1887

Abstract

Transmissible clones potentially pose a serious threat to cystic fibrosis (CF) patients. The AES-1 clone has been found to infect up to 40 % of patients in five CF centres in eastern Australia. Studies were carried out on clonal and non-clonal (NC) isolates from chronically infected CF patients, and the reference strain PAO1, to gain insight into the properties of AES-1. The transcriptomes of AES-1 and NC isolates, and of PAO1, grown planktonically and as a 72 h biofilm were compared using PAO1 microarrays. Microarray data were validated using real-time PCR. Overall, most differentially expressed genes were downregulated. AES-1 differentially expressed bacteriophage genes, novel motility genes, and virulence and quorum-sensing-related genes, compared with both PAO1 and NC. AES-1 but not NC biofilms significantly downregulated aerobic respiration genes compared with planktonic growth, suggesting enhanced anaerobic/microaerophilic growth by AES-1. Biofilm measurement showed that AES-1 formed significantly larger and thicker biofilms than NC or PAO1 isolates. This may be related to expression of the gene PA0729, encoding a biofilm-enhancing bacteriophage, identified by PCR in all AES-1 but few NC isolates (=42). Links with the Liverpool epidemic strain included the presence of PA0729 and the absence of the bacteriophage gene cluster PA0632–PA0639. No common markers were found with the Manchester strain. No particular differentially expressed gene in AES-1 could definitively be ascribed a role in its infectivity, thus increasing the likelihood that AES-1 infectivity is multi-factorial and possibly involves novel genes. This study extends our understanding of the transcriptomic and genetic differences between clonal and NC strains of from CF lung.

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2008-12-01
2024-03-28
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References

  1. Al-Aloul M., Crawley J., Winstanley C., Hart C. A., Ledson M. J., Walshaw M. J. 2004; Increased morbidity associated with chronic infection by an epidemic Pseudomonas aeruginosa strain in CF patients. Thorax 59:334–336 [CrossRef]
    [Google Scholar]
  2. Alvarez-Ortega C., Harwood C. S. 2007; Responses of Pseudomonas aeruginosa to low oxygen indicate that growth in the cystic fibrosis lung is by aerobic respiration. Mol Microbiol 65:153–165 [CrossRef]
    [Google Scholar]
  3. Anthony M., Rose B., Pegler M. B., Elkins M., Service H., Thamotharampillai K., Watson J., Robinson M., Bye P. other authors 2002; Genetic analysis of Pseudomonas aeruginosa isolates from the sputa of Australian adult cystic fibrosis patients. J Clin Microbiol 40:2772–2778 [CrossRef]
    [Google Scholar]
  4. Armstrong D. S., Nixon G. M., Carzino R., Bigham A., Carlin J. B., Robins-Browne R. M., Grimwood K. 2002; Detection of a widespread clone of Pseudomonas aeruginosa in a pediatric cystic fibrosis clinic. Am J Respir Crit Care Med 166:983–987 [CrossRef]
    [Google Scholar]
  5. Armstrong D., Bell S., Robinson M., Bye P., Rose B., Harbour C., Lee C., Service H., Nissen M. other authors 2003; Evidence for spread of a clonal strain of Pseudomonas aeruginosa among cystic fibrosis clinics. J Clin Microbiol 41:2266–2267 [CrossRef]
    [Google Scholar]
  6. Arora S. K., Bangera M., Lory S., Ramphal R. 2001; A genomic island in Pseudomonas aeruginosa carries the determinants of flagellin glycosylation. Proc Natl Acad Sci U S A 98:9342–9347 [CrossRef]
    [Google Scholar]
  7. Ausubel F. M., Brent R., Kingston R. E., Moore D. D., Seidman J. G., Smith J. A., Struhl K. 2003 Current Protocols in Molecular Biology , 3rd edn. New York: John Wiley & Sons;
    [Google Scholar]
  8. Benjamini Y., Hochberg Y. 1995; Controlling the false discovery rate: a practical and powerful approach to multiple testing. J R Stat Soc Ser B Methodological 57:289–300
    [Google Scholar]
  9. Bolstad B. M., Irizarry R. A., Astrand M., Speed T. P. 2003; A comparison of normalization methods for high density oligonucleotide array data based on variance and bias. Bioinformatics 19:185–193 [CrossRef]
    [Google Scholar]
  10. Brockhurst M. A., Buckling A., Rainey P. B. 2005; The effect of a bacteriophage on diversification of the opportunistic bacterial pathogen, Pseudomonas aeruginosa . Proc Biol Sci 272:1385–1391 [CrossRef]
    [Google Scholar]
  11. Buell C. R., Joardar V., Lindeberg M., Selengut J., Paulsen I. T., Gwinn M. L., Dodson R. J., Deboy R. T., Durkin A. S. other authors 2003; The complete genome sequence of the Arabidopsis and tomato pathogen Pseudomonas syringae pv. tomato DC3000. Proc Natl Acad Sci U S A 100:10181–10186 [CrossRef]
    [Google Scholar]
  12. 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 [CrossRef]
    [Google Scholar]
  13. De Kievit T. R., Gillis R., Marx S., Brown C., Iglewski B. H. 2001; Quorum-sensing genes in Pseudomonas aeruginosa biofilms: their role and expression patterns. Appl Environ Microbiol 67:1865–1873 [CrossRef]
    [Google Scholar]
  14. Drenkard E., Ausubel F. M. 2002; Pseudomonas biofilm formation and antibiotic resistance are linked to phenotypic variation. Nature 416:740–743 [CrossRef]
    [Google Scholar]
  15. Dziewit L., Jazurek M., Drewniak L., Baj J., Bartosik D. 2007; The SXT conjugative element and linear prophage N15 encode toxin–antitoxin-stabilizing systems homologous to the tad-ata module of the Paracoccus aminophilus plasmid pAMI2. J Bacteriol 189:1983–1997 [CrossRef]
    [Google Scholar]
  16. Finnan S., Morrissey J. P., O'Gara F., Boyd E. F. 2004; Genome diversity of Pseudomonas aeruginosa isolates from cystic fibrosis patients and the hospital environment. J Clin Microbiol 42:5783–5792 [CrossRef]
    [Google Scholar]
  17. Gautier L., Cope L., Bolstad B. M., Irizarry R. A. 2004; affy – analysis of Affymetrix GeneChip data at the probe level. Bioinformatics 20:307–315 [CrossRef]
    [Google Scholar]
  18. Gentleman R. C., Carey V. J., Bates D. M., Bolstad B., Dettling M., Dudoit S., Ellis B., Gautier L., Ge Y. other authors 2004; Bioconductor: open software development for computational biology and bioinformatics. Genome Biol 5:R80 [CrossRef]
    [Google Scholar]
  19. Head N. E., Yu H. 2004; Cross-sectional analysis of clinical and environmental isolates of Pseudomonas aeruginosa : biofilm formation, virulence, and genome diversity. Infect Immun 72:133–144 [CrossRef]
    [Google Scholar]
  20. Hentzer M., Givskov M., Eberl L. 2004; Quorum sensing in biofilms: gossip in slime city. In Microbial Biofilms pp 118–140 Edited by Ghannoum M., O'Toole G. A. Washington, DC: American Society for Microbiology;
    [Google Scholar]
  21. Hentzer M., Eberl L., Givskov M. 2005; Transcriptome analysis of Pseudomonas aeruginosa biofilm development: anaerobic respiration and iron limitation. Biofilms 2:37–61 [CrossRef]
    [Google Scholar]
  22. Irizarry R. A., Hobbs B., Collin F., Beazer-Barclay Y. D., Antonellis K. J., Scherf U., Speed T. P. 2003; Exploration, normalization, and summaries of high density oligonucleotide array probe level data. Biostatistics 4:249–264 [CrossRef]
    [Google Scholar]
  23. Kim S.-H., Lee K.-B., Lee J.-S., Cho Y.-H. 2003; Genome diversification by phage-derived genomic islands in Pseudomonas aeruginosa . J Microbiol Biotechnol 13:783–788
    [Google Scholar]
  24. Kim S. J., Park R. Y., Kang S. M., Choi M. H., Kim C. M., Shin S. H. 2006; Pseudomonas aeruginosa alkaline protease can facilitate siderophore-mediated iron-uptake via the proteolytic cleavage of transferrins. Biol Pharm Bull 29:2295–2300 [CrossRef]
    [Google Scholar]
  25. Klausen M., Heydorn A., Ragas P., Lambertsen L., Aaes-Jorgensen A., Molin S., Tolker-Nielsen T. 2003; Biofilm formation by Pseudomonas aeruginosa wild type, flagella and type IV pili mutants. Mol Microbiol 48:1511–1524 [CrossRef]
    [Google Scholar]
  26. Klockgether J., Reva O., Larbig K., Tummler B. 2004; Sequence analysis of the mobile genome island pKLC102 of Pseudomonas aeruginosa C. J Bacteriol 186:518–534 [CrossRef]
    [Google Scholar]
  27. Koch C., Hoiby N. 1993; Pathogenesis of cystic fibrosis. Lancet 341:1065–1069 [CrossRef]
    [Google Scholar]
  28. Kosorok M. R., Zeng L., West S. E., Rock M. J., Splaingard M. L., Laxova A., Green C. G., Collins J., Farrell P. M. 2001; Acceleration of lung disease in children with cystic fibrosis after Pseudomonas aeruginosa acquisition. Pediatr Pulmonol 32:277–287 [CrossRef]
    [Google Scholar]
  29. Kukavica-Ibrulj I., Bragonzi A., Paroni M., Winstanley C., Sanschagrin F., O'Toole G. A., Levesque R. C. 2008; In vivo growth of Pseudomonas aeruginosa strains PAO1 and PA14 and the hypervirulent strain LESB58 in a rat model of chronic lung infection. J Bacteriol 190:2804–2813 [CrossRef]
    [Google Scholar]
  30. Lamont I. L., Beare P. A., Ochsner U., Vasil A. I., Vasil M. L. 2002; Siderophore-mediated signaling regulates virulence factor production in Pseudomonas aeruginosa . Proc Natl Acad Sci U S A 99:7072–7077 [CrossRef]
    [Google Scholar]
  31. Lewis D. A., Jones A., Parkhill J., Speert D. P., Govan J. R., Lipuma J. J., Lory S., Webb A. K., Mahenthiralingam E. 2005; Identification of DNA markers for a transmissible Pseudomonas aeruginosa cystic fibrosis strain. Am J Respir Cell Mol Biol 33:56–64 [CrossRef]
    [Google Scholar]
  32. Mack K., Titball R. W. 1998; The detection of insertion sequences within the human pathogen Burkholderia pseudomallei which have been identified previously in Burkholderia cepacia . FEMS Microbiol Lett 162:69–74 [CrossRef]
    [Google Scholar]
  33. Manos J., Artimovich E., Belas R. 2004; Enhanced motility of a Proteus mirabilis strain expressing hybrid FlaAB flagella. Microbiology 150:1291–1299 [CrossRef]
    [Google Scholar]
  34. Mathee K., Narasimhan G., Valdes C., Qiu X., Matewish J. M., Koehrsen M., Rokas A., Yandava C. N., Engels R. other authors 2008; Dynamics of Pseudomonas aeruginosa genome evolution. Proc Natl Acad Sci U S A 105:3100–3105 [CrossRef]
    [Google Scholar]
  35. McCallum S. J., Gallagher M. J., Corkill J. E., Hart C. A., Ledson M. J., Walshaw M. J. 2002; Spread of an epidemic Pseudomonas aeruginosa strain from a patient with cystic fibrosis (CF) to non-CF relatives. Thorax 57:559–560 [CrossRef]
    [Google Scholar]
  36. Nouwens A. S., Beatson S. A., Whitchurch C. B., Walsh B. J., Schweizer H. P., Mattick J. S., Cordwell S. J. 2003; Proteome analysis of extracellular proteins regulated by the las and rhl quorum sensing systems in Pseudomonas aeruginosa PAO1. Microbiology 149:1311–1322 [CrossRef]
    [Google Scholar]
  37. O'Carroll M. R., Syrmis M. W., Wainwright C. E., Greer R. M., Mitchell P., Coulter C., Sloots T. P., Nissen M. D., Bell S. C. 2004; Clonal strains of Pseudomonas aeruginosa in paediatric and adult cystic fibrosis units. Eur Respir J 24:101–106 [CrossRef]
    [Google Scholar]
  38. O'Toole G. A., Kolter R. 1998; Flagellar and twitching motility are necessary for Pseudomonas aeruginosa biofilm development. Mol Microbiol 30:295–304 [CrossRef]
    [Google Scholar]
  39. Ochsner U. A., Wilderman P. J., Vasil A. I., Vasil M. L. 2002; GeneChip expression analysis of the iron starvation response in Pseudomonas aeruginosa : identification of novel pyoverdine biosynthesis genes. Mol Microbiol 45:1277–1287 [CrossRef]
    [Google Scholar]
  40. Panagea S., Winstanley C., Walshaw M. J., Ledson M. J., Hart C. A. 2005; Environmental contamination with an epidemic strain of Pseudomonas aeruginosa in a Liverpool cystic fibrosis centre, and study of its survival on dry surfaces. J Hosp Infect 59:102–107 [CrossRef]
    [Google Scholar]
  41. Pandey D. P., Gerdes K. 2005; Toxin–antitoxin loci are highly abundant in free-living but lost from host-associated prokaryotes. Nucleic Acids Res 33:966–976 [CrossRef]
    [Google Scholar]
  42. Pellegrino F. L., Casali N., Dos Santos K. R., Nouer S. A., Scheidegger E. M., Riley L. W., Moreira B. M. 2006; Pseudomonas aeruginosa epidemic strain carrying bla SPM metallo- β -lactamase detected in Rio de Janeiro. Brazil. J Chemother 18:151–156 [CrossRef]
    [Google Scholar]
  43. Platt M. D., Schurr M. J., Sauer K., Vazquez G., Kukavica-Ibrulj I., Potvin E., Levesque R. C., Fedynak A., Brinkman F. S. other authors 2008; Proteomic, microarray, and signature-tagged mutagenesis analyses of anaerobic Pseudomonas aeruginosa at pH 6.5, likely representing chronic, late-stage cystic fibrosis airway conditions. J Bacteriol 190:2739–2758 [CrossRef]
    [Google Scholar]
  44. Ravin V., Ravin N., Casjens S., Ford M. E., Hatfull G. F., Hendrix R. W. 2000; Genomic sequence and analysis of the atypical temperate bacteriophage N15. J Mol Biol 299:53–73 [CrossRef]
    [Google Scholar]
  45. Romling U., Schmidt K. D., Tummler B. 1997; Large genome rearrangements discovered by the detailed analysis of 21 Pseudomonas aeruginosa clone C isolates found in environment and disease habitats. J Mol Biol 271:386–404 [CrossRef]
    [Google Scholar]
  46. Salunkhe P., Smart C. H., Morgan J. A., Panagea S., Walshaw M. J., Hart C. A., Geffers R., Tummler B., Winstanley C. 2005; A cystic fibrosis epidemic strain of Pseudomonas aeruginosa displays enhanced virulence and antimicrobial resistance. J Bacteriol 187:4908–4920 [CrossRef]
    [Google Scholar]
  47. 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 [CrossRef]
    [Google Scholar]
  48. Scharfman A., Arora S. K., Delmotte P., Van Brussel E., Mazurier J., Ramphal R., Roussel P. 2001; Recognition of Lewis x derivatives present on mucins by flagellar components of Pseudomonas aeruginosa . Infect Immun 69:5243–5248 [CrossRef]
    [Google Scholar]
  49. Schirmer A., Jendrossek D. 1994; Molecular characterization of the extracellular poly(3-hydroxyoctanoic acid) [P(3HO)] depolymerase gene of Pseudomonas fluorescens GK13 and of its gene product. J Bacteriol 176:7065–7073
    [Google Scholar]
  50. 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 [CrossRef]
    [Google Scholar]
  51. Scott F. W., Pitt T. L. 2004; Identification and characterization of transmissible Pseudomonas aeruginosa strains in cystic fibrosis patients in England and Wales. J Med Microbiol 53:609–615 [CrossRef]
    [Google Scholar]
  52. Shen J., Meldrum A., Poole K. 2002; FpvA receptor involvement in pyoverdine biosynthesis in Pseudomonas aeruginosa . J Bacteriol 184:3268–3275 [CrossRef]
    [Google Scholar]
  53. Smyth G. 2003; Statistical issues in cDNA microarray data analysis. In Functional Genomics: Methods and Protocols pp 111–136 Edited by Brownstein M. J., Khodursky A. B. Totowa, NJ: Humana Press;
    [Google Scholar]
  54. Smyth G. K. 2004; Linear models and empirical Bayes methods for assessing differential expression in microarray experiments. Stat Appl Genet Mol Biol 3: (article 3)
    [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 [CrossRef]
    [Google Scholar]
  56. Suh S. J., Silo-Suh L., Woods D. E., Hassett D. J., West S. E., Ohman D. E. 1999; Effect of rpoS mutation on the stress response and expression of virulence factors in Pseudomonas aeruginosa . J Bacteriol 181:3890–3897
    [Google Scholar]
  57. Tenover F. C., Arbeit R. D., Goering R. V., Mickelsen P. A., Murray B. E., Persing D. H., Swaminathan B. 1995; Interpreting chromosomal DNA restriction patterns produced by pulsed-field gel electrophoresis: criteria for bacterial strain typing. J Clin Microbiol 33:2233–2239
    [Google Scholar]
  58. Van Alst N. E., Picardo K. F., Iglewski B. H., Haidaris C. G. 2007; Nitrate sensing and metabolism modulate motility, biofilm formation, and virulence in Pseudomonas aeruginosa . Infect Immun 75:3780–3790 [CrossRef]
    [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 [CrossRef]
    [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 [CrossRef]
    [Google Scholar]
  61. Webb J. S., Lau M., Kjelleberg S. 2004; Bacteriophage and phenotypic variation in Pseudomonas aeruginosa biofilm development. J Bacteriol 186:8066–8073 [CrossRef]
    [Google Scholar]
  62. 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 [CrossRef]
    [Google Scholar]
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