1887

Abstract

is an opportunistic pathogen that forms biofilms on mucous plugs in the lungs of cystic fibrosis (CF) patients, resulting in chronic infections. Pulmonary isolates often display a mucoid (alginate-producing) phenotype, whereas non-mucoid strains are generally associated with acute infections. We characterized the cytosolic proteomes of biofilm-associated and planktonic forms of a CF pulmonary isolate, FRD1, and a non-mucoid strain, PAO1. Since Ca metabolism is altered in CF pulmonary fluids, we also analysed the effect of Ca on the proteome responses of these strains. Both strains altered the abundances of 40–60 % of their proteins in response to biofilm growth and/or [Ca]. Differentially expressed proteins clustered into 12 groups, based on their abundance profiles. From these clusters, 146 proteins were identified by using MALDI-TOF/TOF mass spectrometry. Similarities as well as strain-specific differences were observed. Both strains altered the production of proteins involved in iron acquisition, pyocyanin biosynthesis, quinolone signalling and nitrogen metabolism, proteases, and proteins involved in oxidative and general stress responses. Individual proteins from these classes were highly represented in the biofilm proteomes of both strains. Strain-specific differences concerned the proteins within these functional groups, particularly for enzymes involved in iron acquisition and polysaccharide metabolism, and proteases. The results demonstrate that a mucoid CF isolate of responds to biofilm-associated growth and [Ca] in a fashion similar to strain PAO1, but that strain-specific differences may allow this CF isolate to successfully colonize the pulmonary environment.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.2007/010371-0
2007-11-01
2024-03-28
Loading full text...

Full text loading...

/deliver/fulltext/micro/153/11/3838.html?itemId=/content/journal/micro/10.1099/mic.0.2007/010371-0&mimeType=html&fmt=ahah

References

  1. Alonso-Casajus N., Dauvillee D., Viale A. M., Munoz F. J., Baroja-Fernandez E., Moran-Zorzano M. T., Eydallin G., Ball S., Pozueta-Romero J. 2006; Glycogen phosphorylase, the product of the glgP gene, catalyzes glycogen breakdown by removing glucose units from the nonreducing ends in Escherichia coli. J Bacteriol 188:5266–5272
    [Google Scholar]
  2. Banin E., Vasil M. L., Greenberg E. P. 2005; Iron and Pseudomonas aeruginosa biofilm formation. Proc Natl Acad Sci U S A 102:11076–11081
    [Google Scholar]
  3. Belvisi M., Barnes P. J., Larkin S., Yacoub M., Tadjkarimi S., Williams T. J., Mitchell J. A. 1995; Nitric oxide synthase activity is elevated in inflammatory lung disease in humans. Eur J Pharmacol 283:255–258
    [Google Scholar]
  4. Boles B. R., Thoendel M., Singh P. K. 2004; Self-generated diversity produces “insurance effects” in biofilm communities. Proc Natl Acad Sci U S A 101:16630–16635
    [Google Scholar]
  5. Bredenbruch F., Geffers R., Nimtz M., Buer J., Haussler S. 2006; The Pseudomonas aeruginosa quinolone signal (PQS) has an iron-chelating activity. Environ Microbiol 8:1318–1329
    [Google Scholar]
  6. Brint J. M., Ohman D. E. 1995; Synthesis of multiple exoproducts in Pseudomonas aeruginosa is under the control of RhlR–RhlI, another set of regulators in strain PAO1 with homology to the autoinducer-responsive LuxR–LuxI family. J Bacteriol 177:7155–7163
    [Google Scholar]
  7. Britigan B. E., Roeder T. L., Rasmussen G. T., Shasby D. M., McCormick M. L., Cox C. D. 1992; Interaction of the Pseudomonas aeruginosa secretory products pyocyanin and pyochelin generates hydroxyl radical and causes synergistic damage to endothelial cells. Implications for Pseudomonas-associated tissue injury. J Clin Invest 90:2187–2196
    [Google Scholar]
  8. Britigan B. E., Rasmussen G. T., Cox C. D. 1994; Pseudomonas siderophore pyochelin enhances neutrophil-mediated endothelial cell injury. Am J Physiol 266:L192–L198
    [Google Scholar]
  9. Costerton J. W., Lewandowski Z., Caldwell D. E., Korber D. R., Lappin-Scott H. M. 1995; Microbial biofilms. Annu Rev Microbiol 49:711–745
    [Google Scholar]
  10. Costerton J. W., Stewart P. S., Greenberg E. P. 1999; Bacterial biofilms: a common cause of persistent infections. Science 284:1318–1322
    [Google Scholar]
  11. Davies D. G., Parsek M. R., Pearson J. P., Iglewski B. H., Costerton J. W., Greenberg E. P. 1998; The involvement of cell-to-cell signals in the development of a bacterial biofilm. Science 280:295–298
    [Google Scholar]
  12. Denning G. M., Railsback M. A., Rasmussen G. T., Cox C. D., Britigan B. E. 1998; Pseudomonas pyocyanine alters calcium signaling in human airway epithelial cells. Am J Physiol 274:L893–L900
    [Google Scholar]
  13. Egan M. E., Glockner-Pagel J., Ambrose C., Cahill P. A., Pappoe L., Balamuth N., Cho E., Canny S., Wagner C. A. other authors 2002; Calcium-pump inhibitors induce functional surface expression of ΔF508-CFTR protein in cystic fibrosis epithelial cells. Nat Med 8:485–492
    [Google Scholar]
  14. Elkins J. G., Hassett D. J., Stewart P. S., Schweizer H. P., McDermott T. R. 1999; Protective role of catalase in Pseudomonas aeruginosa biofilm resistance to hydrogen peroxide. Appl Environ Microbiol 65:4594–4600
    [Google Scholar]
  15. Firoved A. M., Deretic V. 2003; Microarray analysis of global gene expression in mucoid Pseudomonas aeruginosa. J Bacteriol 185:1071–1081
    [Google Scholar]
  16. Friedman L., Kolter R. 2004; Two genetic loci produce distinct carbohydrate-rich structural components of the Pseudomonas aeruginosa biofilm matrix. J Bacteriol 186:4457–4465
    [Google Scholar]
  17. Halmerbauer G., Arri S., Schierl M., Strauch E., Koller D. Y. 2000; The relationship of eosinophil granule proteins to ions in the sputum of patients with cystic fibrosis. Clin Exp Allergy 30:1771–1776
    [Google Scholar]
  18. Hanna S. L., Sherman N. E., Kinter M. T., Goldberg J. B. 2000; Comparison of proteins expressed by Pseudomonas aeruginosa strains representing initial and chronic isolates from a cystic fibrosis patient: an analysis by 2-D gel electrophoresis and capillary column liquid chromatography-tandem mass spectrometry. Microbiology 146:2495–2508
    [Google Scholar]
  19. Hassett D. J., Schweizer H. P., Ohman D. E. 1995; Pseudomonas aeruginosa sodA and sodB mutants defective in manganese- and iron-cofactored superoxide dismutase activity demonstrate the importance of the iron-cofactored form in aerobic metabolism. J Bacteriol 177:6330–6337
    [Google Scholar]
  20. Hassett D. J., Sokol P. A., Howell M. L., Ma J. F., Schweizer H. T., Ochsner U., Vasil M. L. 1996; Ferric uptake regulator (Fur) mutants of Pseudomonas aeruginosa demonstrate defective siderophore-mediated iron uptake, altered aerobic growth, and decreased superoxide dismutase and catalase activities. J Bacteriol 178:3996–4003
    [Google Scholar]
  21. Hassett D. J., Ma J. F., Elkins J. G., McDermott T. R., Ochsner U. A., West S. E., Huang C. T., Fredericks J., Burnett S. other authors 1999; Quorum sensing in Pseudomonas aeruginosa controls expression of catalase and superoxide dismutase genes and mediates biofilm susceptibility to hydrogen peroxide. Mol Microbiol 34:1082–1093
    [Google Scholar]
  22. Hentzer M., Teitzel G. M., Balzer G. J., Heydorn A., Molin S., Givskov M., Parsek M. R. 2001; Alginate overproduction affects Pseudomonas aeruginosa biofilm structure and function. J Bacteriol 183:5395–5401
    [Google Scholar]
  23. Jackson K. D., Starkey M., Kremer S., Parsek M. R., Wozniak D. J. 2004; Identification of psl, a locus encoding a potential exopolysaccharide that is essential for Pseudomonas aeruginosa PAO1 biofilm formation. J Bacteriol 186:4466–4475
    [Google Scholar]
  24. Jensen P. O., Bjarnsholt T., Phipps R., Rasmussen T. B., Calum H., Christoffersen L., Moser C., Williams P., Pressler T. other authors 2007; Rapid necrotic killing of polymorphonuclear leukocytes is caused by quorum-sensing-controlled production of rhamnolipid by Pseudomonas aeruginosa. Microbiology 153:1329–1338
    [Google Scholar]
  25. Jesaitis A. J., Franklin M. J., Berglund D., Sasaki M., Lord C. I., Bleazard J. B., Duffy J. E., Beyenal H., Lewandowski Z. 2003; Compromised host defense on Pseudomonas aeruginosa biofilms: characterization of neutrophil and biofilm interactions. J Immunol 171:4329–4339
    [Google Scholar]
  26. Keren I., Shah D., Spoering A., Kaldalu N., Lewis K. 2004; Specialized persister cells and the mechanism of multidrug tolerance in Escherichia coli. J Bacteriol 186:8172–8180
    [Google Scholar]
  27. Lau G. W., Ran H., Kong F., Hassett D. J., Mavrodi D. 2004; Pseudomonas aeruginosa pyocyanin is critical for lung infection in mice. Infect Immun 72:4275–4278
    [Google Scholar]
  28. Lewis K. 2007; Persister cells, dormancy and infectious disease. Nat Rev Microbiol 5:48–56
    [Google Scholar]
  29. Liu L., Ridefelt P., Hakansson L., Venge P. 1999; Regulation of human eosinophil migration across lung epithelial monolayers by distinct calcium signaling mechanisms in the two cell types. J Immunol 163:5649–5655
    [Google Scholar]
  30. Lorin M. I., Gaerlan P. F., Mandel I. D., Denning C. R. 1976; Composition of nasal secretion in patients with cystic fibrosis. J Lab Clin Med 88:114–117
    [Google Scholar]
  31. Lyczak J. B., Cannon C. L., Pier G. B. 2000; Establishment of Pseudomonas aeruginosa infection: lessons from a versatile opportunist. Microbes Infect 2:1051–1060
    [Google Scholar]
  32. Lyczak J. B., Cannon C. L., Pier G. B. 2002; Lung infections associated with cystic fibrosis. Clin Microbiol Rev 15:194–222
    [Google Scholar]
  33. Mah T. F., Pitts B., Pellock B., Walker G. C., Stewart P. S., O'Toole G. A. 2003; A genetic basis for Pseudomonas aeruginosa biofilm antibiotic resistance. Nature 426:306–310
    [Google Scholar]
  34. Mai G. T., Seow W. K., Pier G. B., McCormack J. G., Thong Y. H. 1993; Suppression of lymphocyte and neutrophil functions by Pseudomonas aeruginosa mucoid exopolysaccharide (alginate): reversal by physicochemical, alginase, and specific monoclonal antibody treatments. Infect Immun 61:559–564
    [Google Scholar]
  35. Malhotra S., Silo-Suh L. A., Mathee K., Ohman D. E. 2000; Proteome analysis of the effect of mucoid conversion on global protein expression in Pseudomonas aeruginosa strain PAO1 shows induction of the disulfide bond isomerase, dsbA. J Bacteriol 182:6999–7006
    [Google Scholar]
  36. Marquart M. E., Dajcs J. J., Caballero A. R., Thibodeaux B. A., O'Callaghan R. J. 2005; Calcium and magnesium enhance the production of Pseudomonas aeruginosa protease IV, a corneal virulence factor. Med Microbiol Immunol 194:39–45
    [Google Scholar]
  37. McKnight S. L., Iglewski B. H., Pesci E. C. 2000; The Pseudomonas quinolone signal regulates rhl quorum sensing in Pseudomonas aeruginosa. J Bacteriol 182:2702–2708
    [Google Scholar]
  38. Musk D. J., Banko D. A., Hergenrother P. J. 2005; Iron salts perturb biofilm formation and disrupt existing biofilms of Pseudomonas aeruginosa. Chem Biol 12:789–796
    [Google Scholar]
  39. Nivens D. E., Ohman D. E., Williams J., Franklin M. J. 2001; Role of alginate and its O acetylation in formation of Pseudomonas aeruginosa microcolonies and biofilms. J Bacteriol 183:1047–1057
    [Google Scholar]
  40. Ochsner U. A., Vasil M. L. 1996; Gene repression by the ferric uptake regulator in Pseudomonas aeruginosa: cycle selection of iron-regulated genes. Proc Natl Acad Sci U S A 93:4409–4414
    [Google Scholar]
  41. Ochsner U. A., Vasil A. I., Vasil M. L. 1995; Role of the ferric uptake regulator of Pseudomonas aeruginosa in the regulation of siderophores and exotoxin A expression: purification and activity on iron-regulated promoters. J Bacteriol 177:7194–7201
    [Google Scholar]
  42. Ohman D. E., Chakrabarty A. M. 1981; Genetic mapping of chromosomal determinants for the production of the exopolysaccharide alginate in a Pseudomonas aeruginosa cystic fibrosis isolate. Infect Immun 33:142–148
    [Google Scholar]
  43. Ojha A., Anand M., Bhatt A., Kremer L., Jacobs W. R. Jr, Hatfull G. F. 2005; GroEL1: a dedicated chaperone involved in mycolic acid biosynthesis during biofilm formation in mycobacteria. Cell 123:861–873
    [Google Scholar]
  44. Ojoo J. C., Mulrennan S. A., Kastelik J. A., Morice A. H., Redington A. E. 2005; Exhaled breath condensate pH and exhaled nitric oxide in allergic asthma and in cystic fibrosis. Thorax 60:22–26
    [Google Scholar]
  45. Olson J. C., Ohman D. E. 1992; Efficient production and processing of elastase and LasA by Pseudomonas aeruginosa require zinc and calcium ions. J Bacteriol 174:4140–4147
    [Google Scholar]
  46. Pier G. B., Coleman F., Grout M., Franklin M., Ohman D. E. 2001; Role of alginate O acetylation in resistance of mucoid Pseudomonas aeruginosa to opsonic phagocytosis. Infect Immun 69:1895–1901
    [Google Scholar]
  47. Rani S. A., Pitts B., Stewart P. S. 2005; Rapid diffusion of fluorescent tracers into Staphylococcus epidermidis biofilms visualized by time lapse microscopy. Antimicrob Agents Chemother 49:728–732
    [Google Scholar]
  48. Redly G. A., Poole K. 2003; Pyoverdine-mediated regulation of FpvA synthesis in Pseudomonas aeruginosa: involvement of a probable extracytoplasmic-function sigma factor, FpvI. J Bacteriol 185:1261–1265
    [Google Scholar]
  49. Roberts M. E., Stewart P. S. 2004; Modeling antibiotic tolerance in biofilms by accounting for nutrient limitation. Antimicrob Agents Chemother 48:48–52
    [Google Scholar]
  50. Sarkisova S., Patrauchan M. A., Berglund D., Nivens D. E., Franklin M. J. 2005; Calcium-induced virulence factors associated with the extracellular matrix of mucoid Pseudomonas aeruginosa biofilms. J Bacteriol 187:4327–4337
    [Google Scholar]
  51. Sauer K., Camper A. K., Ehrlich G. D., Costerton J. W., Davies D. G. 2002; Pseudomonas aeruginosa displays multiple phenotypes during development as a biofilm. J Bacteriol 184:1140–1154
    [Google Scholar]
  52. Schuster M., Lostroh C. P., Ogi T., Greenberg E. P. 2003; Identification, timing, and signal specificity of Pseudomonas aeruginosa quorum-controlled genes: a transcriptome analysis. J Bacteriol 185:2066–2079
    [Google Scholar]
  53. Southey-Pillig C. J., Davies D. G., Sauer K. 2005; Characterization of temporal protein production in Pseudomonas aeruginosa biofilms. J Bacteriol 187:8114–8126
    [Google Scholar]
  54. Stewart P. S., Costerton J. W. 2001; Antibiotic resistance of bacteria in biofilms. Lancet 358:135–138
    [Google Scholar]
  55. Stover C. K., Pham X. Q., Erwin A. L., Mizoguchi S. D., Warrener P., Hickey M. J., Brinkman F. S., Hufnagle W. O., Kowalik D. J. other authors 2000; Complete genome sequence of Pseudomonas aeruginosa PA01, an opportunistic pathogen. Nature 406:959–964
    [Google Scholar]
  56. Vasil M. L., Ochsner U. A., Johnson Z., Colmer J. A., Hamood A. N. 1998; The fur-regulated gene encoding the alternative sigma factor PvdS is required for iron-dependent expression of the LysR-type regulator ptxR in Pseudomonas aeruginosa. J Bacteriol 180:6784–6788
    [Google Scholar]
  57. Viebrock A., Zumft W. G. 1988; Molecular cloning, heterologous expression, and primary structure of the structural gene for the copper enzyme nitrous oxide reductase from denitrifying Pseudomonas stutzeri. J Bacteriol 170:4658–4668
    [Google Scholar]
  58. von Ruecker A. A., Bertele R., Harms H. K. 1984; Calcium metabolism and cystic fibrosis: mitochondrial abnormalities suggest a modification of the mitochondrial membrane. Pediatr Res 18:594–599
    [Google Scholar]
  59. Wagner V. E., Bushnell D., Passador L., Brooks A. I., Iglewski B. H. 2003; Microarray analysis of Pseudomonas aeruginosa quorum-sensing regulons: effects of growth phase and environment. J Bacteriol 185:2080–2095
    [Google Scholar]
  60. Waite R. D., Papakonstantinopoulou A., Littler E., Curtis M. A. 2005; Transcriptome analysis of Pseudomonas aeruginosa growth: comparison of gene expression in planktonic cultures and developing and mature biofilms. J Bacteriol 187:6571–6576
    [Google Scholar]
  61. Waite R. D., Paccanaro A., Papakonstantinopoulou A., Hurst J. M., Saqi M., Littler E., Curtis M. A. 2006; Clustering of Pseudomonas aeruginosa transcriptomes from planktonic cultures, developing and mature biofilms reveals distinct expression profiles. BMC Genomics 7:162
    [Google Scholar]
  62. Walters M. C. III, Roe F., Bugnicourt A., Franklin M. J., Stewart P. S. 2003; Contributions of antibiotic penetration, oxygen limitation, and low metabolic activity to tolerance of Pseudomonas aeruginosa biofilms to ciprofloxacin and tobramycin. Antimicrob Agents Chemother 47:317–323
    [Google Scholar]
  63. Whiteley M., Bangera M. G., Bumgarner R. E., Parsek M. R., Teitzel G. M., Lory S., Greenberg E. P. 2001; Gene expression in Pseudomonas aeruginosa biofilms. Nature 413:860–864
    [Google Scholar]
  64. Wilderman P. J., Vasil A. I., Johnson Z., Wilson M. J., Cunliffe H. E., Lamont I. L., Vasil M. L. 2001; Characterization of an endoprotease (PrpL) encoded by a PvdS-regulated gene in Pseudomonas aeruginosa. Infect Immun 69:5385–5394
    [Google Scholar]
  65. Wozniak D. J., Wyckoff T. J., Starkey M., Keyser R., Azadi P., O'Toole G. A., Parsek M. R. 2003; Alginate is not a significant component of the extracellular polysaccharide matrix of PA14 and PAO1 Pseudomonas aeruginosa biofilms. Proc Natl Acad Sci U S A 100:7907–7912
    [Google Scholar]
  66. Yahr T. L., Mende-Mueller L. M., Friese M. B., Frank D. W. 1997; Identification of type III secreted products of the Pseudomonas aeruginosa exoenzyme S regulon. J Bacteriol 179:7165–7168
    [Google Scholar]
  67. Yoon S. S., Hennigan R. F., Hilliard G. M., Ochsner U. A., Parvatiyar K., Kamani M. C., Allen H. L., DeKievit T. R., Gardner P. R. other authors 2002; Pseudomonas aeruginosa anaerobic respiration in biofilms: relationships to cystic fibrosis pathogenesis. Dev Cell 3:593–603
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.2007/010371-0
Loading
/content/journal/micro/10.1099/mic.0.2007/010371-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error