Advertisement

Quorum Sensing Autoinducer(s) and Flagellum Independently Mediate EPS Signaling in Vibrio cholerae Through LuxO-Independent Mechanism

  • Smritikana Biswas
  • Prithwiraj Mukherjee
  • Tuhin Manna
  • Kunal Dutta
  • Kartik Chandra Guchhait
  • Amit Karmakar
  • Monalisha Karmakar
  • Parimal Dua
  • Amiya Kumar Panda
  • Chandradipa Ghosh
Environmental Microbiology

Abstract

Vibrio cholerae, the Gram-negative bacterium causing lethal diarrheal disease cholera, forms biofilm on solid surfaces to gain adaptive advantage for successful survival in aquatic reservoirs. Expression of exopolysaccharide (EPS), an extracellular matrix material, has been found critical for biofilm-based environmental persistence. In a subset of epidemic-causing V. cholerae, absence of flagellum but not motility was identified to induce elevated exopolysaccharide expression. Identification of the role played by quorum sensing autoinducer molecules, i.e., cholera autoinducer 1 (CAI-1) and autoinducer 2 (AI-2) as well as central regulator LuxO on EPS expression in the subset was explored. Deletion mutations were introduced in vital genes responsible for synthesizing CAI-1 (cqsA), AI-2 (luxS), flagellum (flaA), LuxO (luxO), flagellar motor (motX), and VpsR (vpsR) in the model strain MO10. Subsequent phenotypic alterations in terms of colony morphology, EPS expression, biofilm formation, and transcription level of relevant genes were analyzed. Autoinducer cross-feeding experiment confirmed the role of autoinducers in EPS signaling. Results reveal that autoinducers and flagellum are the two major EPS signaling units in this subset where one unit becomes predominant for EPS production in absence of the other. Moreover, either unit exerts negative influence on EPS induction by the other. Both the EPS signaling cascades are independent of LuxO contribution and essentially involve sodium-driven flagellar motor and VpsR. A cell density and flagellum-mediated, but LuxO-independent, EPS signaling mechanism is considered to be functional in these organisms that confers their survival fitness.

Keywords

Vibrio cholerae Quorum sensing Exopolysaccharide Biofilm Flagellum Cholera autoinducer 1 Autoinducer 2 Rugose colony 

Notes

Acknowledgements

Authors thank Prof. Karl E. Klose (Dept. of Biology, UTSA, USA) for gifts of strains and plasmid vectors and Prof. Sunando Bandyopadhyay, Department of Geography, University of Calcutta, for technical support.

Funding information

This study is financially supported by the Science and Engineering Research Council, Department of Science and Technology (DST), Government of India, in the form of a research grant (SR/SO/HS-43/2006), and the Department of Biotechnology, Govt. of India (Project Number BT/PR3802/BRB/10/981/2011).

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interest.

References

  1. 1.
    Organization WH (2013) Monthly report on dracunculiasis cases, January-May 2013, Rapport mensuel des cas de dracunculose, janvier-mai 2013. Wkly Epidemiol. Rec. 88:335–336Google Scholar
  2. 2.
    Levine MM, Kaper JB, Black R, Clements ML (1983) New knowledge on pathogenesis of bacterial enteric infections as applied to vaccine development. Microbiol. Rev. 47:510PubMedPubMedCentralGoogle Scholar
  3. 3.
    Taylor RK, Miller VL, Furlong DB, Mekalanos JJ (1987) Use of phoA gene fusions to identify a pilus colonization factor coordinately regulated with cholera toxin. Proc. Natl. Acad. Sci. U. S. A. 84:2833–2837CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Skorupski K, Taylor RK (1997) Control of the ToxR virulence regulon in Vibrio cholerae by environmental stimuli. Mol. Microbiol. 25:1003–1009CrossRefPubMedGoogle Scholar
  5. 5.
    Watnick PI, Lauriano CM, Klose KE, Croal L, Kolter R (2001) The absence of a flagellum leads to altered colony morphology, biofilm development and virulence in Vibrio cholerae O139. Mol. Microbiol. 39:223–235CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Lutz C, Erken M, Noorian P, Sun S, McDougald D (2013) Environmental reservoirs and mechanisms of persistence of Vibrio cholerae. Front. Microbiol. 4:375CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Yildiz FH, Visick KL (2009) Vibrio biofilms: so much the same yet so different. Trends Microbiol. 17:109–118CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Yildiz FH, Schoolnik GK (1999) Vibrio cholerae O1 El Tor: identification of a gene cluster required for the rugose colony type, exopolysaccharide production, chlorine resistance, and biofilm formation. Proc. Natl. Acad. Sci. U. S. A. 96:4028–4033CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    White PB (1938) The rugose variant of Vibrios. J. Pathol. 46:1–6CrossRefGoogle Scholar
  10. 10.
    Zhu J, Mekalanos JJ (2003) Quorum sensing-dependent biofilms enhance colonization in Vibrio cholerae. Dev. Cell 5:647–656CrossRefPubMedGoogle Scholar
  11. 11.
    Miller MB, Skorupski K, Lenz DH, Taylor RK, Bassler BL (2002) Parallel quorum sensing systems converge to regulate virulence in Vibrio cholerae. Cell 110:303–314CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Henke JM, Bassler BL (2004) Three parallel quorum-sensing systems regulate gene expression in Vibrio harveyi. J. Bacteriol. 186:6902–6914CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Ng WL, Perez LJ, Wei Y, Kraml C, Semmelhack MF, Bassler BL (2011) Signal production and detection specificity in Vibrio CqsA/CqsS quorum-sensing systems. Mol. Microbiol. 79:1407–1417CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Wei Y, Perez LJ, Ng W-L, Semmelhack MF, Bassler BL (2011) Mechanism of Vibrio cholerae autoinducer-1 biosynthesis. ACS Chem. Biol. 6:356–365CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Schauder S, Shokat K, Surette MG, Bassler BL (2001) The LuxS family of bacterial autoinducers: biosynthesis of a novel quorum-sensing signal molecule. Mol. Microbiol. 41:463–476CrossRefPubMedGoogle Scholar
  16. 16.
    Bassler BL (1999) How bacteria talk to each other: regulation of gene expression by quorum sensing. Curr. Opin. Microbiol. 2:582–587CrossRefPubMedGoogle Scholar
  17. 17.
    Higgins DA, Pomianek ME, Kraml CM, Taylor RK, Semmelhack MF, Bassler BL (2007) The major Vibrio cholerae autoinducer and its role in virulence factor production. Nature 450:883–886CrossRefPubMedGoogle Scholar
  18. 18.
    Jobling MG, Holmes RK (1997) Characterization of hapR, a positive regulator of the Vibrio cholerae HA/protease gene hap, and its identification as a functional homologue of the Vibrio harveyi luxR gene. Mol. Microbiol. 26:1023–1034CrossRefPubMedGoogle Scholar
  19. 19.
    Römling U, Amikam D (2006) Cyclic di-GMP as a second messenger. Curr. Opin. Microbiol. 9:218–228CrossRefPubMedGoogle Scholar
  20. 20.
    Hammer BK, Bassler BL (2009) Distinct sensory pathways in Vibrio cholerae El Tor and classical biotypes modulate cyclic dimeric GMP levels to control biofilm formation. J. Bacteriol. 191:169–177CrossRefPubMedGoogle Scholar
  21. 21.
    Jung SA, Chapman CA, Ng W-L (2015) Quadruple quorum-sensing inputs control Vibrio cholerae virulence and maintain system robustness. PLoS Pathog. 11:e1004837CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Papenfort K, Bassler BL (2016) Quorum sensing signal-response systems in gram-negative bacteria. Nat Rev Microbiol 14:576–588CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Lauriano CM, Ghosh C, Correa NE, Klose KE (2004) The sodium-driven flagellar motor controls exopolysaccharide expression in Vibrio cholerae. J. Bacteriol. 186:4864–4874CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Shao Y, Bassler BL (2014) Quorum regulatory small RNAs repress type VI secretion in Vibrio cholerae. Mol. Microbiol. 92:921–930CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Joelsson A, Liu Z, Zhu J (2006) Genetic and phenotypic diversity of quorum-sensing systems in clinical and environmental isolates of Vibrio cholerae. Infect. Immun. 74:1141–1147CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Hanahan D (1983) Studies on transformation of Escherichia coli with plasmids. J. Mol. Biol. 166:557–580CrossRefPubMedGoogle Scholar
  27. 27.
    Miller VL, Mekalanos JJ (1988) A novel suicide vector and its use in construction of insertion mutations: osmoregulation of outer membrane proteins and virulence determinants in Vibrio cholerae requires toxR. J. Bacteriol. 170:2575–2583CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Correa NE, Lauriano CM, McGee R, Klose KE (2000) Phosphorylation of the flagellar regulatory protein FlrC is necessary for Vibrio cholerae motility and enhanced colonization. Mol. Microbiol. 35:743–755CrossRefPubMedGoogle Scholar
  29. 29.
    Simons RW, Houman F, Kleckner N (1987) Improved single and multicopy lac-based cloning vectors for protein and operon fusions. Gene 53:85–96CrossRefPubMedGoogle Scholar
  30. 30.
    Sambrook J, Russel DW, Russel DW (2001) Molecular cloning: a laboratory manual (Ed. 3). Cold Spring Harbor Laboratory Press, NY, USAGoogle Scholar
  31. 31.
    Heidelberg JF, Eisen JA, Nelson WC, Clayton RA, Gwinn ML, Dodson RJ, Haft DH, Hickey EK, Peterson JD, Umayam L (2000) DNA sequence of both chromosomes of the cholera pathogen Vibrio cholerae. Nature 406:477–483CrossRefGoogle Scholar
  32. 32.
    Miller JH (1992) A short course in bacterial genetics : a laboratory manual and handbook for Escherichia coli and related bacteria. Cold Spring Harbor laboratory press. New York (N.Y.)Google Scholar
  33. 33.
    Sun M-L, Zhao F, Shi M, Zhang X-Y, Zhou B-C, Zhang Y-Z, Chen X-L (2015) Characterization and biotechnological potential analysis of a new exopolysaccharide from the Arctic marine bacterium Polaribacter sp. SM1127. Sci. Rep. 5:18435CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Dubois M, Gilles KA, Hamilton JK, Pt R, Smith F (1956) Colorimetric method for determination of sugars and related substances. Anal. Chem. 28:350–356CrossRefGoogle Scholar
  35. 35.
    Fong JC, Yildiz FH (2007) The rbmBCDEF gene cluster modulates development of rugose colony morphology and biofilm formation in Vibrio cholerae. J. Bacteriol. 189:2319–2330CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Mok KC, Wingreen NS, Bassler BL (2003) Vibrio harveyi quorum sensing: a coincidence detector for two autoinducers controls gene expression. EMBO J. 22:870–881CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Antonova ES, Hammer BK (2011) Quorum-sensing autoinducer molecules produced by members of a multispecies biofilm promote horizontal gene transfer to Vibrio cholerae. FEMS Microbiol. Lett. 322:68–76CrossRefPubMedGoogle Scholar
  38. 38.
    Bassler BL, Wright M, Silverman MR (1994) Multiple signalling systems controlling expression of luminescence in Vibrio harveyi: sequence and function of genes encoding a second sensory pathway. Mol. Microbiol. 13:273–286CrossRefPubMedGoogle Scholar
  39. 39.
    Kawagishi I, Imagawa M, Imae Y, McCarter L, Homma M (1996) The sodium-driven polar flagellar motor of marine Vibrio as the mechanosensor that regulates lateral flagellar expression. Mol. Microbiol. 20:693–699CrossRefPubMedGoogle Scholar
  40. 40.
    Yildiz FH, Dolganov NA, Schoolnik GK (2001) VpsR, a member of the response regulators of the two-component regulatory systems, is required for expression of vps biosynthesis genes and EPSETr-associated phenotypes in Vibrio cholerae O1 El Tor. J. Bacteriol. 183:1716–1726CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Smritikana Biswas
    • 1
  • Prithwiraj Mukherjee
    • 1
  • Tuhin Manna
    • 1
  • Kunal Dutta
    • 1
    • 2
  • Kartik Chandra Guchhait
    • 1
  • Amit Karmakar
    • 1
  • Monalisha Karmakar
    • 1
  • Parimal Dua
    • 1
  • Amiya Kumar Panda
    • 2
  • Chandradipa Ghosh
    • 1
  1. 1.Department of Human Physiology with Community HealthVidyasagar UniversityMidnaporeIndia
  2. 2.Department of Chemistry and Chemical TechnologyVidyasagar UniversityMidnaporeIndia

Personalised recommendations