scholarly journals Structures, Compositions, and Activities of Live Shewanella Biofilms Formed on Graphite Electrodes in Electrochemical Flow Cells

2017 ◽  
Vol 83 (17) ◽  
Author(s):  
Miho Kitayama ◽  
Ryota Koga ◽  
Takuya Kasai ◽  
Atsushi Kouzuma ◽  
Kazuya Watanabe
Keyword(s):  
Wastewater Treatment ◽  
Water Flow ◽  
Biofilm Formation ◽  
Shewanella Oneidensis ◽  
Flow Conditions ◽  
Current Generation ◽  
Content Type ◽  
Flow Cells ◽  
Electrode Potentials ◽  
Over Time

ABSTRACT An electrochemical flow cell equipped with a graphite working electrode (WE) at the bottom was inoculated with Shewanella oneidensis MR-1 expressing an anaerobic fluorescent protein, and biofilm formation on the WE was observed over time during current generation at WE potentials of +0.4 and 0 V (versus standard hydrogen electrodes), under electrolyte-flow conditions. Electrochemical analyses suggested the presence of unique electron-transfer mechanisms in the +0.4-V biofilm. Microscopic analyses revealed that, in contrast to aerobic biofilms, current-generating biofilm (at +0.4 V) was thin and flat (∼10 μm in thickness), and cells were evenly and densely distributed in the biofilm. In contrast, cells were unevenly distributed in biofilm formed at 0 V. In situ fluorescence staining and biofilm recovery experiments showed that the amounts of extracellular polysaccharides (EPSs) in the +0.4-V biofilm were much smaller than those in the aerobic and 0-V biofilms, suggesting that Shewanella cells suppress the production of EPSs at +0.4 V under flow conditions. We suggest that Shewanella cells perceive electrode potentials and modulate the structure and composition of biofilms to efficiently transfer electrons to electrodes. IMPORTANCE A promising application of microbial fuel cells (MFCs) is to save energy in wastewater treatment. Since current is generated in these MFCs by biofilm microbes under horizontal flows of wastewater, it is important to understand the mechanisms for biofilm formation and current generation under water-flow conditions. Although massive work has been done to analyze the molecular mechanisms for current generation by model exoelectrogenic bacteria, such as Shewanella oneidensis, limited information is available regarding the formation of current-generating biofilms over time under water-flow conditions. The present study developed electrochemical flow cells and used them to examine the electrochemical and structural features of current-generating biofilms under water-flow conditions. We show unique features of mature biofilms actively generating current, creating opportunities to search for as-yet-undiscovered current-generating mechanisms in Shewanella biofilms. Furthermore, information provided in the present study is useful for researchers attempting to develop anode architectures suitable for wastewater treatment MFCs.

scholarly journals Identification of a Diguanylate Cyclase That Facilitates Biofilm Formation on Electrodes by Shewanella oneidensis MR-1

2021 ◽  
Vol 87 (9) ◽  
Author(s):  
Akiho Matsumoto ◽  
Ryota Koga ◽  
Robert A. Kanaly ◽  
Atsushi Kouzuma ◽  
Kazuya Watanabe
Keyword(s):  
Microbial Fuel Cells ◽  
Biofilm Formation ◽  
Shewanella Oneidensis ◽  
Electric Energy ◽  
Diguanylate Cyclase ◽  
Flow Conditions ◽  
Content Type ◽  
Electrochemically Active ◽  
Medium Flow

ABSTRACT In many bacteria, cyclic diguanosine monophosphate (c-di-GMP), synthesized by diguanylate cyclase (DGC), serves as a second messenger involved in the regulation of biofilm formation. Although studies have suggested that c-di-GMP also regulates the formation of electrochemically active biofilms (EABFs) by Shewanella oneidensis MR-1, DGCs involved in this process remained to be identified. Here, we report that the SO_1646 gene, hereafter named dgcS, is upregulated under medium flow conditions in electrochemical flow cells (EFCs), and its product (DgcS) functions as a major DGC in MR-1. In vitro assays demonstrated that purified DgcS catalyzed the synthesis of c-di-GMP from GTP. Comparisons of intracellular c-di-GMP levels in the wild-type strain and a dgcS deletion mutant (ΔdgcS mutant) showed that production of c-di-GMP was markedly reduced in the ΔdgcS mutant when cells were grown in batch cultures and on electrodes in EFCs. Cultivation of the ΔdgcS mutant in EFCs also revealed that the loss of DgcS resulted in impaired biofilm formation and decreased current generation. These findings demonstrate that MR-1 uses DgcS to synthesize c-di-GMP under medium flow conditions, thereby activating biofilm formation on electrodes. IMPORTANCE Bioelectrochemical systems (BESs) have attracted wide attention owing to their utility in sustainable biotechnology processes, such as microbial fuel cells and electrofermentation systems. In BESs, electrochemically active bacteria (EAB) form biofilms on electrode surfaces, thereby serving as effective catalysts for the interconversion between chemical and electric energy. It is therefore important to understand mechanisms for the formation of biofilm by EAB grown on electrodes. Here, we show that a model EAB, S. oneidensis MR-1, expresses DgcS as a major DGC, thereby activating the formation of biofilms on electrodes via c-di-GMP-dependent signal transduction cascades. The findings presented herein provide the molecular basis for improving electrochemical interactions between EAB and electrodes in BESs. The results also offer molecular insights into how Shewanella regulates biofilm formation on solid surfaces in the natural environment.

scholarly journals Quorum Sensing Influences Burkholderia thailandensis Biofilm Development and Matrix Production

2016 ◽  
Vol 198 (19) ◽  
pp. 2643-2650 ◽  
Author(s):  
Boo Shan Tseng ◽  
Charlotte D. Majerczyk ◽  
Daniel Passos da Silva ◽  
Josephine R. Chandler ◽  
E. Peter Greenberg ◽  
...  
Keyword(s):  
Quorum Sensing ◽  
Biofilm Formation ◽  
Bacterial Species ◽  
Matrix Material ◽  
Close Relative ◽  
Wild Type ◽  
Flow Conditions ◽  
Burkholderia Thailandensis ◽  
Content Type ◽  
Biofilm Development

ABSTRACTMembers of the genusBurkholderiaare known to be adept at biofilm formation, which presumably assists in the survival of these organisms in the environment and the host. Biofilm formation has been linked to quorum sensing (QS) in several bacterial species. In this study, we characterizedBurkholderia thailandensisbiofilm development under flow conditions and sought to determine whether QS contributes to this process.B. thailandensisbiofilm formation exhibited an unusual pattern: the cells formed small aggregates and then proceeded to produce mature biofilms characterized by “dome” structures filled with biofilm matrix material. We showed that this process was dependent on QS.B. thailandensishas three acyl-homoserine lactone (AHL) QS systems (QS-1, QS-2, and QS-3). An AHL-negative strain produced biofilms consisting of cell aggregates but lacking the matrix-filled dome structures. This phenotype was rescued via exogenous addition of the three AHL signals. Of the threeB. thailandensisQS systems, we show that QS-1 is required for proper biofilm development, since abtaR1mutant, which is defective in QS-1 regulation, forms biofilms without these dome structures. Furthermore, our data show that the wild-type biofilm biomass, as well as the material inside the domes, stains with a fucose-binding lectin. ThebtaR1mutant biofilms, however, are negative for fucose staining. This suggests that the QS-1 system regulates the production of a fucose-containing exopolysaccharide in wild-type biofilms. Finally, we present data showing that QS ability during biofilm development produces a biofilm that is resistant to dispersion under stress conditions.IMPORTANCEThe saprophyteBurkholderia thailandensisis a close relative of the pathogenic bacteriumBurkholderia pseudomallei, the causative agent of melioidosis, which is contracted from its environmental reservoir. Since most bacteria in the environment reside in biofilms,B. thailandensisis an ideal model organism for investigating questions inBurkholderiaphysiology. In this study, we characterizedB. thailandensisbiofilm development and sought to determine if quorum sensing (QS) contributes to this process. Our work shows thatB. thailandensisproduces biofilms with unusual dome structures under flow conditions. Our findings suggest that these dome structures are filled with a QS-regulated, fucose-containing exopolysaccharide that may be involved in the resilience ofB. thailandensisbiofilms against changes in the nutritional environment.

Increased biofilm in Shewanella oneidensis MR-1 leads to higher current generation in METs

2020 ◽  
Author(s):  
Ana V. Silva ◽  
Miriam Edel ◽  
Johannes Gescher ◽  
Catarina M. Paquete
Keyword(s):  
Biofilm Formation ◽  
Large Scale ◽  
Shewanella Oneidensis ◽  
Extracellular Electron Transfer ◽  
Central Process ◽  
Current Generation ◽  
Promising Alternative ◽  
Over Expression ◽  
Current Production ◽  
Low Levels

<p>Biofilm formation is a central process in the function of Microbial Electrochemical Technologies (METs). These technologies have emerged in recent years as a promising alternative green source of energy, in which microbes consume organic matter to produce energy or valuable by-products. It is the ability of performing extracellular electron transfer that allows these microbes, called electroactive organisms, to exchange electrons with an electrode in these systems. The low levels of current achieved have been the set-back for the large-scale application of METs. <em>Shewanella oneidensis</em> MR-1 is one of the most studied electroactive organisms, and it has been demonstrated that its increased biofilm formation can lead to higher current generation. The <em>bolA</em> gene has been identified as a central player in biofilm formation in different organisms, with its overexpression leading to increased biofilm production. In this work, we explored the effect of this gene in biofilm formation and current production by <em>S. oneidensis</em> MR-1. Our results demonstrate that this gene is involved in the biofilm formation by this organism, with its over expression leading to an increased biofilm formation. We could also show that this increase in biofilm formation lead to a consequent higher current generation. This information is a relevant step for the optimization of electroactive organisms towards their practical application in METs.</p>

scholarly journals Biofilm Formation by the Fish Pathogen Flavobacterium columnare: Development and Parameters Affecting Surface Attachment

2013 ◽  
Vol 79 (18) ◽  
pp. 5633-5642 ◽  
Author(s):  
Wenlong Cai ◽  
Leonardo De La Fuente ◽  
Covadonga R. Arias
Keyword(s):  
Biofilm Formation ◽  
Laser Scanning ◽  
Time Course ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Freshwater Species ◽  
Flavobacterium Columnare ◽  
Fish Pathogen ◽  
Flow Conditions ◽  
Content Type

ABSTRACTFlavobacterium columnareis a bacterial fish pathogen that affects many freshwater species worldwide. The natural reservoir of this pathogen is unknown, but its resilience in closed aquaculture systems posits biofilm as the source of contagion for farmed fish. The objectives of this study were (i) to characterize the dynamics of biofilm formation and morphology under static and flow conditions and (ii) to evaluate the effects of temperature, pH, salinity, hardness, and carbohydrates on biofilm formation. NineteenF. columnarestrains, including representatives of all of the defined genetic groups (genomovars), were compared in this study. The structure of biofilm was characterized by light microscopy, confocal laser scanning microscopy, and scanning electron microscopy.F. columnarewas able to attach to and colonize inert surfaces by producing biofilm. Surface colonization started within 6 h postinoculation, and microcolonies were observed within 24 h. Extracellular polysaccharide substances and water channels were observed in mature biofilms (24 to 48 h). A similar time course was observed whenF. columnareformed biofilm in microfluidic chambers under flow conditions. The virulence potential of biofilm was confirmed by cutaneous inoculation of channel catfish fingerlings with mature biofilm. Several physicochemical parameters modulate attachment to surfaces, with the largest influence being exerted by hardness, salinity, and the presence of mannose. Maintenance of hardness and salinity values within certain ranges could prevent biofilm formation byF. columnarein aquaculture systems.

scholarly journals σBInhibits Poly-N-Acetylglucosamine Exopolysaccharide Synthesis and Biofilm Formation inStaphylococcus aureus

2019 ◽  
Vol 201 (11) ◽  
Author(s):  
Jaione Valle ◽  
Maite Echeverz ◽  
Iñigo Lasa
Keyword(s):  
Staphylococcus Aureus ◽  
Biofilm Formation ◽  
Flow Conditions ◽  
Content Type ◽  
Environmental Signals ◽  
General Stress Response ◽  
Biofilm Development ◽  
Type Strains ◽  
Complex Regulatory Network

ABSTRACTStaphylococcus aureusclinical strains are able to produce at least two distinct types of biofilm matrixes: biofilm matrixes made of the polysaccharide intercellular adhesin (PIA) or poly-N-acetylglucosamine (PNAG), whose synthesis is mediated by theicaADBClocus, and biofilm matrixes built of proteins (polysaccharide independent). σBis a conserved alternative sigma factor that regulates the expression of more than 100 genes in response to changes in environmental conditions. While numerous studies agree that σBis required for polysaccharide-independent biofilms, controversy persists over the role of σBin the regulation of PIA/PNAG-dependent biofilm development. Here, we show that genetically unrelatedS. aureusσB-deficient strains produced stronger biofilms under both static and flow conditions and accumulated higher levels of PIA/PNAG exopolysaccharide than their corresponding wild-type strains. The increased accumulation of PIA/PNAG in the σBmutants correlated with a greater accumulation of the IcaC protein showed that it was not due to adjustments inicaADBCoperon transcription and/oricaADBCmRNA stability. Overall, our results reveal that in the presence of active σB, the turnover of Ica proteins is accelerated, reducing the synthesis of PIA/PNAG exopolysaccharide and consequently the PIA/PNAG-dependent biofilm formation capacity.IMPORTANCEDue to its multifaceted lifestyle,Staphylococcus aureusneeds a complex regulatory network to connect environmental signals with cellular physiology. One particular transcription factor, named σB(SigB), is involved in the general stress response and the expression of virulence factors. For many years, great confusion has existed about the role of σBin the regulation of the biofilm lifestyle inS. aureus. Our study demonstrated that σBis not necessary for exopolysaccharide-dependent biofilms and, even more, thatS. aureusproduces stronger biofilms in the absence of σB. The increased accumulation of exopolysaccharide correlates with higher stability of the proteins responsible for its synthesis. The present findings reveal an additional regulatory layer to control biofilm exopolysaccharide synthesis under stress conditions.

scholarly journals Roles of Two Shewanella oneidensis MR-1 Extracellular Endonucleases

2011 ◽  
Vol 77 (15) ◽  
pp. 5342-5351 ◽  
Author(s):  
Julia Gödeke ◽  
Magnus Heun ◽  
Sebastian Bubendorfer ◽  
Kristina Paul ◽  
Kai M. Thormann
Keyword(s):  
Biofilm Formation ◽  
Cell Envelope ◽  
Shewanella Oneidensis ◽  
Thick Layer ◽  
Sole Source ◽  
Extracellular Dna ◽  
Matrix Component ◽  
Content Type ◽  
Static Conditions

ABSTRACTThe dissimilatory iron-reducing bacteriumShewanella oneidensisMR-1 is capable of using extracellular DNA (eDNA) as the sole source of carbon, phosphorus, and nitrogen. In addition, we recently demonstrated thatS. oneidensisMR-1 requires eDNA as a structural component during all stages of biofilm formation. In this study, we characterize the roles of twoShewanellaextracellular endonucleases, ExeS and ExeM. While ExeS is likely secreted into the medium, ExeM is predicted to remain associated with the cell envelope. BothexeMandexeSare highly expressed under phosphate-limited conditions. Mutants lackingexeSand/orexeMexhibit decreased eDNA degradation; however, the capability ofS. oneidensisMR-1 to use DNA as the sole source of phosphorus is only affected in mutants lackingexeM. Neither of the two endonucleases alleviates toxic effects of increased eDNA concentrations. The deletion ofexeMand/orexeSsignificantly affects biofilm formation ofS. oneidensisMR-1 under static conditions, and expression ofexeMandexeSdrastically increases during static biofilm formation. Under hydrodynamic conditions, a deletion ofexeMleads to altered biofilms that consist of densely packed structures which are covered by a thick layer of eDNA. Based on these results, we hypothesize that a major role of ExeS and, in particular, ExeM ofS. oneidensisMR-1, is to degrade eDNA as a matrix component during biofilm formation to improve nutrient supply and to enable detachment.

scholarly journals Disruption of Putrescine Biosynthesis in Shewanella oneidensis Enhances Biofilm Cohesiveness and Performance in Cr(VI) Immobilization

2013 ◽  
Vol 80 (4) ◽  
pp. 1498-1506 ◽  
Author(s):  
Yuanzhao Ding ◽  
Ni Peng ◽  
Yonghua Du ◽  
Lianghui Ji ◽  
Bin Cao
Keyword(s):  
Biofilm Formation ◽  
Shewanella Oneidensis ◽  
Cell Detachment ◽  
Library Screening ◽  
Wild Type ◽  
Content Type ◽  
Biosynthesis Gene ◽  
And Performance

ABSTRACTAlthough biofilm-based bioprocesses have been increasingly used in various applications, the long-term robust and efficient biofilm performance remains one of the main bottlenecks. In this study, we demonstrated that biofilm cohesiveness and performance ofShewanella oneidensiscan be enhanced through disrupting putrescine biosynthesis. Through random transposon mutagenesis library screening, one hyperadherent mutant strain, CP2-1-S1, exhibiting an enhanced capability in biofilm formation, was obtained. Comparative analysis of the performance of biofilms formed byS. oneidensisMR-1 wild type (WT) and CP2-1-S1 in removing dichromate (Cr2O72−), i.e., Cr(VI), from the aqueous phase showed that, compared with the WT biofilms, CP2-1-S1 biofilms displayed a substantially lower rate of cell detachment upon exposure to Cr(VI), suggesting a higher cohesiveness of the mutant biofilms. In addition, the amount of Cr(III) immobilized by CP2-1-S1 biofilms was much larger, indicating an enhanced performance in Cr(VI) bioremediation. We further showed thatspeF, a putrescine biosynthesis gene, was disrupted in CP2-1-S1 and that the biofilm phenotypes could be restored by both genetic and chemical complementations. Our results also demonstrated an important role of putrescine in mediating matrix disassembly inS. oneidensisbiofilms.

scholarly journals Functional Specificity of Extracellular Nucleases of Shewanella oneidensis MR-1

2012 ◽  
Vol 78 (12) ◽  
pp. 4400-4411 ◽  
Author(s):  
Magnus Heun ◽  
Lucas Binnenkade ◽  
Maximilian Kreienbaum ◽  
Kai M. Thormann
Keyword(s):  
Biofilm Formation ◽  
Bacterial Species ◽  
Shewanella Oneidensis ◽  
Large Cell ◽  
Sole Source ◽  
Extracellular Dna ◽  
Divalent Metal Ions ◽  
Content Type ◽  
Highly Active ◽  
Extracellular Nuclease

ABSTRACTBacterial species such asShewanella oneidensisMR-1 require extracellular nucleolytic activity for the utilization of extracellular DNA (eDNA) as a source of nutrients and for the turnover of eDNA as a structural matrix component during biofilm formation. We have previously characterized two extracellular nucleases ofS. oneidensisMR-1, ExeM and ExeS. Although both are involved in biofilm formation, they are not specifically required for the utilization of eDNA as a nutrient. Here we identified and characterized EndA, a third extracellular nuclease ofShewanella. The heterologously overproduced and purified protein was highly active and rapidly degraded linear and supercoiled DNAs of various origins. Divalent metal ions (Mg2+or Mn2+) were required for function.endAis cotranscribed withphoA, an extracellular phosphatase, and is not upregulated upon phosphostarvation. Deletion ofendAabolished both extracellular degradation of DNA byS. oneidensisMR-1 and the ability to use eDNA as a sole source of phosphorus. PhoA is not strictly required for the exploitation of eDNA as a nutrient. The activity of EndA prevents the formation of large cell aggregates during planktonic growth. However, in contrast to the findings for ExeM,endAdeletion had only minor effects on biofilm formation. The findings strongly suggest that the extracellular nucleases ofS. oneidensisexert specific functions required under different conditions.

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