1887

Abstract

Pseudomonas aeruginosa is an environmental bacterium but is also an opportunistic pathogen. The aim of this work is to evaluate the contribution of P. aeruginosa LasR and RhlR transcriptional regulators of the quorum-sensing response (QSR) to the production of virulence factors, and to its virulence in a mouse abscess model. The QSR is a complex regulatory network that modulates the expression of several virulence factors, including elastase, pyocyanin and rhamnolipids. LasR, when complexed with the auto-inducer 3-oxo-dodecanoyl lactone (3O-C12-HSL), produced by LasI, is at the top of the QSR regulatory cascade since it activates transcription of some genes encoding virulence factors (such as the gene coding for elastase, lasB) and also transcription of both rhlR and rhlI, encoding the synthase of the auto-inducer butanoyl-homoserine lactone (C4-HSL). In turn RhlR, coupled with C4-HSL, activates the transcription of genes encoding for the enzymes involved in pyocyanin and rhamnolipid production. Several efforts have been made to obtain inhibitors of LasR activity that would suppress the QSR. However, these attempts have used chemical compounds that might not be specific for LasR inactivation. In this work we show that individual inactivation of either lasR or rhlR did not block the QSR, nor did it impair P. aeruginosa virulence, and that even a lasR rhlR double mutant still presented residual virulence, even lacking the production of virulence factors. These results show that the inhibition of either lasR or rhlR is not a straightforward approach to blocking P. aeruginosa virulence, due to the great complexity of the QSR.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.000778
2019-02-01
2020-01-27
Loading full text...

Full text loading...

References

  1. Driscoll JA, Brody SL, Kollef MH. The epidemiology, pathogenesis and treatment of Pseudomonas aeruginosa infections. Drugs 2007;67:351–368 [CrossRef][PubMed]
    [Google Scholar]
  2. Gellatly SL, Hancock RE. Pseudomonas aeruginosa: new insights into pathogenesis and host defenses. Pathog Dis 2013;67:159–173 [CrossRef][PubMed]
    [Google Scholar]
  3. Williams P, Cámara M. Quorum sensing and environmental adaptation in Pseudomonas aeruginosa: a tale of regulatory networks and multifunctional signal molecules. Curr Opin Microbiol 2009;12:182–191 [CrossRef][PubMed]
    [Google Scholar]
  4. Lee J, Zhang L. The hierarchy quorum sensing network in Pseudomonas aeruginosa. Protein Cell 2015;6:26–41 [CrossRef][PubMed]
    [Google Scholar]
  5. Mukherjee S, Moustafa D, Smith CD, Goldberg JB, Bassler BL et al. The RhlR quorum-sensing receptor controls Pseudomonas aeruginosa pathogenesis and biofilm development independently of its canonical homoserine lactone autoinducer. PLoS Pathog 2017;13:e1006504 [CrossRef][PubMed]
    [Google Scholar]
  6. Mukherjee S, Moustafa DA, Stergioula V, Smith CD, Goldberg JB et al. The PqsE and RhlR proteins are an autoinducer synthase-receptor pair that control virulence and biofilm development in Pseudomonas aeruginosa. Proc Natl Acad Sci USA 2018;115:E9411E9418 [CrossRef][PubMed]
    [Google Scholar]
  7. Grosso-Becerra MV, Santos-Medellín C, González-Valdez A, Méndez JL, Delgado G et al. Pseudomonas aeruginosa clinical and environmental isolates constitute a single population with high phenotypic diversity. BMC Genomics 2014;15:318 [CrossRef][PubMed]
    [Google Scholar]
  8. García-Contreras R, Peréz-Eretza B, Jasso-Chávez R, Lira-Silva E, Roldán-Sánchez JA et al. High variability in quorum quenching and growth inhibition by furanone C-30 in Pseudomonas aeruginosa clinical isolates from cystic fibrosis patients. Pathog Dis 2015;73:ftv040 [CrossRef][PubMed]
    [Google Scholar]
  9. Feltner JB, Wolter DJ, Pope CE, Groleau MC, Smalley NE et al. LasR variant cystic fibrosis isolates reveal an adaptable quorum-sensing hierarchy in Pseudomonas aeruginosa. MBio 2016;7:e0151301516 [CrossRef][PubMed]
    [Google Scholar]
  10. Morales E, González-Valdez A, Servín-González L, Soberón-Chávez G. Pseudomonas aeruginosa quorum-sensing response in the absence of functional LasR and LasI proteins: the case of strain 148, a virulent dolphin isolate. FEMS Microbiol Lett 2017;364: [CrossRef][PubMed]
    [Google Scholar]
  11. Schaber JA, Carty NL, McDonald NA, Graham ED, Cheluvappa R et al. Analysis of quorum sensing-deficient clinical isolates of Pseudomonas aeruginosa. J Med Microbiol 2004;53:841–853 [CrossRef][PubMed]
    [Google Scholar]
  12. Köhler T, Ouertatani-Sakouhi H, Cosson P, van Delden C. QsrO a novel regulator of quorum-sensing and virulence in Pseudomonas aeruginosa. PLoS One 2014;9:e87814 [CrossRef][PubMed]
    [Google Scholar]
  13. Amara N, Mashiach R, Amar D, Krief P, Spieser SA et al. Covalent inhibition of bacterial quorum sensing. J Am Chem Soc 2009;131:10610–10619 [CrossRef][PubMed]
    [Google Scholar]
  14. Jakobsen TH, Bjarnsholt T, Jensen , Givskov M, Høiby N. Targeting quorum sensing in Pseudomonas aeruginosa biofilms: current and emerging inhibitors. Future Microbiol 2013;8:901–921 [CrossRef][PubMed]
    [Google Scholar]
  15. O'Brien KT, Noto JG, Nichols-O'Neill L, Perez LJ. Potent Irreversible Inhibitors of LasR Quorum Sensing in Pseudomonas aeruginosa. ACS Med Chem Lett 2015;6:162–167 [CrossRef][PubMed]
    [Google Scholar]
  16. O'Loughlin CT, Miller LC, Siryaporn A, Drescher K, Semmelhack MF et al. A quorum-sensing inhibitor blocks Pseudomonas aeruginosa virulence and biofilm formation. Proc Natl Acad Sci USA 2013;110:17981–17986 [CrossRef][PubMed]
    [Google Scholar]
  17. Eibergen NR, Moore JD, Mattmann ME, Blackwell HE. Potent and Selective Modulation of the RhlR Quorum Sensing Receptor by Using Non-native Ligands: An Emerging Target for Virulence Control in Pseudomonas aeruginosa. Chembiochem 2015;16:2348–2356 [CrossRef][PubMed]
    [Google Scholar]
  18. Gi M, Jeong J, Lee K, Lee KM, Toyofuku M et al. A drug-repositioning screening identifies pentetic acid as a potential therapeutic agent for suppressing the elastase-mediated virulence of Pseudomonas aeruginosa. Antimicrob Agents Chemother 2014;58:7205–7214 [CrossRef][PubMed]
    [Google Scholar]
  19. Chang CY, Krishnan T, Wang H, Chen Y, Yin WF et al. Non-antibiotic quorum sensing inhibitors acting against N-acyl homoserine lactone synthase as druggable target. Sci Rep 2014;4:7245 [CrossRef][PubMed]
    [Google Scholar]
  20. Allegretta G, Weidel E, Empting M, Hartmann RW. Catechol-based substrates of chalcone synthase as a scaffold for novel inhibitors of PqsD. Eur J Med Chem 2015;90:351–359 [CrossRef][PubMed]
    [Google Scholar]
  21. Köhler T, Perron GG, Buckling A, van Delden C. Quorum Sensing inhibition selects for virulence and cooperation in Pseudomonas aeruginosa. PLoS Pathog 2010;6:e1000883 [CrossRef][PubMed]
    [Google Scholar]
  22. García-Contreras R. Is Quorum Sensing Interference a Viable Alternative to Treat Pseudomonas aeruginosa Infections?. Front Microbiol 2016;7:1454 [CrossRef][PubMed]
    [Google Scholar]
  23. Dacheux D, Attree I, Schneider C, Toussaint B. Cell death of human polymorphonuclear neutrophils induced by a Pseudomonas aeruginosa cystic fibrosis isolate requires a functional type III secretion system. Infect Immun 1999;67:6164–6167[PubMed]
    [Google Scholar]
  24. Mikkelsen H, McMullan R, Filloux A. The Pseudomonas aeruginosa reference strain PA14 displays increased virulence due to a mutation in ladS. PLoS One 2011;6:e29113 [CrossRef][PubMed]
    [Google Scholar]
  25. Pletzer D, Mansour SC, Wuerth K, Rahanjam N, Hancock RE. New mouse model for chronic infections by Gram-negative bacteria enabling the study of anti-infective efficacy and host-microbe interactions. MBio 2017;8:e0014017 [CrossRef][PubMed]
    [Google Scholar]
  26. Granato ET, Harrison F, Kümmerli R, Ross-Gillespie A. Do bacterial "virulence factors" always increase virulence? A meta-analysis of pyoverdine production in Pseudomonas aeruginosa as a test case. Front Microbiol 2016;7:7 [CrossRef][PubMed]
    [Google Scholar]
  27. Essar DW, Eberly L, Hadero A, Crawford IP. Identification and characterization of genes for a second anthranilate synthase in Pseudomonas aeruginosa: interchangeability of the two anthranilate synthases and evolutionary implications. J Bacteriol 1990;172:884–900 [CrossRef][PubMed]
    [Google Scholar]
  28. Beatson SA, Whitchurch CB, Sargent JL, Levesque RC, Mattick JS. Differential regulation of twitching motility and elastase production by Vfr in Pseudomonas aeruginosa. J Bacteriol 2002;184:3605–3613 [CrossRef][PubMed]
    [Google Scholar]
  29. Matsuyama T, Sogawa M, Yano I. Direct colony thin-layer chromatography and rapid characterization of Serratia marcescens mutants defective in production of wetting agents. Appl Environ Microbiol 1987;53:1186–1188[PubMed]
    [Google Scholar]
  30. Coffey BM, Anderson GG. Biofilm formation in the 96-well microtitre plate. In Filloux A, Ramos J-L. (editors) Pseudomonas Methods and Protocols. Methods in Molecular Biology 1149 New York, NY: Springer Protocols, Humana Press,; 2014; pp.631–641
    [Google Scholar]
  31. Diggle SP, Fletcher MP, Cámara M, Williams P. Detection of 2-alkyl-4-quinolone using biosensors. In Rumbaugh Kendra P. (editor) Quorum Sensing: Methods and Protocolsvol. 692 Methods in Molecular Biology; 2011
    [Google Scholar]
  32. Sambrook J, Fritsch EF, Maniatis T. Molecular Cloning: A Laboratory Manual, 2nd ed. Cold Spring Harbor, NY: (Cold Spring Harbor Lab Press); 1989
    [Google Scholar]
  33. Cadoret F, Soscia C, Voulhoux R. Gene transfer: transformation/electroporation. In Filloux A, Ramos J-L. (editors) Pseudomonas Methods and Protocols. Methods in Molecular Biology 1149 New York, NY: Springer Protocols, Humana Press; 2014; pp.271–279
    [Google Scholar]
  34. Choi KH, Schweizer HP. An improved method for rapid generation of unmarked Pseudomonas aeruginosa deletion mutants. BMC Microbiol 2005;5:30 [CrossRef][PubMed]
    [Google Scholar]
  35. Toska J, Sun Y, Carbonell DA, Foster AN, Jacobs MR et al. Diversity of virulence phenotypes among type III secretion negative Pseudomonas aeruginosa clinical isolates. PLoS One 2014;9:e86829 [CrossRef][PubMed]
    [Google Scholar]
  36. Heeb S, Blumer C, Haas D. Regulatory RNA as mediator in GacA/RsmA-dependent global control of exoproduct formation in Pseudomonas fluorescens CHA0. J Bacteriol 2002;184:1046–1056 [CrossRef][PubMed]
    [Google Scholar]
  37. Sharma V, Yamamura A, Yokobayashi Y. Engineering artificial small RNAs for conditional gene silencing in Escherichia coli. ACS Synth Biol 2012;1:6–13 [CrossRef][PubMed]
    [Google Scholar]
  38. Miller JH. Experiments in Molecular GeneticsVol. 433 Cold Spring Harb NY: Cold Spring Harb Lab Press; 1972; pp.352–355
    [Google Scholar]
  39. Berube BJ, Murphy KR, Torhan MC, Bowlin NO, Williams JD et al. Impact of type III secretion effectors and of phenoxyacetamide inhibitors of type III secretion on abscess formation in a mouse model of Pseudomonas aeruginosa infection. Antimicrob Agents Chemother 2017;61:e0120201217 [CrossRef][PubMed]
    [Google Scholar]
  40. Gong FY, Zhang DY, Zhang JG, Wang LL, Zhan WL et al. siRNA-mediated gene silencing of MexB from the MexA-MexB-OprM efflux pump in Pseudomonas aeruginosa. BMB Rep 2014;47:203–208 [CrossRef][PubMed]
    [Google Scholar]
  41. Gottesman S. Micros for microbes: non-coding regulatory RNAs in bacteria. Trends Genet 2005;21:399–404 [CrossRef][PubMed]
    [Google Scholar]
  42. Rumbaugh KP, Griswold JA, Iglewski BH, Hamood AN. Contribution of quorum sensing to the virulence of Pseudomonas aeruginosa in burn wound infections. Infect Immun 1999;67:5854–5862[PubMed]
    [Google Scholar]
  43. Castillo-Juarez I, López-Jácome LE, Soberón-Chávez G, Tomás M, Lee J et al. Exploiting Quorum Sensing inhibition for the control of Pseudomonas aeruginosa and Acinetobacter baumanni biofilms. Curr Top Med Chem 2017;17:1915–1927 [CrossRef][PubMed]
    [Google Scholar]
  44. Wang S, Yu S, Zhang Z, Wei Q, Yan L et al. Coordination of swarming motility, biosurfactant synthesis, and biofilm matrix exopolysaccharide production in Pseudomonas aeruginosa. Appl Environ Microbiol 2014;80:6724–6732 [CrossRef][PubMed]
    [Google Scholar]
  45. Cornforth DM, Dees JL, Ibberson CB, Huse HK, Mathiesen IH et al. Pseudomonas aeruginosa transcriptome during human infection. Proc Natl Acad Sci USA 2018;115:E5125E5134 [CrossRef][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.000778
Loading
/content/journal/micro/10.1099/mic.0.000778
Loading

Data & Media loading...

Supplements

Supplementary File 1

PDF

Supplementary File 2

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