioelectron.
(2013)
H.J. Kim et al.
A mediator-less microbial fuel cell using a metal reducing bacterium, Shewanella putrefaciens
Enzyme Microb. Technol.
(2002)
B.E. Logan et al.
Electricity-producing bacterial communities in microbial fuel cells
Trends Microbiol.
(2006)
V.B. Oliveira et al.
Overview on the developments of microbial fuel cells
Biochem. Eng. J.
(2013)
D. Prasad et al.
Direct electron transfer with yeast cells and construction of a mediatorless microbial fuel cell
Biosens. Bioelectron.
(2007)
S.V. Raghavulu et al.
Saccharomyces cerevisiae as anodic biocatalyst for power generation in biofuel cell: influence of redox condition and substrate load
Bioresour. Technol.
(2011)
M. Rahimnejad et al.
Methylene blue as electron promoters in microbial fuel cell
Int. J. Hydrogen Energy
(2011)
A. Pizzariello et al.
A glucose/hydrogen peroxide biofuel cell that uses oxidase and peroxidase as catalysts by composite bulk-modified bioelectrodes based on a solid binding matrix
Bioelectrochemistry
(2002)

F.R. Van der Zee et al.

Impact and application of electron shuttles on the redox (bio)transformation of contaminants: a review

Biotechnol. Adv.

(2009)

K. Watanabe et al.

Electron shuttles in biotechnology

Curr. Opin. Biotechnol.

(2009)

L.L. Wei et al.

Effects of cathodic electron acceptors and potassium ferricyanide concentrations on the performance of microbial fuel cell

Int. J. Hydrogen Energy

(2012)

Q. Wen et al.

Production of electricity from the treatment of continuous brewery wastewater using a microbial fuel cell

Fuel

(2010)

P. Aelterman et al.

Continuous electricity generation at high voltages and currents using stacked microbial fuel cells

Environ. Sci. Technol.

(2006)

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Novel method for screening microbes for application in microbial fuel cell

Highlights

Abstract

Introduction

Section snippets

Microorganisms

Inoculum preparation

Results and discussion

Conclusions

Acknowledgements

Biosens. Bioelectron.

Enzyme Microb. Technol.

Biosens. Bioelectron.

Enzyme Microb. Technol.

Trends Microbiol.

Biochem. Eng. J.

Biosens. Bioelectron.

Bioresour. Technol.

Int. J. Hydrogen Energy

Bioelectrochemistry

Biotechnol. Adv.

Curr. Opin. Biotechnol.

Int. J. Hydrogen Energy

Fuel

Continuous electricity generation at high voltages and currents using stacked microbial fuel cells

Environ. Sci. Technol.