Identification of Anti-virulence Compounds That Disrupt Quorum-Sensing Regulated Acute and Persistent Pathogenicity

English version České info

Antibiotic resistant and tolerant bacterial pathogens are responsible for acute, chronic and persistent human infections recalcitrant to any current treatments. Therefore, there is an urgent need to identify new antimicrobial drugs that will help circumvent the current antibiotic resistance crisis. Bacterial pathogens often develop resistance to antibiotic drugs that target bacterial growth or viability. In contrast, strategies that specifically target virulence pathways non-essential for growth could limit selective resistance, and thus are candidates for the development of next-generation antimicrobial therapeutics. In this study we target the bacterial communication system MvfR (PqsR), which is known to control virulence of the opportunistic bacterial pathogen Pseudomonas aeruginosa. We identified and improved upon new small molecules that effectively silence the MvfR communication system, and as a result block P. aeruginosa virulence both in vitro and in vivo. Moreover, these new compounds are the first known to restrict the ability of bacteria to form antibiotic-tolerant cells and consequently proved to be very effective at preventing persistent infection in a mammalian infection model. Because of their ability to simultaneously block acute and persistent infections, these new molecules may provide a very strong basis for the development of next generation antimicrobials.

Vyšlo v časopise: Identification of Anti-virulence Compounds That Disrupt Quorum-Sensing Regulated Acute and Persistent Pathogenicity. PLoS Pathog 10(8): e32767. doi:10.1371/journal.ppat.1004321
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.ppat.1004321

Souhrn

Zdroje

1. BoucherHW, TalbotGH, BradleyJS, EdwardsJE, GilbertD, et al. (2009) Bad bugs, no drugs: no ESKAPE! An update from the Infectious Diseases Society of America. Clin Infect Dis 48 : 1–12.

2. LivermoreDM (2012) Fourteen years in resistance. Int J Antimicrob Agents 39 : 283–294.

3. SpellbergB, GuidosR, GilbertD, BradleyJ, BoucherHW, et al. (2008) The epidemic of antibiotic-resistant infections: a call to action for the medical community from the Infectious Diseases Society of America. Clin Infect Dis 46 : 155–164.

4. ParkerCT, SperandioV (2009) Cell-to-cell signalling during pathogenesis. Cell Microbiol 11 : 363–369.

5. NgWL, BasslerBL (2009) Bacterial quorum-sensing network architectures. Annu Rev Genet 43 : 197–222.

6. BandyopadhayaA, KesarwaniM, QueYA, HeJ, PadfieldK, et al. (2012) The quorum sensing volatile molecule 2-amino acetophenon modulates host immune responses in a manner that promotes life with unwanted guests. PLoS Pathog 8: e1003024.

7. KesarwaniM, HazanR, HeJ, QueY, ApidianakisY, et al. (2011) A Quorum Sensing Regulated Small Volatile Molecule Reduces Acute Virulence and Promotes Chronic Infection Phenotypes. PLoS Pathogens 7: e1002192.

8. QueYA, HazanR, StrobelB, MauraD, HeJ, et al. (2013) A quorum sensing small volatile molecule promotes antibiotic tolerance in bacteria. PLoS One 8: e80140.

9. MökerN, DeanCR, TaoJ (2010) Pseudomonas aeruginosa increases formation of multidrug-tolerant persister cells in response to quorum-sensing signaling molecules. Journal of bacteriology 192 : 1946–1955.

10. VegaNM, AllisonKR, KhalilAS, CollinsJJ (2012) Signaling-mediated bacterial persister formation. Nat Chem Biol 8 : 431–433.

11. RaskoDA, SperandioV (2010) Anti-virulence strategies to combat bacteria-mediated disease. Nat Rev Drug Discov 9 : 117–128.

12. DefoirdtT, BrackmanG, CoenyeT (2013) Quorum sensing inhibitors: how strong is the evidence? Trends Microbiol 21 : 619–624.

13. NjorogeJ, SperandioV (2009) Jamming bacterial communication: new approaches for the treatment of infectious diseases. EMBO Mol Med 1 : 201–210.

14. WilliamsP, CamaraM (2009) Quorum sensing and environmental adaptation in Pseudomonas aeruginosa: a tale of regulatory networks and multifunctional signal molecules. Curr Opin Microbiol 12 : 182–191.

15. DezielE, LepineF, MilotS, HeJ, MindrinosMN, et al. (2004) Analysis of Pseudomonas aeruginosa 4-hydroxy-2-alkylquinolines (HAQs) reveals a role for 4-hydroxy-2-heptylquinoline in cell-to-cell communication. Proc Natl Acad Sci U S A 101 : 1339–1344.

16. XiaoG, DezielE, HeJ, LepineF, LesicB, et al. (2006) MvfR, a key Pseudomonas aeruginosa pathogenicity LTTR-class regulatory protein, has dual ligands. Mol Microbiol 62 : 1689–1699.

17. SchellMA (1993) Molecular biology of the LysR family of transcriptional regulators. Annu Rev Microbiol 47 : 597–626.

18. TangHB, DiMangoE, BryanR, GambelloM, IglewskiBH, et al. (1996) Contribution of specific Pseudomonas aeruginosa virulence factors to pathogenesis of pneumonia in a neonatal mouse model of infection. Infect Immun 64 : 37–43.

19. CaoH, KrishnanG, GoumnerovB, TsongalisJ, TompkinsR, et al. (2001) A quorum sensing-associated virulence gene of Pseudomonas aeruginosa encodes a LysR-like transcription regulator with a unique self-regulatory mechanism. Proceedings of the National Academy of Sciences 98 : 14613.

20. RahmeLG, StevensEJ, WolfortSF, ShaoJ, TompkinsRG, et al. (1995) Common virulence factors for bacterial pathogenicity in plants and animals. Science (New York, NY) 268 : 1899–1902.

21. PearsonJP, PesciEC, IglewskiBH (1997) Roles of Pseudomonas aeruginosa las and rhl quorum-sensing systems in control of elastase and rhamnolipid biosynthesis genes. J Bacteriol 179 : 5756–5767.

22. D'ArgenioDA, WuM, HoffmanLR, KulasekaraHD, DezielE, et al. (2007) Growth phenotypes of Pseudomonas aeruginosa lasR mutants adapted to the airways of cystic fibrosis patients. Mol Microbiol 64 : 512–533.

23. SmithEE, BuckleyDG, WuZ, SaenphimmachakC, HoffmanLR, et al. (2006) Genetic adaptation by Pseudomonas aeruginosa to the airways of cystic fibrosis patients. Proc Natl Acad Sci U S A 103 : 8487–8492.

24. LauGW, GoumnerovBC, WalendziewiczCL, HewitsonJ, XiaoW, et al. (2003) The Drosophila melanogaster toll pathway participates in resistance to infection by the gram-negative human pathogen Pseudomonas aeruginosa. Infect Immun 71 : 4059–4066.

25. DézielE, GopalanS, TampakakiAP, LépineF, PadfieldKE, et al. (2005) The contribution of MvfR to Pseudomonas aeruginosa pathogenesis and quorum sensing circuitry regulation: multiple quorum sensing-regulated genes are modulated without affecting lasRI, rhlRI or the production of N-acyl-L-homoserine lactones. Mol Microbiol 55 : 998–1014.

26. WadeDS, CalfeeMW, RochaER, LingEA, EngstromE, et al. (2005) Regulation of Pseudomonas quinolone signal synthesis in Pseudomonas aeruginosa. J Bacteriol 187 : 4372–4380.

27. QueY, HazanR, RyanCM, MilotS, LépineF, et al. (2011) Production of Pseudomonas aeruginosa Intercellular Small Signaling Molecules in Human Burn Wounds. Journal of Pathogens 2011 : 1–5.

28. TaylorGW, MachanZA, MehmetS, ColePJ, WilsonR (1995) Rapid identification of 4-hydroxy-2-alkylquinolines produced by Pseudomonas aeruginosa using gas chromatography-electron-capture mass spectrometry. J Chromatogr B Biomed Appl 664 : 458–462.

29. DiggleSP, MatthijsS, WrightVJ, FletcherMP, ChhabraSR, et al. (2007) The Pseudomonas aeruginosa 4-quinolone signal molecules HHQ and PQS play multifunctional roles in quorum sensing and iron entrapment. Chemistry & biology 14 : 87–96.

30. HeebS, FletcherMP, ChhabraSR, DiggleSP, WilliamsP, et al. (2011) Quinolones: from antibiotics to autoinducers. FEMS Microbiol Rev 35 : 247–274.

31. HazanR, HeJ, XiaoG, DekimpeV, ApidianakisY, et al. (2010) Homeostatic interplay between bacterial cell-cell signaling and iron in virulence. PLoS Pathogens 6: e1000810.

32. Scott-ThomasAJ, SyhreM, PattemorePK, EptonM, LaingR, et al. (2010) 2-Aminoacetophenone as a potential breath biomarker for Pseudomonas aeruginosa in the cystic fibrosis lung. BMC Pulm Med 10 : 56.

33. TzikaAA, ConstantinouC, BandyopadhayaA, PsychogiosN, LeeS, et al. (2013) A small volatile bacterial molecule triggers mitochondrial dysfunction in murine skeletal muscle. PLoS One 8: e74528.

34. LewisK (2010) Persister Cells. Annual review of microbiology 64 : 357–72.

35. FauvartM, De GrooteVN, MichielsJ (2011) Role of persister cells in chronic infections: clinical relevance and perspectives on anti-persister therapies. J Med Microbiol 60 : 699–709.

36. GefenO, BalabanNQ (2009) The importance of being persistent: heterogeneity of bacterial populations under antibiotic stress. FEMS microbiology reviews 33 : 704–717.

37. PelicicV, ReyratJM, GicquelB (1996) Expression of the Bacillus subtilis sacB gene confers sucrose sensitivity on mycobacteria. J Bacteriol 178 : 1197–1199.

38. YangL, BarkenKB, SkindersoeME, ChristensenAB, GivskovM, et al. (2007) Effects of iron on DNA release and biofilm development by Pseudomonas aeruginosa. Microbiology (Reading, England) 153 : 1318–1328.

39. FarrowJM, PesciEC (2007) Two distinct pathways supply anthranilate as a precursor of the Pseudomonas quinolone signal. Journal of bacteriology 189 : 3425–3433.

40. LesicB, LepineF, DezielE, ZhangJ, ZhangQ, et al. (2007) Inhibitors of pathogen intercellular signals as selective anti-infective compounds. PLoS Pathog 3 : 1229–1239.

41. HazanR, QueYA, MauraD, RahmeLG (2012) A method for high throughput determination of viable bacteria cell counts in 96-well plates. BMC Microbiol 12 : 259.

42. LepineF, DekimpeV, LesicB, MilotS, LesimpleA, et al. (2007) PqsA is required for the biosynthesis of 2,4-dihydroxyquinoline (DHQ), a newly identified metabolite produced by Pseudomonas aeruginosa and Burkholderia thailandensis. Biol Chem 388 : 839–845.

43. ZhangYM, FrankMW, ZhuK, MayasundariA, RockCO (2008) PqsD is responsible for the synthesis of 2,4-dihydroxyquinoline, an extracellular metabolite produced by Pseudomonas aeruginosa. J Biol Chem 283 : 28788–28794.

44. XiaoG, HeJ, RahmeLG (2006) Mutation analysis of the Pseudomonas aeruginosa mvfR and pqsABCDE gene promoters demonstrates complex quorum-sensing circuitry. Microbiology 152 : 1679–1686.

45. RahmeLG, AusubelFM, CaoH, DrenkardE, GoumnerovBC, et al. (2000) Plants and animals share functionally common bacterial virulence factors. Proc Natl Acad Sci U S A 97 : 8815–8821.

46. RahmeLG, TanMW, LeL, WongSM, TompkinsRG, et al. (1997) Use of model plant hosts to identify Pseudomonas aeruginosa virulence factors. Proc Natl Acad Sci U S A 94 : 13245–13250.

47. Mahajan-MiklosS, TanMW, RahmeLG, AusubelFM (1999) Molecular mechanisms of bacterial virulence elucidated using a Pseudomonas aeruginosa-Caenorhabditis elegans pathogenesis model. Cell 96 : 47–56.

48. LavoieEG, WangdiT, KazmierczakBI (2011) Innate immune responses to Pseudomonas aeruginosa infection. Microbes Infect 13 : 1133–1145.

49. BjarnsholtT, Kirketerp-MollerK, JensenPO, MadsenKG, PhippsR, et al. (2008) Why chronic wounds will not heal: a novel hypothesis. Wound Repair Regen 16 : 2–10.

50. MulcahyLR, BurnsJL, LoryS, LewisK (2010) Emergence of Pseudomonas aeruginosa strains producing high levels of persister cells in patients with cystic fibrosis. Journal of bacteriology 192 : 6191–6199.

51. UbedaC, PamerEG (2012) Antibiotics, microbiota, and immune defense. Trends Immunol 33 : 459–466.

52. MellbyeB, SchusterM (2011) The sociomicrobiology of antivirulence drug resistance: a proof of concept. MBio 2: e00131–11.

53. CalfeeMW, ColemanJP, PesciEC (2001) Interference with Pseudomonas quinolone signal synthesis inhibits virulence factor expression by Pseudomonas aeruginosa. Proc Natl Acad Sci U S A 98 : 11633–11637.

54. LeeJH, KimYG, ChoMH, KimJA, LeeJ (2012) 7-fluoroindole as an antivirulence compound against Pseudomonas aeruginosa. FEMS Microbiol Lett 329 : 36–44.

55. TashiroY, ToyofukuM, Nakajima-KambeT, UchiyamaH, NomuraN (2010) Bicyclic compounds repress membrane vesicle production and Pseudomonas quinolone signal synthesis in Pseudomonas aeruginosa. FEMS Microbiol Lett 304 : 123–130.

56. YangL, LiuY, SternbergC, MolinS (2010) Evaluation of enoyl-acyl carrier protein reductase inhibitors as Pseudomonas aeruginosa quorum-quenching reagents. Molecules 15 : 780–792.

57. YangL, RybtkeMT, JakobsenTH, HentzerM, BjarnsholtT, et al. (2009) Computer-aided identification of recognized drugs as Pseudomonas aeruginosa quorum-sensing inhibitors. Antimicrob Agents Chemother 53 : 2432–2443.

58. CuginiC, CalfeeMW, FarrowJM3rd, MoralesDK, PesciEC, et al. (2007) Farnesol, a common sesquiterpene, inhibits PQS production in Pseudomonas aeruginosa. Mol Microbiol 65 : 896–906.

59. LuC, KirschB, ZimmerC, de JongJC, HennC, et al. (2012) Discovery of antagonists of PqsR, a key player in 2-alkyl-4-quinolone-dependent quorum sensing in Pseudomonas aeruginosa. Chem Biol 19 : 381–390.

60. IlangovanA, FletcherM, RampioniG, PustelnyC, RumbaughK, et al. (2013) Structural Basis for Native Agonist and Synthetic Inhibitor Recognition by the Pseudomonas aeruginosa Quorum Sensing Regulator PqsR (MvfR). PLoS Pathog 9: e1003508 doi:1003510.1001371/journal.ppat.1003508

61. KleinT, HennC, de JongJC, ZimmerC, KirschB, et al. (2012) Identification of small-molecule antagonists of the Pseudomonas aeruginosa transcriptional regulator PqsR: biophysically guided hit discovery and optimization. ACS Chem Biol 7 : 1496–1501.

62. LuC, MaurerCK, KirschB, SteinbachA, HartmannRW (2013) Overcoming the Unexpected Functional Inversion of a PqsR Antagonist in Pseudomonas aeruginosa: An In Vivo Potent Antivirulence Agent Targeting pqs Quorum Sensing. Angew Chem Int Ed Engl 126 : 1127–1130.

63. ChungPY, TohYS (2014) Anti-biofilm agents: recent breakthrough against multi-drug resistant Staphylococcus aureus. Pathog Dis 70 : 231–239.

64. BjarnsholtT, CiofuO, MolinS, GivskovM, HoibyN (2013) Applying insights from biofilm biology to drug development -⁠ can a new approach be developed? Nat Rev Drug Discov 12 : 791–808.

65. AllisonKR, BrynildsenMP, CollinsJJ (2011) Metabolite-enabled eradication of bacterial persisters by aminoglycosides. Nature 473 : 216–220.

66. GrantSS, KaufmannBB, ChandNS, HaseleyN, HungDT (2012) Eradication of bacterial persisters with antibiotic-generated hydroxyl radicals. Proc Natl Acad Sci U S A 109 : 12147–12152.

67. ConlonBP, NakayasuES, FleckLE, LaFleurMD, IsabellaVM, et al. (2013) Activated ClpP kills persisters and eradicates a chronic biofilm infection. Nature 503 : 365–370.

68. KimJS, HeoP, YangTJ, LeeKS, JinYS, et al. (2011) Bacterial persisters tolerate antibiotics by not producing hydroxyl radicals. Biochem Biophys Res Commun 413 : 105–110.

69. KrushkalJ, QuY, LovleyDR, AdkinsRM (2012) Phylogenetic classification of diverse LysR-type transcriptional regulators of a model prokaryote Geobacter sulfurreducens. J Mol Evol 74 : 187–205.

70. DerbiseA, LesicB, DacheuxD, GhigoJM, CarnielE (2003) A rapid and simple method for inactivating chromosomal genes in Yersinia. FEMS Immunol Med Microbiol 38 : 113–116.

71. LesicB, RahmeLG (2008) Use of the lambda Red recombinase system to rapidly generate mutants in Pseudomonas aeruginosa. BMC Mol Biol 9 : 20.

72. ColemanJP, HudsonLL, McKnightSL, FarrowJM, CalfeeMW, et al. (2008) Pseudomonas aeruginosa PqsA is an anthranilate-coenzyme A ligase. Journal of bacteriology 190 : 1247–1255.

73. GallagherLA, McKnightSL, KuznetsovaMS, PesciEC, ManoilC (2002) Functions required for extracellular quinolone signaling by Pseudomonas aeruginosa. Journal of bacteriology 184 : 6472.

74. PelicicV, ReyratJM, GicquelB (1996) Generation of unmarked directed mutations in mycobacteria, using sucrose counter-selectable suicide vectors. Mol Microbiol 20 : 919–925.

75. ZhangJH, ChungTD, OldenburgKR (1999) A Simple Statistical Parameter for Use in Evaluation and Validation of High Throughput Screening Assays. J Biomol Screen 4 : 67–73.

76. LepineF, DezielE, MilotS, RahmeLG (2003) A stable isotope dilution assay for the quantification of the Pseudomonas quinolone signal in Pseudomonas aeruginosa cultures. Biochim Biophys Acta 1622 : 36–41.

77. EssarDW, EberlyL, HaderoA, CrawfordIP (1990) Identification and characterization of genes for a second anthranilate synthase in Pseudomonas aeruginosa: interchangeability of the two anthranilate synthases and evolutionary implications. J Bacteriol 172 : 884–900.

78. CastangS, McManusHR, TurnerKH, DoveSL (2008) H-NS family members function coordinately in an opportunistic pathogen. Proc Natl Acad Sci U S A 105 : 18947–18952.

79. Vallet-GelyI, DonovanKE, FangR, JoungJK, DoveSL (2005) Repression of phase-variable cup gene expression by H-NS-like proteins in Pseudomonas aeruginosa. Proc Natl Acad Sci U S A 102 : 11082–11087.

80. SavliH, KaradenizliA, KolayliF, GundesS, OzbekU, et al. (2003) Expression stability of six housekeeping genes: A proposal for resistance gene quantification studies of Pseudomonas aeruginosa by real-time quantitative RT-PCR. J Med Microbiol 52 : 403–408.

81. MosmannT (1983) Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 65 : 55–63.

82. ComolliJC, HauserAR, WaiteL, WhitchurchCB, MattickJS, et al. (1999) Pseudomonas aeruginosa gene products PilT and PilU are required for cytotoxicity in vitro and virulence in a mouse model of acute pneumonia. Infect Immun 67 : 3625–3630.

83. GarwoodM, DelaBarreL (2001) The return of the frequency sweep: designing adiabatic pulses for contemporary NMR. J Magn Reson 153 : 155–177.

84. LevittMH, FreemanR, FrenkielT (1982) Broadband heteronuclear decoupling. J Magn Reson 47 : 328–330.

85. Rankin ID (2005) MIC testing. In: Coyle MB, editor. Manual of Antimicrobial Susceptibility Testing: American Society for Microbiology. pp. 53–62.

86. BauerAW, RobertsCEJr, KirbyWM (1959) Single disc versus multiple disc and plate dilution techniques for antibiotic sensitivity testing. Antibiot Annu 7 : 574–580.