Phenazines and Other Redox-Active Antibiotics Promote Microbial Mineral Reduction

ABSTRACT Natural products with important therapeutic properties are known to be produced by a variety of soil bacteria, yet the ecological function of these compounds is not well understood. Here we show that phenazines and other redox-active antibiotics can promote microbial mineral reduction. Pseudomonas chlororaphis PCL1391, a root isolate that produces phenazine-1-carboxamide (PCN), is able to reductively dissolve poorly crystalline iron and manganese oxides, whereas a strain carrying a mutation in one of the phenazine-biosynthetic genes (phzB) is not; the addition of purified PCN restores this ability to the mutant strain. The small amount of PCN produced relative to the large amount of ferric iron reduced in cultures of P. chlororaphis implies that PCN is recycled multiple times; moreover, poorly crystalline iron (hydr)oxide can be reduced abiotically by reduced PCN. This ability suggests that PCN functions as an electron shuttle rather than an iron chelator, a finding that is consistent with the observation that dissolved ferric iron is undetectable in culture fluids. Multiple phenazines and the glycopeptidic antibiotic bleomycin can also stimulate mineral reduction by the dissimilatory iron-reducing bacterium Shewanella oneidensis MR1. Because diverse bacterial strains that cannot grow on iron can reduce phenazines, and because thermodynamic calculations suggest that phenazines have lower redox potentials than those of poorly crystalline iron (hydr)oxides in a range of relevant environmental pH (5 to 9), we suggest that natural products like phenazines may promote microbial mineral reduction in the environment.

[1]  T. Beveridge,et al.  Bacterial Recognition of Mineral Surfaces: Nanoscale Interactions Between Shewanella and α-FeOOH , 2001, Science.

[2]  M. Winson,et al.  Multiple homologues of LuxR and LuxI control expression of virulence determinants and secondary metabolites through quorum sensing in Pseudomonas aeruginosa PAO1 , 1995, Molecular microbiology.

[3]  K. Nealson,et al.  Bacterial Manganese Reduction and Growth with Manganese Oxide as the Sole Electron Acceptor , 1988, Science.

[4]  R. D. Demoss,et al.  ON THE BIOSYNTHESIS OF PYOCYANINE , 1959, Journal of bacteriology.

[5]  U. Schwertmann,et al.  Iron Oxides , 2003, SSSA Book Series.

[6]  S. Hecht Bleomycin: new perspectives on the mechanism of action. , 2000, Journal of natural products.

[7]  N. A. Seveno,et al.  Growth of Pseudomonas aureofaciens PGS12 and the dynamics of HHL and phenazine production in liquid culture, on nutrient agar, and on plant roots , 2001, Microbial Ecology.

[8]  J. M. Meyer,et al.  The Fluorescent Pigment of Pseudomonas fluorescens : Biosynthesis, Purification and Physicochemical Properties , 1978 .

[9]  E. Friedheim PYOCYANINE, AN ACCESSORY RESPIRATORY ENZYME , 1931, The Journal of experimental medicine.

[10]  Janet G. Hering,et al.  Principles and Applications of Aquatic Chemistry , 1993 .

[11]  L. Thomashow,et al.  Production of the Antibiotic Phenazine-1-Carboxylic Acid by Fluorescent Pseudomonas Species in the Rhizosphere of Wheat , 1990, Applied and environmental microbiology.

[12]  J. Ottow Evaluation of iron-reducing bacteria in soil and the physiological mechanism of iron-reduction in Aerobacter aerogenes. , 1968, Zeitschrift fur allgemeine Mikrobiologie.

[13]  D. Lovley Environmental Microbe-Metal Interactions , 2000 .

[14]  C. Myers,et al.  Isolation and sequence of omcA, a gene encoding a decaheme outer membrane cytochrome c of Shewanella putrefaciens MR-1, and detection of omcA homologs in other strains of S. putrefaciens. , 1998, Biochimica et biophysica acta.

[15]  M. Mazzola,et al.  Contribution of phenazine antibiotic biosynthesis to the ecological competence of fluorescent pseudomonads in soil habitats , 1992, Applied and environmental microbiology.

[16]  J. M. Turner,et al.  Occurrence, biochemistry and physiology of phenazine pigment production. , 1986, Advances in microbial physiology.

[17]  K. Straub,et al.  The use of biologically produced ferrihydrite for the isolation of novel iron-reducing bacteria. , 1998, Systematic and applied microbiology.

[18]  D. Lovley,et al.  Availability of Ferric Iron for Microbial Reduction in Bottom Sediments of the Freshwater Tidal Potomac River , 1986, Applied and environmental microbiology.

[19]  Derek R. Lovley,et al.  Geobacter metallireducens accesses insoluble Fe(iii) oxide by chemotaxis , 2002, Nature.

[20]  Dianne K. Newman,et al.  A role for excreted quinones in extracellular electron transfer , 2000, Nature.

[21]  S. Bromfield THE REDUCTION OF IRON OXIDE BY BACTERIA , 1954 .

[22]  H. Korth Einfluß von Eisen und Sauerstoff auf die Pigmentbildung bei verschiedenen Pseudomonas-Spezies , 2004, Archiv für Mikrobiologie.

[23]  P. Maurice,et al.  Siderophore Production and Iron Reduction by Pseudomonas mendocina in Response to Iron Deprivation , 2000 .

[24]  J. Thomas-Oates,et al.  Phenazine-1-carboxamide production in the biocontrol strain Pseudomonas chlororaphis PCL1391 is regulated by multiple factors secreted into the growth medium. , 2001, Molecular plant-microbe interactions : MPMI.

[25]  L. Stookey Ferrozine---a new spectrophotometric reagent for iron , 1970 .

[26]  F. Morel,et al.  Microbial Mobilization of Arsenic from Sediments of the Aberjona Watershed , 1997 .

[27]  M. Madigan,et al.  Brock Biology of Microorganisms , 1996 .

[28]  D. Newman,et al.  Extracellular electron transfer , 2001, Cellular and Molecular Life Sciences CMLS.

[29]  D. Lovley,et al.  Novel Mode of Microbial Energy Metabolism: Organic Carbon Oxidation Coupled to Dissimilatory Reduction of Iron or Manganese , 1988, Applied and environmental microbiology.

[30]  D. Lovley,et al.  Humic Substances as a Mediator for Microbially Catalyzed Metal Reduction , 1998 .

[31]  D. Haas,et al.  Oxygen-Sensing Reporter Strain of Pseudomonas fluorescens for Monitoring the Distribution of Low-Oxygen Habitats in Soil , 1999, Applied and Environmental Microbiology.

[32]  T. Chin-A-Woeng,et al.  Phenazines and their role in biocontrol by Pseudomonas bacteria. , 2003, The New phytologist.

[33]  Kelly P. Nevin,et al.  Mechanisms for Fe(III) Oxide Reduction in Sedimentary Environments , 2002 .

[34]  V. M. Young,et al.  A new method of preparation of pyocyanin and demonstration of an unusual bacterial sensitivity. , 1979, Analytical biochemistry.

[35]  Kelly P. Nevin,et al.  Dissimilatory Fe(III) and Mn(IV) reduction. , 1991, Advances in microbial physiology.

[36]  S. Baron,et al.  Antibiotic action of pyocyanin , 1981, Antimicrobial Agents and Chemotherapy.

[37]  Lovley DerekR. Organic matter mineralization with the reduction of ferric iron: A review , 1987 .

[38]  Charles M. Moore,et al.  Dissimilatory Fe(III) and Mn(IV) Reduction by Shewanella putrefaciens Requires ferE, a Homolog of the pulE (gspE) Type II Protein Secretion Gene , 2002, Journal of bacteriology.

[39]  L. Birkofer Konstitution von Dihydrophenazin‐Derivaten , 1952 .

[40]  C. Leang,et al.  OmcB, a c-Type Polyheme Cytochrome, Involved in Fe(III) Reduction in Geobacter sulfurreducens , 2003, Journal of bacteriology.

[41]  P. Bakker,et al.  Biocontrol by Phenazine-1-carboxamide-Producing Pseudomonas chlororaphis PCL1391 of Tomato Root Rot Caused by Fusarium oxysporum f. sp. radicis-lycopersici , 1998 .

[42]  L. Hersman The Role of Siderophores in Iron Oxide Dissolution , 2000 .

[43]  S. Vartivarian,et al.  Extracellular iron reductases: identification of a new class of enzymes by siderophore-producing microorganisms. , 1999, Archives of biochemistry and biophysics.

[44]  A. Beliaev,et al.  Shewanella putrefaciens mtrB Encodes an Outer Membrane Protein Required for Fe(III) and Mn(IV) Reduction , 1998, Journal of bacteriology.

[45]  U. Deppenmeier,et al.  Isolation and Characterization of Methanophenazine and Function of Phenazines in Membrane-Bound Electron Transport ofMethanosarcina mazei Gö1 , 1998, Journal of bacteriology.

[46]  F. J. Stevenson HUmus Chemistry Genesis, Composition, Reactions , 1982 .

[47]  L. Pierson,et al.  Phenazine antibiotic production in Pseudomonas aureofaciens: role in rhizosphere ecology and pathogen suppression , 1996 .

[48]  D. Lovley,et al.  Organic Matter Mineralization with Reduction of Ferric Iron in Anaerobic Sediments , 1986, Applied and environmental microbiology.

[49]  King Eo,et al.  Two simple media for the demonstration of pyocyanin and fluorescin. , 1954 .

[50]  D. Wood,et al.  Phenazine antibiotic biosynthesis in Pseudomonas aureofaciens 30-84 is regulated by PhzR in response to cell density , 1994, Journal of bacteriology.

[51]  C. D. Cox Role of pyocyanin in the acquisition of iron from transferrin , 1986, Infection and immunity.

[52]  A. McLoughlin,et al.  The regulation of pyocyanin production in Pseudomonas aeruginosa , 1982, European journal of applied microbiology and biotechnology.

[53]  I. Fridovich,et al.  Mechanism of the antibiotic action pyocyanine , 1980, Journal of bacteriology.