Transcriptomic and Proteomic Responses of the Organohalide-Respiring Bacterium Desulfoluna spongiiphila to Growth with 2,6-Dibromophenol as the Electron Acceptor

ABSTRACT Organohalide respiration is an important process in the global halogen cycle and for bioremediation. In this study, we compared the global transcriptomic and proteomic analyses of Desulfoluna spongiiphila strain AA1, an organohalide-respiring member of the Desulfobacterota isolated from a marine sponge, with 2,6-dibromophenol or with sulfate as an electron acceptor. The most significant difference of the transcriptomic analysis was the expression of one reductive dehalogenase gene cluster (rdh16), which was significantly upregulated with the addition of 2,6-dibromophenol. The corresponding protein, reductive dehalogenase RdhA16032, was detected in the proteome under treatment with 2,6-dibromophenol but not with sulfate only. There was no significant difference in corrinoid biosynthesis gene expression levels between the two treatments, indicating that the production of corrinoid in D. spongiiphila is constitutive or not specific for organohalide versus sulfate respiration. Electron-transporting proteins or mediators unique for reductive dehalogenation were not revealed in our analysis, and we hypothesize that reductive dehalogenation may share an electron-transporting system with sulfate reduction. The metabolism of D. spongiiphila, predicted from transcriptomic and proteomic results, demonstrates high metabolic versatility and provides insights into the survival strategies of a marine sponge symbiont in an environment rich in organohalide compounds and other secondary metabolites. IMPORTANCE Respiratory reductive dehalogenation is an important process in the overall cycling of both anthropogenic and natural organohalide compounds. Marine sponges produce a vast array of bioactive compounds as secondary metabolites, including diverse halogenated compounds that may enrich for dehalogenating bacteria. Desulfoluna spongiiphila strain AA1 was originally enriched and isolated from the marine sponge Aplysina aerophoba and can grow with both brominated compounds and sulfate as electron acceptors for respiration. An understanding of the overall gene expression and the protein production profile in response to organohalides is needed to identify the full complement of genes or enzymes involved in organohalide respiration. Elucidating the metabolic capacity of this sponge-associated bacterium lays the foundation for understanding how dehalogenating bacteria may control the fate of organohalide compounds in sponges and their role in a symbiotic organobromine cycle.

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Genome-Guided Identification of Organohalide-RespiringDeltaproteobacteriafrom the Marine Environment

mBio ◽

10.1128/mbio.02471-18 ◽

2018 ◽

Vol 9 (6) ◽

Cited By ~ 3

Author(s):

Jie Liu ◽

Max M. Häggblom

Keyword(s):

Marine Environment ◽

Gene Diversity ◽

Anthropogenic Activities ◽

Reductive Dehalogenation ◽

Important Process ◽

Organohalide Respiration ◽

Content Type ◽

Reductive Dehalogenase ◽

Comprehensive Survey ◽

Reductive Dehalogenase Genes

ABSTRACTOrganohalide compounds are widespread in the environment as a result of both anthropogenic activities and natural production. The marine environment, in particular, is a major reservoir of organohalides, and reductive dehalogenation is thought to be an important process in the overall cycling of these compounds.Deltaproteobacteriaare important members of the marine microbiota with diverse metabolic capacities, and reductive dehalogenation has been observed in someDeltaproteobacteria. In this study, a comprehensive survey ofDeltaproteobacteriagenomes revealed that approximately 10% contain reductive dehalogenase (RDase) genes, which are found within a common gene neighborhood. The dehalogenating potential of select RDase A-containingDeltaproteobacteriaand their gene expression were experimentally verified. ThreeDeltaproteobacteriastrains isolated from marine environments representing diverse species,Halodesulfovibrio marinisediminis,Desulfuromusa kysingii, andDesulfovibrio bizertensis, were shown to reductively dehalogenate bromophenols and utilize them as terminal electron acceptors in organohalide respiration. Their debrominating activity was not inhibited by sulfate or elemental sulfur, and these species are either sulfate- or sulfur-reducing bacteria. The analysis of RDase A gene transcripts indicated significant upregulation induced by 2,6-dibromophenol. This study extends our knowledge of the phylogenetic diversity of organohalide-respiring bacteria and their functional RDase A gene diversity. The identification of reductive dehalogenase genes in diverseDeltaproteobacteriaand confirmation of their organohalide-respiring capability suggest thatDeltaproteobacteriaplay an important role in natural organohalide cycling.IMPORTANCEThe marine environment is a major reservoir for both anthropogenic and natural organohalides, and reductive dehalogenation is thought to be an important process in the overall cycling of these compounds. Here we demonstrate that the capacity of organohalide respiration appears to be widely distributed in members of marineDeltaproteobacteria. The identification of reductive dehalogenase genes in diverseDeltaproteobacteriaand the confirmation of their dehalogenating activity through functional assays and transcript analysis in select isolates extend our knowledge of organohalide-respiringDeltaproteobacteriadiversity. The presence of functional reductive dehalogenase genes in diverseDeltaproteobacteriaimplies that they may play an important role in organohalide respiration in the environment.

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Dehalococcoides mccartyi Strain DCMB5 Respires a Broad Spectrum of Chlorinated Aromatic Compounds

Applied and Environmental Microbiology ◽

10.1128/aem.02597-14 ◽

2014 ◽

Vol 81 (2) ◽

pp. 587-596 ◽

Cited By ~ 36

Author(s):

Marlén Pöritz ◽

Christian L. Schiffmann ◽

Gerd Hause ◽

Ulrike Heinemann ◽

Jana Seifert ◽

...

Keyword(s):

Electron Acceptor ◽

Aromatic Compounds ◽

Lag Phase ◽

Organohalide Respiration ◽

Content Type ◽

Reductive Dehalogenase ◽

Dehalococcoides Mccartyi ◽

Type Iv ◽

Transmission Electron ◽

Almost All

ABSTRACTPolyhalogenated aromatic compounds are harmful environmental contaminants and tend to persist in anoxic soils and sediments.Dehalococcoides mccartyistrain DCMB5, a strain originating from dioxin-polluted river sediment, was examined for its capacity to dehalogenate diverse chloroaromatic compounds. Strain DCMB5 used hexachlorobenzenes, pentachlorobenzenes, all three tetrachlorobenzenes, and 1,2,3-trichlorobenzene as well as 1,2,3,4-tetra- and 1,2,4-trichlorodibenzo-p-dioxin as electron acceptors for organohalide respiration. In addition, 1,2,3-trichlorodibenzo-p-dioxin and 1,3-, 1,2-, and 1,4-dichlorodibenzo-p-dioxin were dechlorinated, the latter to the nonchlorinated congener with a remarkably short lag phase of 1 to 4 days following transfer. Strain DCMB5 also dechlorinated pentachlorophenol and almost all tetra- and trichlorophenols. Tetrachloroethene was dechlorinated to trichloroethene and served as an electron acceptor for growth. To relate selected dechlorination activities to the expression of specific reductive dehalogenase genes, the proteomes of 1,2,3-trichlorobenzene-, pentachlorobenzene-, and tetrachloroethene-dechlorinating cultures were analyzed. Dcmb_86, an ortholog of the chlorobenzene reductive dehalogenase CbrA, was the most abundant reductive dehalogenase during growth with each electron acceptor, suggesting its pivotal role in organohalide respiration of strain DCMB5. Dcmb_1041 was specifically induced, however, by both chlorobenzenes, whereas 3 putative reductive dehalogenases, Dcmb_1434, Dcmb_1339, and Dcmb_1383, were detected only in tetrachloroethene-grown cells. The proteomes also harbored a type IV pilus protein and the components for its assembly, disassembly, and secretion. In addition, transmission electron microscopy of DCMB5 revealed an irregular mode of cell division as well as the presence of pili, indicating that pilus formation is a feature ofD. mccartyiduring organohalide respiration.

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The MarR-Type Regulator Rdh2R RegulatesrdhGene Transcription in Dehalococcoides mccartyi Strain CBDB1

Journal of Bacteriology ◽

10.1128/jb.00419-16 ◽

2016 ◽

Vol 198 (23) ◽

pp. 3130-3141 ◽

Cited By ~ 7

Author(s):

Lydia Krasper ◽

Hauke Lilie ◽

Anja Kublik ◽

Lorenz Adrian ◽

Ralph Golbik ◽

...

Keyword(s):

Heterologous Expression ◽

Direct Repeat ◽

Negative Regulator ◽

Control Of Gene Expression ◽

Organohalide Respiration ◽

Content Type ◽

Reductive Dehalogenase ◽

Dehalococcoides Mccartyi ◽

Transcriptional Start Sites

ABSTRACTReductive dehalogenases are essential enzymes in organohalide respiration and consist of a catalytic subunit A and a membrane protein B, encoded byrdhABgenes. Thirty-twordhABgenes exist in the genome ofDehalococcoides mccartyistrain CBDB1. To gain a first insight into the regulation ofrdhoperons, the control of gene expression of twordhABgenes (cbdbA1453/cbdbA1452 and cbdbA1455/cbdbA1454) by the MarR-type regulator Rdh2R (cbdbA1456) encoded directly upstream was studied using heterologous expression andin vitrostudies. Promoter-lacZreporter fusions were generated and integrated into the genome of theEscherichia colihost. ThelacZreporter activities of bothrdhApromoters decreased upon transformation of the cells with a plasmid carrying therdh2Rgene, suggesting that Rdh2R acts as repressor, whereas thelacZreporter activity of therdh2Rpromoter was not affected. The transcriptional start sites of bothrdhAgenes in strain CBDB1 and/or the heterologous host mapped to a conserved direct repeat with 11- to 13-bp half-sites. DNase I footprinting revealed binding of Rdh2R to a ∼30-bp sequence covering the complete direct repeat in both promoters, including the transcriptional start sites. Equilibrium sedimentation ultracentrifugation revealed that Rdh2R binds as tetramer to the direct-repeat motif of therdhA(cbdbA1455) promoter. Using electrophoretic mobility shift assays, a similar binding affinity was found for bothrdhApromoters. In the presence of only one half-site of the direct repeat, the interaction was strongly reduced, suggesting a positive cooperativity of binding, for which unusual short palindromes within the direct-repeat half-sites might play an important role.IMPORTANCEDehalococcoides mccartyistrains are obligate anaerobes that grow by organohalide respiration. They have an important bioremediation potential because they are capable of reducing a multitude of halogenated compounds to less toxic products. We are now beginning to understand how these organisms make use of this large catabolic potential, wherebyD. mccartyiexpresses dehalogenases in a compound-specific fashion. MarR-type regulators are often encoded in the vicinity of reductive dehalogenase genes. In this study, we made use of heterologous expression andin vitrostudies to demonstrate that the MarR-type transcription factor Rdh2R acts as a negative regulator. We identify its binding site on the DNA, which suggests a mechanism by which it controls the expression of two adjacent reductive dehalogenase operons.

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Impact of Ammonium on Syntrophic Organohalide-Respiring and Fermenting Microbial Communities

mSphere ◽

10.1128/msphere.00053-16 ◽

2016 ◽

Vol 1 (2) ◽

Cited By ~ 7

Author(s):

Anca G. Delgado ◽

Devyn Fajardo-Williams ◽

Kylie L. Kegerreis ◽

Prathap Parameswaran ◽

Rosa Krajmalnik-Brown

Keyword(s):

Chlorinated Solvents ◽

Reductive Dehalogenation ◽

Ammonium Concentration ◽

Organohalide Respiration ◽

Content Type ◽

Dehalococcoides Mccartyi ◽

Metabolic Capability ◽

Subsurface Environments ◽

Fermentative Bacteria ◽

High Concentrations

ABSTRACT Contamination with ammonium and chlorinated solvents has been reported in numerous subsurface environments, and these chemicals bring significant challenges for in situ bioremediation. Dehalococcoides mccartyi is able to reduce the chlorinated solvent trichloroethene to the nontoxic end product ethene. Fermentative bacteria are of central importance for organohalide respiration and bioremediation to provide D. mccartyi with H2, their electron donor, acetate, their carbon source, and other micronutrients. In this study, we found that high concentrations of ammonium negatively correlated with rates of trichloroethene reductive dehalogenation and fermentation. However, detoxification of trichloroethene to nontoxic ethene occurred even at ammonium concentrations typical of those found in animal waste (up to 2 g liter−1 NH4 +-N). To date, hundreds of subsurface environments have been bioremediated through the unique metabolic capability of D. mccartyi. These findings extend our knowledge of D. mccartyi and provide insight for bioremediation of sites contaminated with chlorinated solvents and ammonium. Syntrophic interactions between organohalide-respiring and fermentative microorganisms are critical for effective bioremediation of halogenated compounds. This work investigated the effect of ammonium concentration (up to 4 g liter−1 NH4 +-N) on trichloroethene-reducing Dehalococcoides mccartyi and Geobacteraceae in microbial communities fed lactate and methanol. We found that production of ethene by D. mccartyi occurred in mineral medium containing ≤2 g liter−1 NH4 +-N and in landfill leachate. For the partial reduction of trichloroethene (TCE) to cis-dichloroethene (cis-DCE) at ≥1 g liter−1 NH4 +-N, organohalide-respiring dynamics shifted from D. mccartyi and Geobacteraceae to mainly D. mccartyi. An increasing concentration of ammonium was coupled to lower metabolic rates, longer lag times, and lower gene abundances for all microbial processes studied. The methanol fermentation pathway to acetate and H2 was conserved, regardless of the ammonium concentration provided. However, lactate fermentation shifted from propionic to acetogenic at concentrations of ≥2 g liter−1 NH4 +-N. Our study findings strongly support a tolerance of D. mccartyi to high ammonium concentrations, highlighting the feasibility of organohalide respiration in ammonium-contaminated subsurface environments. IMPORTANCE Contamination with ammonium and chlorinated solvents has been reported in numerous subsurface environments, and these chemicals bring significant challenges for in situ bioremediation. Dehalococcoides mccartyi is able to reduce the chlorinated solvent trichloroethene to the nontoxic end product ethene. Fermentative bacteria are of central importance for organohalide respiration and bioremediation to provide D. mccartyi with H2, their electron donor, acetate, their carbon source, and other micronutrients. In this study, we found that high concentrations of ammonium negatively correlated with rates of trichloroethene reductive dehalogenation and fermentation. However, detoxification of trichloroethene to nontoxic ethene occurred even at ammonium concentrations typical of those found in animal waste (up to 2 g liter−1 NH4 +-N). To date, hundreds of subsurface environments have been bioremediated through the unique metabolic capability of D. mccartyi. These findings extend our knowledge of D. mccartyi and provide insight for bioremediation of sites contaminated with chlorinated solvents and ammonium.

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Physiological Adaptation of Desulfitobacterium hafniense Strain TCE1 to Tetrachloroethene Respiration

Applied and Environmental Microbiology ◽

10.1128/aem.02471-10 ◽

2011 ◽

Vol 77 (11) ◽

pp. 3853-3859 ◽

Cited By ~ 28

Author(s):

Laure Prat ◽

Julien Maillard ◽

Régis Grimaud ◽

Christof Holliger

Keyword(s):

Electron Acceptor ◽

Growth Conditions ◽

Mass Spectrometry Analysis ◽

Physiological Adaptation ◽

Terminal Electron Acceptor ◽

Organohalide Respiration ◽

Content Type ◽

Spectrometry Analysis ◽

Metabolic Versatility ◽

Desulfitobacterium Hafniense

ABSTRACTDesulfitobacteriumspp. are ubiquitous organisms with a broad metabolic versatility, and some isolates have the ability to use tetrachloroethene (PCE) as terminal electron acceptor. In order to identify proteins involved in this organohalide respiration process, a comparative proteomic analysis was performed. Soluble and membrane-associated proteins obtained from cells ofDesulfitobacterium hafniensestrain TCE1 that were growing on different combinations of the electron donors lactate and hydrogen and the electron acceptors PCE and fumarate were analyzed. Among proteins increasingly expressed in the presence of PCE compared to fumarate as electron acceptor, a total of 57 proteins were identified by mass spectrometry analysis, revealing proteins involved in stress response and associated regulation pathways, such as PspA, GroEL, and CodY, and also proteins potentially participating in carbon and energy metabolism, such as proteins of the Wood-Ljungdahl pathway and electron transfer flavoproteins. These proteomic results suggest thatD. hafniensestrain TCE1 adapts its physiology to face the relative unfavorable growth conditions during an apparent opportunistic organohalide respiration.

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Genomic, Proteomic, and Biochemical Analysis of the Organohalide Respiratory Pathway in Desulfitobacterium dehalogenans

Journal of Bacteriology ◽

10.1128/jb.02370-14 ◽

2014 ◽

Vol 197 (5) ◽

pp. 893-904 ◽

Cited By ~ 25

Author(s):

Thomas Kruse ◽

Bram A. van de Pas ◽

Ariane Atteia ◽

Klaas Krab ◽

Wilfred R. Hagen ◽

...

Keyword(s):

Electron Transport ◽

Electron Acceptor ◽

Electron Transport Chain ◽

Biochemical Analysis ◽

Active Components ◽

Organohalide Respiration ◽

Content Type ◽

Membrane Complex ◽

Redox Active ◽

Transport Chain

Desulfitobacterium dehalogenansis able to grow by organohalide respiration using 3-chloro-4-hydroxyphenyl acetate (Cl-OHPA) as an electron acceptor. We used a combination of genome sequencing, biochemical analysis of redox active components, and shotgun proteomics to study elements of the organohalide respiratory electron transport chain. The genome ofDesulfitobacterium dehalogenansJW/IU-DC1Tconsists of a single circular chromosome of 4,321,753 bp with a GC content of 44.97%. The genome contains 4,252 genes, including six rRNA operons and six predicted reductive dehalogenases. One of the reductive dehalogenases, CprA, is encoded by a well-characterizedcprTKZEBACDgene cluster. Redox active components were identified in concentrated suspensions of cells grown on formate and Cl-OHPA or formate and fumarate, using electron paramagnetic resonance (EPR), visible spectroscopy, and high-performance liquid chromatography (HPLC) analysis of membrane extracts. In cell suspensions, these components were reduced upon addition of formate and oxidized after addition of Cl-OHPA, indicating involvement in organohalide respiration. Genome analysis revealed genes that likely encode the identified components of the electron transport chain from formate to fumarate or Cl-OHPA. Data presented here suggest that the first part of the electron transport chain from formate to fumarate or Cl-OHPA is shared. Electrons are channeled from an outward-facing formate dehydrogenase via menaquinones to a fumarate reductase located at the cytoplasmic face of the membrane. When Cl-OHPA is the terminal electron acceptor, electrons are transferred from menaquinones to outward-facing CprA, via an as-yet-unidentified membrane complex, and potentially an extracellular flavoprotein acting as an electron shuttle between the quinol dehydrogenase membrane complex and CprA.

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Comparative Single-Cell Genomics of Chloroflexi from the Okinawa Trough Deep-Subsurface Biosphere

Applied and Environmental Microbiology ◽

10.1128/aem.00624-16 ◽

2016 ◽

Vol 82 (10) ◽

pp. 3000-3008 ◽

Cited By ~ 32

Author(s):

Heather Fullerton ◽

Craig L. Moyer

Keyword(s):

Single Cell ◽

Okinawa Trough ◽

Reductive Dehalogenation ◽

Rrna Gene ◽

Ssu Rrna ◽

Metabolic Potential ◽

Deep Subsurface ◽

Content Type ◽

Reductive Dehalogenase ◽

The Okinawa Trough

ABSTRACTChloroflexismall-subunit (SSU) rRNA gene sequences are frequently recovered from subseafloor environments, but the metabolic potential of the phylum is poorly understood. The phylumChloroflexiis represented by isolates with diverse metabolic strategies, including anoxic phototrophy, fermentation, and reductive dehalogenation; therefore, function cannot be attributed to these organisms based solely on phylogeny. Single-cell genomics can provide metabolic insights into uncultured organisms, like the deep-subsurfaceChloroflexi. Nine SSU rRNA gene sequences were identified from single-cell sorts of whole-round core material collected from the Okinawa Trough at Iheya North hydrothermal field as part of Integrated Ocean Drilling Program (IODP) expedition 331 (Deep Hot Biosphere). Previous studies of subsurfaceChloroflexisingle amplified genomes (SAGs) suggested heterotrophic or lithotrophic metabolisms and provided no evidence for growth by reductive dehalogenation. Our nineChloroflexiSAGs (seven of which are from the orderAnaerolineales) indicate that, in addition to genes for the Wood-Ljungdahl pathway, exogenous carbon sources can be actively transported into cells. At least one subunit for pyruvate ferredoxin oxidoreductase was found in four of theChloroflexiSAGs. This protein can provide a link between the Wood-Ljungdahl pathway and other carbon anabolic pathways. Finally, one of the sevenAnaerolinealesSAGs contains a distinct reductive dehalogenase homologous (rdhA) gene.IMPORTANCEThrough the use of single amplified genomes (SAGs), we have extended the metabolic potential of an understudied group of subsurface microbes, theChloroflexi. These microbes are frequently detected in the subsurface biosphere, though their metabolic capabilities have remained elusive. In contrast to previously examinedChloroflexiSAGs, our genomes (several are from the orderAnaerolineales) were recovered from a hydrothermally driven system and therefore provide a unique window into the metabolic potential of this type of habitat. In addition, a reductive dehalogenase gene (rdhA) has been directly linked to marine subsurfaceChloroflexi, suggesting that reductive dehalogenation is not limited to the classDehalococcoidia. This discovery expands the nutrient-cycling and metabolic potential present within the deep subsurface and provides functional gene information relating to this enigmatic group.

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Reductive Dehalogenation of Brominated Phenolic Compounds by Microorganisms Associated with the Marine Sponge Aplysina aerophoba

Applied and Environmental Microbiology ◽

10.1128/aem.69.7.4159-4166.2003 ◽

2003 ◽

Vol 69 (7) ◽

pp. 4159-4166 ◽

Cited By ~ 92

Author(s):

Young-Beom Ahn ◽

Sung-Keun Rhee ◽

Donna E. Fennell ◽

Lee J. Kerkhof ◽

Ute Hentschel ◽

...

Keyword(s):

Marine Sponge ◽

Denaturing Gradient Gel Electrophoresis ◽

Dry Weight ◽

Marine Sponges ◽

Reductive Dehalogenation ◽

Reductive Dehalogenase ◽

Anaerobic Microorganisms ◽

Large Numbers ◽

Gradient Gel Electrophoresis ◽

Aplysina Aerophoba

ABSTRACT Marine sponges are natural sources of brominated organic compounds, including bromoindoles, bromophenols, and bromopyrroles, that may comprise up to 12% of the sponge dry weight. Aplysina aerophoba sponges harbor large numbers of bacteria that can amount to 40% of the biomass of the animal. We postulated that there might be mechanisms for microbially mediated degradation of these halogenated chemicals within the sponges. The capability of anaerobic microorganisms associated with the marine sponge to transform haloaromatic compounds was tested under different electron-accepting conditions (i.e., denitrifying, sulfidogenic, and methanogenic). We observed dehalogenation activity of sponge-associated microorganisms with various haloaromatics. 2-Bromo-, 3-bromo-, 4-bromo-, 2,6-dibromo-, and 2,4,6-tribromophenol, and 3,5-dibromo-4-hydroxybenzoate were reductively debrominated under methanogenic and sulfidogenic conditions with no activity observed in the presence of nitrate. Monochlorinated phenols were not transformed over a period of 1 year. Debromination of 2,4,6-tribromophenol, and 2,6-dibromophenol to 2-bromophenol was more rapid than the debromination of the monobrominated phenols. Ampicillin and chloramphenicol inhibited activity, suggesting that dehalogenation was mediated by bacteria. Characterization of the debrominating methanogenic consortia by using terminal restriction fragment length polymorphism (TRFLP) and denaturing gradient gel electrophoresis analysis indicated that different 16S ribosomal DNA (rDNA) phylotypes were enriched on the different halogenated substrates. Sponge-associated microorganisms enriched on organobromine compounds had distinct 16S rDNA TRFLP patterns and were most closely related to the δ subgroup of the proteobacteria. The presence of homologous reductive dehalogenase gene motifs in the sponge-associated microorganisms suggested that reductive dehalogenation might be coupled to dehalorespiration.

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Adaptive Responses ofShewanella decolorationisto Toxic Organic Extracellular Electron Acceptor Azo Dyes in Anaerobic Respiration

Applied and Environmental Microbiology ◽

10.1128/aem.00550-19 ◽

2019 ◽

Vol 85 (16) ◽

Cited By ~ 2

Author(s):

Yun Fang ◽

Jun Liu ◽

Guannan Kong ◽

Xueduan Liu ◽

Yonggang Yang ◽

...

Keyword(s):

Gene Expression ◽

Electron Acceptor ◽

Organic Pollutant ◽

Anaerobic Respiration ◽

Energy Resource ◽

Site Directed Mutagenesis ◽

Adaptive Responses ◽

Material Transport ◽

Content Type ◽

Toxic Organics

ABSTRACTBacterial anaerobic respiration using an extracellular electron acceptor plays a predominant role in global biogeochemical cycles. However, the mechanisms of bacterial adaptation to the toxic organic pollutant as the extracellular electron acceptor during anaerobic respiration are not clear, which limits our ability to optimize the strategies for the bioremediation of a contaminated environment. Here, we report the physiological characteristics and the global gene expression of an ecologically successful bacterium,Shewanella decolorationisS12, when using a typical toxic organic pollutant, amaranth, as the extracellular electron acceptor. Our results revealed that filamentous shift (the cells stretched to fiber-like shapes as long as 18 μm) occurred under amaranth stress. Persistent stress led to a higher filamentous cell rate and decolorization ability in subcultural cells compared to parental strains. In addition, the expression of genes involved in cell division, the chemotaxis system, energy conservation, damage repair, and material transport in filamentous cells was significantly stimulated. The detailed roles of some genes with significantly elevated expressions in filamentous cells, such as the outer membrane porin genesompAandompW, the cytochromecgenesarpCandarpD, the global regulatory factor generpoS, and the methyl-accepting chemotaxis proteins genesSHD_2793andSHD_0015, were identified by site-directed mutagenesis. Finally, a conceptual model was proposed to help deepen our insights into both the bacterial survival strategy when toxic organics were present and the mechanisms by which these toxic organics were biodegraded as the extracellular electron acceptors.IMPORTANCEKeeping toxic organic pollutants (TOPs) in tolerable levels is a huge challenge for bacteria in extremely unfavorable environments since TOPs could serve as energy substitutes but also as survival stresses when they are beyond some thresholds. This study focused on the underlying adaptive mechanisms of ecologically successful bacteriumShewanella decolorationisS12 when exposed to amaranth, a typical toxic organic pollutant, as the extracellular electron acceptor. Our results suggest that filamentous shift is a flexible and valid way to solve the dilemma between the energy resource and toxic stress. Filamentous cells regulate gene expression to enhance their degradation and detoxification capabilities, resulting in a strong viability. These novel adaptive responses to TOPs are believed to be an evolutionary achievement to succeed in harsh habitats and thus have great potential to be applied to environment engineering or synthetic biology if we could picture every unknown node in this pathway.

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

Applied and Environmental Microbiology ◽

10.1128/aem.01615-16 ◽

2016 ◽

Vol 82 (17) ◽

pp. 5428-5443 ◽

Cited By ~ 23

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.

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