scholarly journals Identification of Different Putative Outer Membrane Electron Conduits Necessary for Fe(III) Citrate, Fe(III) Oxide, Mn(IV) Oxide, or Electrode Reduction byGeobacter sulfurreducens

2018 ◽  
Vol 200 (19) ◽  
Author(s):  
Fernanda Jiménez Otero ◽  
Chi Ho Chan ◽  
Daniel R. Bond
Keyword(s):  
Electron Transfer ◽  
Outer Membrane ◽  
Electron Acceptor ◽  
Gene Clusters ◽  
Geobacter Sulfurreducens ◽  
Transcriptional Analysis ◽  
Metal Reduction ◽  
Wild Type ◽  
Redox Proteins ◽  
Content Type

ABSTRACTAt least five gene clusters in theGeobacter sulfurreducensgenome encode putative “electron conduits” implicated in electron transfer across the outer membrane, each containing a periplasmic multihemec-type cytochrome, integral outer membrane anchor, and outer membrane redox lipoprotein(s). Markerless single-gene-cluster deletions and all possible multiple-deletion combinations were constructed and grown with soluble Fe(III) citrate, Fe(III) and Mn(IV) oxides, and graphite electrodes poised at +0.24 V and −0.1 V versus the standard hydrogen electrode (SHE). Different gene clusters were necessary for reduction of each electron acceptor. During metal oxide reduction, deletion of the previously describedomcBCcluster caused defects, but deletion of additional components in an ΔomcBCbackground, such asextEFG, were needed to produce defects greater than 50% compared to findings with the wild type. Deletion of all five gene clusters abolished all metal reduction. During electrode reduction, only the ΔextABCDmutant had a severe growth defect at both redox potentials, while this mutation did not affect Fe(III) oxide, Mn(IV) oxide, or Fe(III) citrate reduction. Some mutants containing only one cluster were able to reduce particular terminal electron acceptors better than the wild type, suggesting routes for improvement by targeting specific electron transfer pathways. Transcriptomic comparisons between fumarate and electrode-based growth conditions showed all of theseextclusters to be constitutive, and transcriptional analysis of the triple-deletion strain containing onlyextABCDdetected no significant changes in expression of genes encoding known redox proteins or pilus components. These genetic experiments reveal new outer membrane conduit complexes necessary for growth ofG. sulfurreducens, depending on the available extracellular electron acceptor.IMPORTANCEGram-negative metal-reducing bacteria utilize electron conduits, chains of redox proteins spanning the outer membrane, to transfer electrons to the extracellular surface. Only one pathway for electron transfer across the outer membrane ofGeobacter sulfurreducenshas been linked to Fe(III) reduction. However,G. sulfurreducensis able to respire a wide array of extracellular substrates. Here we present the first combinatorial genetic analysis of five different electron conduits via creation of new markerless deletion strains and complementation vectors. Multiple conduit gene clusters appear to have overlapping roles, including two that have never been linked to metal reduction. Another recently described cluster (ExtABCD) was the only electron conduit essential during electrode reduction, a substrate of special importance to biotechnological applications of this organism.

scholarly journals Identification of different putative outer membrane electron conduits necessary for Fe(III) citrate, Fe(III) oxide, Mn(IV) oxide, or electrode reduction by Geobacter sulfurreducens

2017 ◽  
Author(s):  
Fernanda Jiménez Otero ◽  
Chi Ho Chan ◽  
Daniel R. Bond
Keyword(s):  
Electron Transfer ◽  
Outer Membrane ◽  
Electron Acceptor ◽  
Single Gene ◽  
Gene Clusters ◽  
Geobacter Sulfurreducens ◽  
Transcriptional Analysis ◽  
Growth Defect ◽  
Redox Potentials ◽  
Wild Type

AbstractAt least five gene clusters in the Geobacter sulfurreducens genome encode putative ‘electron conduits’ implicated in electron transfer across the outer membrane, each containing a periplasmic multiheme c-type cytochrome, integral outer membrane anchor, and outer membrane redox lipoprotein(s). Markerless single gene cluster deletions and all possible multiple deletion combinations were constructed and grown with soluble Fe(III) citrate, Fe(III)- and Mn(IV)-oxides, and graphite electrodes poised at +0.24 V and −0.1 V vs. SHE. Different gene clusters were necessary for reduction of each electron acceptor. During metal oxide reduction, deletion of the previously described omcBC cluster caused defects, but deletion of additional components in an ΔomcBC background, such as extEFG, were needed to produce defects greater than 50% compared to wild type. Deletion of all five gene clusters abolished all metal reduction. During electrode reduction, only the ΔextABCD mutant had a severe growth defect at both redox potentials, while this mutation did not affect Fe(III)-oxide, Mn(IV)-oxide, or Fe(III) citrate reduction. Some mutants containing only one cluster were able to reduce particular terminal electron acceptors better than wild type, suggesting routes for improvement by targeting specific electron transfer pathways. Transcriptomic comparisons between fumarate and electrode-based growth showed all of these ext clusters to be constitutive, and transcriptional analysis of the triple-deletion strain containing only extABCD detected no significant changes in expression of known redox proteins or pili components. These genetic experiments reveal new outer membrane conduit complexes necessary for growth of G. sulfurreducens, depending on the available extracellular electron acceptor.

scholarly journals Identification of different putative outer membrane electron conduits necessary for Fe(III) citrate, Fe(III)-oxide, Mn(IV)-oxide, or electrode reduction by Geobacter sulfurreducens .

2018 ◽  
Author(s):  
Fernanda Jiménez Otero ◽  
Chi Ho Chan ◽  
Daniel R Bond
Keyword(s):  
Electron Transfer ◽  
Outer Membrane ◽  
Electron Acceptor ◽  
Single Gene ◽  
Gene Clusters ◽  
Geobacter Sulfurreducens ◽  
Transcriptional Analysis ◽  
Growth Defect ◽  
Redox Potentials ◽  
Wild Type

At least five gene clusters in the Geobacter sulfurreducens genome encode putative ‘electron conduits’ implicated in electron transfer across the outer membrane, each containing a periplasmic multiheme c -type cytochrome, integral outer membrane anchor, and outer membrane redox lipoprotein(s). Markerless single gene cluster deletions and all possible multiple deletion combinations were constructed and grown with soluble Fe(III) citrate, Fe(III)- and Mn(IV)-oxides, and graphite electrodes poised at +0.24 V and -0.1 V vs. SHE. Different gene clusters were necessary for reduction of each electron acceptor. During metal oxide reduction, deletion of the previously described omcBC cluster caused defects, but deletion of additional components in an Δ omcBC background, such as extEFG , were needed to produce defects greater than 50% compared to wild type. Deletion of all five gene clusters abolished all metal reduction. During electrode reduction, only the Δ extABCD mutant had a severe growth defect at both redox potentials, while this mutation did not affect Fe(III)-oxide, Mn(IV)-oxide, or Fe(III) citrate reduction. Some mutants containing only one cluster were able to reduce particular terminal electron acceptors better than wild type, suggesting routes for improvement by targeting specific electron transfer pathways. Transcriptomic comparisons between fumarate and electrode-based growth showed all of these ext clusters to be constitutive, and transcriptional analysis of the triple-deletion strain containing only extABCD detected no significant changes in expression of known redox proteins or pili components. These genetic experiments reveal new outer membrane conduit complexes necessary for growth of G. sulfurreducens , depending on the available extracellular electron acceptor.

scholarly journals Divergent Nrf Family Proteins and MtrCAB Homologs Facilitate Extracellular Electron Transfer inAeromonas hydrophila

2018 ◽  
Vol 84 (23) ◽  
Author(s):  
Bridget E. Conley ◽  
Peter J. Intile ◽  
Daniel R. Bond ◽  
Jeffrey A. Gralnick
Keyword(s):  
Electron Transfer ◽  
Outer Membrane ◽  
Aeromonas Hydrophila ◽  
Shewanella Oneidensis ◽  
Geobacter Sulfurreducens ◽  
Model Systems ◽  
Extracellular Electron Transfer ◽  
Metal Reduction ◽  
Inner Membrane ◽  
Content Type

ABSTRACTExtracellular electron transfer (EET) is a strategy for respiration in which electrons generated from metabolism are moved outside the cell to a terminal electron acceptor, such as iron or manganese oxide. EET has primarily been studied in two model systems,Shewanella oneidensisandGeobacter sulfurreducens. Metal reduction has also been reported in numerous microorganisms, includingAeromonasspp., which are ubiquitousGammaproteobacteriafound in aquatic ecosystems, with some species capable of pathogenesis in humans and fish. Genomic comparisons ofAeromonasspp. revealed a potential outer membrane conduit homologous toS. oneidensisMtrCAB. While the ability to respire metals and mineral oxides is not widespread in the genusAeromonas, 90% of the sequencedAeromonas hydrophilaisolates contain MtrCAB homologs.A. hydrophilaATCC 7966 mutants lackingmtrAare unable to reduce metals. Expression ofA. hydrophila mtrCABin anS. oneidensismutant lacking homologous components restored metal reduction. Although the outer membrane conduits for metal reduction were similar, homologs of theS. oneidensisinner membrane and periplasmic EET components CymA, FccA, and CctA were not found inA. hydrophila. We characterized a cluster of genes predicted to encode components related to a formate-dependent nitrite reductase (NrfBCD), here named NetBCD (forNrf-likeelectrontransfer), and a predicted diheme periplasmic cytochrome, PdsA (periplasmicdihemeshuttle). We present genetic evidence that proteins encoded by this cluster facilitate electron transfer from the cytoplasmic membrane across the periplasm to the MtrCAB conduit and function independently from an authentic NrfABCD system.A. hydrophilamutants lackingpdsAandnetBCDwere unable to reduce metals, while heterologous expression of these genes could restore metal reduction in anS. oneidensismutant background. EET may therefore allowA. hydrophilaand other species ofAeromonasto persist and thrive in iron- or manganese-rich oxygen-limited environments.IMPORTANCEMetal-reducing microorganisms are used for electricity production, bioremediation of toxic compounds, wastewater treatment, and production of valuable compounds. Despite numerous microorganisms being reported to reduce metals, the molecular mechanism has primarily been studied in two model systems,Shewanella oneidensisandGeobacter sulfurreducens. We have characterized the mechanism of extracellular electron transfer inAeromonas hydrophila, which uses the well-studiedShewanellasystem, MtrCAB, to move electrons across the outer membrane; however, mostAeromonasspp. appear to use a novel mechanism to transfer electrons from the inner membrane through the periplasm and to the outer membrane. The conserved use of MtrCAB inShewanellaspp. andAeromonasspp. for metal reduction and conserved genomic architecture of metal reduction genes inAeromonasspp. may serve as genomic markers for identifying metal-reducing microorganisms from genomic or transcriptomic sequencing. Understanding the variety of pathways used to reduce metals can allow for optimization and more efficient design of microorganisms used for practical applications.

scholarly journals Interspecies Electron Transfer via Hydrogen and Formate Rather than Direct Electrical Connections in Cocultures of Pelobacter carbinolicus and Geobacter sulfurreducens

2012 ◽  
Vol 78 (21) ◽  
pp. 7645-7651 ◽  
Author(s):  
Amelia-Elena Rotaru ◽  
Pravin M. Shrestha ◽  
Fanghua Liu ◽  
Toshiyuki Ueki ◽  
Kelly Nevin ◽  
...  
Keyword(s):  
Electron Transfer ◽  
Formate Dehydrogenase ◽  
Geobacter Sulfurreducens ◽  
Close Relative ◽  
Wild Type ◽  
Geobacter Metallireducens ◽  
Content Type ◽  
Interspecies Electron Transfer ◽  
Essential Components ◽  
Electrical Connections

ABSTRACTDirect interspecies electron transfer (DIET) is an alternative to interspecies H2/formate transfer as a mechanism for microbial species to cooperatively exchange electrons during syntrophic metabolism. To understand what specific properties contribute to DIET, studies were conducted withPelobacter carbinolicus, a close relative ofGeobacter metallireducens, which is capable of DIET.P. carbinolicusgrew in coculture withGeobacter sulfurreducenswith ethanol as the electron donor and fumarate as the electron acceptor, conditions under whichG. sulfurreducensformed direct electrical connections withG. metallireducens. In contrast to the cell aggregation associated with DIET,P. carbinolicusandG. sulfurreducensdid not aggregate. Attempts to initiate cocultures with a genetically modified strain ofG. sulfurreducensincapable of both H2and formate utilization were unsuccessful, whereas cocultures readily grew with mutant strains capable of formate but not H2uptake or vice versa. The hydrogenase mutant ofG. sulfurreducenscompensated, in cocultures, with significantly increased formate dehydrogenase gene expression. In contrast, the transcript abundance of a hydrogenase gene was comparable in cocultures with that for the formate dehydrogenase mutant ofG. sulfurreducensor the wild type, suggesting that H2was the primary electron carrier in the wild-type cocultures. Cocultures were also initiated with strains ofG. sulfurreducensthat could not produce pili or OmcS, two essential components for DIET. The finding thatP. carbinolicusexchanged electrons withG. sulfurreducensvia interspecies transfer of H2/formate rather than DIET demonstrates that not all microorganisms that can grow syntrophically are capable of DIET and that closely related microorganisms may use significantly different strategies for interspecies electron exchange.

scholarly journals Adaptation to Disruption of the Electron Transfer Pathway for Fe(III) Reduction in Geobacter sulfurreducens

2005 ◽  
Vol 187 (17) ◽  
pp. 5918-5926 ◽  
Author(s):  
Ching Leang ◽  
L. A. Adams ◽  
K.-J. Chin ◽  
K. P. Nevin ◽  
B. A. Methé ◽  
...  
Keyword(s):  
Electron Transfer ◽  
Steady State ◽  
Electron Transport ◽  
Outer Membrane ◽  
Geobacter Sulfurreducens ◽  
Significant Shift ◽  
Deficient Mutant ◽  
Wild Type ◽  
Transcript Levels ◽  
Electron Transfer Pathway

ABSTRACT Previous studies demonstrated that an outer membrane c-type cytochrome, OmcB, was involved in Fe(III) reduction in Geobacter sulfurreducens. An OmcB-deficient mutant was greatly impaired in its ability to reduce both soluble and insoluble Fe(III). Reintroducing omcB restored the capacity for Fe(III) reduction at a level proportional to the level of OmcB production. Here, we report that the OmcB-deficient mutant gradually adapted to grow on soluble Fe(III) but not insoluble Fe(III). The adapted OmcB-deficient mutant reduced soluble Fe(III) at a rate comparable to that of the wild type, but the cell yield of the mutant was only ca. 60% of that of the wild type under steady-state culturing conditions. Analysis of proteins and transcript levels demonstrated that expression of several membrane-associated cytochromes was higher in the adapted mutant than in the wild type. Further comparison of transcript levels during steady-state growth on Fe(III) citrate with a whole-genome DNA microarray revealed a significant shift in gene expression in an apparent attempt to adapt metabolism to the impaired electron transport to Fe(III). These results demonstrate that, although there are many other membrane-bound c-type cytochromes in G. sulfurreducens, increased expression of these cytochromes cannot completely compensate for the loss of OmcB. The concept that outer membrane cytochromes are promiscuous reductases that are interchangeable in function appears to be incorrect. Furthermore, the results indicate that there may be different mechanisms for electron transfer to soluble Fe(III) and insoluble Fe(III) oxides in G. sulfurreducens, which emphasizes the importance of studying electron transport to the environmentally relevant Fe(III) oxides.

scholarly journals GeobacterStrains Expressing Poorly Conductive Pili Reveal Constraints on Direct Interspecies Electron Transfer Mechanisms

mBio ◽  
2018 ◽  
Vol 9 (4) ◽  
Author(s):  
Toshiyuki Ueki ◽  
Kelly P. Nevin ◽  
Amelia-Elena Rotaru ◽  
Li-Ying Wang ◽  
Joy E. Ward ◽  
...  
Keyword(s):  
Electron Transfer ◽  
Electron Flow ◽  
Geobacter Sulfurreducens ◽  
Wild Type ◽  
Anaerobic Digesters ◽  
Content Type ◽  
Direct Interspecies Electron Transfer ◽  
Interspecies Electron Transfer ◽  
Anaerobic Soils ◽  
Pilin Gene

ABSTRACTCytochrome-to-cytochrome electron transfer and electron transfer along conduits of multiple extracellular magnetite grains are often proposed as strategies for direct interspecies electron transfer (DIET) that do not require electrically conductive pili (e-pili). However, physical evidence for these proposed DIET mechanisms has been lacking. To investigate these possibilities further, we constructedGeobacter metallireducensstrain Aro-5, in which the wild-type pilin gene was replaced with thearo-5pilin gene that was previously shown to yield poorly conductive pili inGeobacter sulfurreducensstrain Aro-5.G. metallireducensstrain Aro-5 did not reduce Fe(III) oxide and produced only low current densities, phenotypes consistent with expression of poorly conductive pili. LikeG. sulfurreducensstrain Aro-5,G. metallireducensstrain Aro-5 displayed abundant outer surface cytochromes. Cocultures initiated with wild-typeG. metallireducensas the electron-donating strain andG. sulfurreducensstrain Aro-5 as the electron-accepting strain grew via DIET. However,G. metallireducensAro-5/G. sulfurreducenswild-type cocultures did not. Cocultures initiated with the Aro-5 strains of both species grew only when amended with granular activated carbon (GAC), a conductive material known to be a conduit for DIET. Magnetite could not substitute for GAC. The inability of the two Aro-5 strains to adapt for DIET in the absence of GAC suggests that there are physical constraints on establishing DIET solely through cytochrome-to-cytochrome electron transfer or along chains of magnetite. The finding that DIET is possible with electron-accepting partners that lack highly conductive pili greatly expands the range of potential electron-accepting partners that might participate in DIET.IMPORTANCEDIET is thought to be an important mechanism for interspecies electron exchange in natural anaerobic soils and sediments in which methane is either produced or consumed, as well as in some photosynthetic mats and anaerobic digesters converting organic wastes to methane. Understanding the potential mechanisms for DIET will not only aid in modeling carbon and electron flow in these geochemically significant environments but will also be helpful for interpreting meta-omic data from as-yet-uncultured microbes in DIET-based communities and for designing strategies to promote DIET in anaerobic digesters. The results demonstrate the need to develop a better understanding of the diversity of types of e-pili in the microbial world to identify potential electron-donating partners for DIET. Novel methods for recovering as-yet-uncultivated microorganisms capable of DIET in culture will be needed to further evaluate whether DIET is possible without e-pili in the absence of conductive materials such as GAC.

scholarly journals Metal Reduction and Protein Secretion Genes Required for Iodate Reduction by Shewanella oneidensis

2018 ◽  
Vol 85 (3) ◽  
Author(s):  
Yael J. Toporek ◽  
Jung Kee Mok ◽  
Hyun Dong Shin ◽  
Brady D. Lee ◽  
M. Hope Lee ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Protein Secretion ◽  
Electron Donor ◽  
Shewanella Oneidensis ◽  
Electron Donors ◽  
Metal Reduction ◽  
Type Ii ◽  
Wild Type ◽  
Content Type ◽  
Reduction Activity

ABSTRACT The metal-reducing gammaproteobacterium Shewanella oneidensis reduces iodate (IO3−) as an anaerobic terminal electron acceptor. Microbial IO3− electron transport pathways are postulated to terminate with nitrate (NO3−) reductase, which reduces IO3− as an alternative electron acceptor. Recent studies with S. oneidensis, however, have demonstrated that NO3− reductase is not involved in IO3− reduction. The main objective of the present study was to determine the metal reduction and protein secretion genes required for IO3− reduction by Shewanella oneidensis with lactate, formate, or H2 as the electron donor. With all electron donors, the type I and type V protein secretion mutants retained wild-type IO3− reduction activity, while the type II protein secretion mutant lacking the outer membrane secretin GspD was impaired in IO3− reduction. Deletion mutants lacking the cyclic AMP receptor protein (CRP), cytochrome maturation permease CcmB, and inner membrane-tethered c-type cytochrome CymA were impaired in IO3− reduction with all electron donors, while deletion mutants lacking c-type cytochrome MtrA and outer membrane β-barrel protein MtrB of the outer membrane MtrAB module were impaired in IO3− reduction with only lactate as an electron donor. With all electron donors, mutants lacking the c-type cytochromes OmcA and MtrC of the metal-reducing extracellular electron conduit MtrCAB retained wild-type IO3− reduction activity. These findings indicate that IO3− reduction by S. oneidensis involves electron donor-dependent metal reduction and protein secretion pathway components, including the outer membrane MtrAB module and type II protein secretion of an unidentified IO3− reductase to the S. oneidensis outer membrane. IMPORTANCE Microbial iodate (IO3−) reduction is a major component in the biogeochemical cycling of iodine and the bioremediation of iodine-contaminated environments; however, the molecular mechanism of microbial IO3− reduction is poorly understood. Results of the present study indicate that outer membrane (type II) protein secretion and metal reduction genes encoding the outer membrane MtrAB module of the extracellular electron conduit MtrCAB are required for IO3− reduction by S. oneidensis. On the other hand, the metal-reducing c-type cytochrome MtrC of the extracellular electron conduit is not required for IO3− reduction by S. oneidensis. These findings indicate that the IO3− electron transport pathway terminates with an as yet unidentified IO3− reductase that associates with the outer membrane MtrAB module to deliver electrons extracellularly to IO3−.

scholarly journals Regulation of Gene Expression in Shewanella oneidensis MR-1 during Electron Acceptor Limitation and Bacterial Nanowire Formation

2016 ◽  
Vol 82 (17) ◽  
pp. 5428-5443 ◽  
Author(s):  
Sarah E. Barchinger ◽  
Sahand Pirbadian ◽  
Christine Sambles ◽  
Carol S. Baker ◽  
Kar Man Leung ◽  
...  
Keyword(s):  
Gene Expression ◽  
Electron Transfer ◽  
Outer Membrane ◽  
Electron Acceptor ◽  
Shewanella Oneidensis ◽  
Oxygen Limitation ◽  
Extracellular Electron Transfer ◽  
Transport Pathways ◽  
Content Type ◽  
Genes Encoding

ABSTRACTIn limiting oxygen as an electron acceptor, the dissimilatory metal-reducing bacteriumShewanella oneidensisMR-1 rapidly forms nanowires, extensions of its outer membrane containing the cytochromes MtrC and OmcA needed for extracellular electron transfer. RNA sequencing (RNA-Seq) analysis was employed to determine differential gene expression over time from triplicate chemostat cultures that were limited for oxygen. We identified 465 genes with decreased expression and 677 genes with increased expression. The coordinated increased expression of heme biosynthesis, cytochrome maturation, and transport pathways indicates thatS. oneidensisMR-1 increases cytochrome production, including the transcription of genes encoding MtrA, MtrC, and OmcA, and transports these decaheme cytochromes across the cytoplasmic membrane during electron acceptor limitation and nanowire formation. In contrast, the expression of themtrAandmtrChomologsmtrFandmtrDeither remains unaffected or decreases under these conditions. TheompWgene, encoding a small outer membrane porin, has 40-fold higher expression during oxygen limitation, and it is proposed that OmpW plays a role in cation transport to maintain electrical neutrality during electron transfer. The genes encoding the anaerobic respiration regulator cyclic AMP receptor protein (CRP) and the extracytoplasmic function sigma factor RpoE are among the transcription factor genes with increased expression. RpoE might function by signaling the initial response to oxygen limitation. Our results show that RpoE activates transcription from promoters upstream ofmtrCandomcA. The transcriptome and mutant analyses ofS. oneidensisMR-1 nanowire production are consistent with independent regulatory mechanisms for extending the outer membrane into tubular structures and for ensuring the electron transfer function of the nanowires.IMPORTANCEShewanella oneidensisMR-1 has the capacity to transfer electrons to its external surface using extensions of the outer membrane called bacterial nanowires. These bacterial nanowires link the cell's respiratory chain to external surfaces, including oxidized metals important in bioremediation, and explain whyS. oneidensiscan be utilized as a component of microbial fuel cells, a form of renewable energy. In this work, we use differential gene expression analysis to focus on which genes function to produce the nanowires and promote extracellular electron transfer during oxygen limitation. Among the genes that are expressed at high levels are those encoding cytochrome proteins necessary for electron transfer.Shewanellacoordinates the increased expression of regulators, metabolic pathways, and transport pathways to ensure that cytochromes efficiently transfer electrons along the nanowires.

scholarly journals Going Wireless: Fe(III) Oxide Reduction without Pili by Geobacter sulfurreducens Strain JS-1

2014 ◽  
Vol 80 (14) ◽  
pp. 4331-4340 ◽  
Author(s):  
Jessica A. Smith ◽  
Pier-Luc Tremblay ◽  
Pravin Malla Shrestha ◽  
Oona L. Snoeyenbos-West ◽  
Ashley E. Franks ◽  
...  
Keyword(s):  
Electron Transfer ◽  
Electron Transport ◽  
Type Strain ◽  
Culture Medium ◽  
Wild Type Strain ◽  
Geobacter Sulfurreducens ◽  
Electron Shuttle ◽  
Oxide Reduction ◽  
Wild Type ◽  
Content Type

ABSTRACTPrevious studies have suggested that the conductive pili ofGeobacter sulfurreducensare essential for extracellular electron transfer to Fe(III) oxides and for optimal long-range electron transport through current-producing biofilms. The KN400 strain ofG. sulfurreducensreduces poorly crystalline Fe(III) oxide more rapidly than the more extensively studied DL-1 strain. Deletion of the gene encoding PilA, the structural pilin protein, in strain KN400 inhibited Fe(III) oxide reduction. However, low rates of Fe(III) reduction were detected after extended incubation (>30 days) in the presence of Fe(III) oxide. After seven consecutive transfers, the PilA-deficient strain adapted to reduce Fe(III) oxide as fast as the wild type. Microarray, whole-genome resequencing, proteomic, and gene deletion studies indicated that this adaptation was associated with the production of larger amounts of thec-type cytochrome PgcA, which was released into the culture medium. It is proposed that the extracellular cytochrome acts as an electron shuttle, promoting electron transfer from the outer cell surface to Fe(III) oxides. The adapted PilA-deficient strain competed well with the wild-type strain when both were grown together on Fe(III) oxide. However, when 50% of the culture medium was replaced with fresh medium every 3 days, the wild-type strain outcompeted the adapted strain. A possible explanation for this is that the necessity to produce additional PgcA, to replace the PgcA being continually removed, put the adapted strain at a competitive disadvantage, similar to the apparent selection against electron shuttle-producing Fe(III) reducers in many anaerobic soils and sediments. Despite increased extracellular cytochrome production, the adapted PilA-deficient strain produced low levels of current, consistent with the concept that long-range electron transport throughG. sulfurreducensbiofilms is more effective via pili.

scholarly journals The Serratia marcescens Siderophore Serratiochelin Is Necessary for Full Virulence during Bloodstream Infection

2020 ◽  
Vol 88 (8) ◽  
Author(s):  
Danelle R. Weakland ◽  
Sara N. Smith ◽  
Bailey Bell ◽  
Ashootosh Tripathi ◽  
Harry L. T. Mobley
Keyword(s):  
Serratia Marcescens ◽  
Bloodstream Infection ◽  
Iron Acquisition ◽  
Gene Clusters ◽  
Clinical Isolates ◽  
Wild Type ◽  
Serratia Plymuthica ◽  
Content Type ◽  
Cas Assay ◽  
Essential Nutrient

ABSTRACT Serratia marcescens is a bacterium frequently found in the environment, but over the last several decades it has evolved into a concerning clinical pathogen, causing fatal bacteremia. To establish such infections, pathogens require specific nutrients; one very limited but essential nutrient is iron. We sought to characterize the iron acquisition systems in S. marcescens isolate UMH9, which was recovered from a clinical bloodstream infection. Using RNA sequencing (RNA-seq), we identified two predicted siderophore gene clusters (cbs and sch) that were regulated by iron. Mutants were constructed to delete each iron acquisition locus individually and in conjunction, generating both single and double mutants for the putative siderophore systems. Mutants lacking the sch gene cluster lost their iron-chelating ability as quantified by the chrome azurol S (CAS) assay, whereas the cbs mutant retained wild-type activity. Mass spectrometry-based analysis identified the chelating siderophore to be serratiochelin, a siderophore previously identified in Serratia plymuthica. Serratiochelin-producing mutants also displayed a decreased growth rate under iron-limited conditions created by dipyridyl added to LB medium. Additionally, mutants lacking serratiochelin were significantly outcompeted during cochallenge with wild-type UMH9 in the kidneys and spleen after inoculation via the tail vein in a bacteremia mouse model. This result was further confirmed by an independent challenge, suggesting that serratiochelin is required for full S. marcescens pathogenesis in the bloodstream. Nine other clinical isolates have at least 90% protein identity to the UMH9 serratiochelin system; therefore, our results are broadly applicable to emerging clinical isolates of S. marcescens causing bacteremia.

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