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. 1996 Jun;64(6):2246–2255. doi: 10.1128/iai.64.6.2246-2255.1996

Alterations in Vibrio cholerae motility phenotypes correlate with changes in virulence factor expression.

C L Gardel 1, J J Mekalanos 1
PMCID: PMC174063  PMID: 8675334

Abstract

Motility is thought to contribute to the virulence of Vibrio cholerae, but the role it plays in pathogenesis is not completely understood. To investigate the influence of motility on virulence gene expression and intestinal colonization, we have isolated mutants with altered swarming abilities in soft agar medium. Both spontaneous hyperswarmer (exhibiting faster swarm rates) and spontaneous or transposon-induced nonmotile mutants of strain 0395 were obtained. Surprisingly, we found that two of three classes of hyperswarmer mutants were defective in autoagglutination, a phenotype associated with expression of toxin-coregulated pili (TCP), an essential ToxR-regulated colonization factor of V. cholerae. In contrast, nonmotile mutants exhibited autoagglutination under growth conditions that normally repress this phenotype. Further characterization of mutant strains revealed differences in the expression of other virulence determinants. Class I hyperswarmer mutants were defective in production of TCP, cholera toxin, and a cell-associated hemolysin but showed increased levels of protease and fucose-sensitive hemagglutinin. All nonmotile mutants examined, including those with insertions in a sequence homologous to motB, exhibited increased expression of TCP pilin, cholera toxin, and cell-associated hemolysin but dramatically decreased levels of fucose-sensitive hemagglutinin and HEp-2 adhesins. In general, nonmotile mutants displayed few or no defects in intestinal colonization, while class I hypermotile mutants were highly defective in colonization. These results suggest that the motility phenotype of V. cholerae is tightly coupled to the expression of multiple ToxR-regulated and non-ToxR-regulated virulence determinants.

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Selected References

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  1. Akerley B. J., Cotter P. A., Miller J. F. Ectopic expression of the flagellar regulon alters development of the Bordetella-host interaction. Cell. 1995 Feb 24;80(4):611–620. doi: 10.1016/0092-8674(95)90515-4. [DOI] [PubMed] [Google Scholar]
  2. Alm R. A., Stroeher U. H., Manning P. A. Extracellular proteins of Vibrio cholerae: nucleotide sequence of the structural gene (hlyA) for the haemolysin of the haemolytic El Tor strain 017 and characterization of the hlyA mutation in the non-haemolytic classical strain 569B. Mol Microbiol. 1988 Jul;2(4):481–488. doi: 10.1111/j.1365-2958.1988.tb00054.x. [DOI] [PubMed] [Google Scholar]
  3. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  4. Armstrong J. B., Adler J., Dahl M. M. Nonchemotactic mutants of Escherichia coli. J Bacteriol. 1967 Jan;93(1):390–398. doi: 10.1128/jb.93.1.390-398.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Attridge S. R., Rowley D. The role of the flagellum in the adherence of Vibrio cholerae. J Infect Dis. 1983 May;147(5):864–872. doi: 10.1093/infdis/147.5.864. [DOI] [PubMed] [Google Scholar]
  6. Beattie D. T., Knapp S., Mekalanos J. J. Evidence that modulation requires sequences downstream of the promoters of two vir-repressed genes of Bordetella pertussis. J Bacteriol. 1990 Dec;172(12):6997–7004. doi: 10.1128/jb.172.12.6997-7004.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Blair D. F., Kim D. Y., Berg H. C. Mutant MotB proteins in Escherichia coli. J Bacteriol. 1991 Jul;173(13):4049–4055. doi: 10.1128/jb.173.13.4049-4055.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  9. Butterton J. R., Stoebner J. A., Payne S. M., Calderwood S. B. Cloning, sequencing, and transcriptional regulation of viuA, the gene encoding the ferric vibriobactin receptor of Vibrio cholerae. J Bacteriol. 1992 Jun;174(11):3729–3738. doi: 10.1128/jb.174.11.3729-3738.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Coster T. S., Killeen K. P., Waldor M. K., Beattie D. T., Spriggs D. R., Kenner J. R., Trofa A., Sadoff J. C., Mekalanos J. J., Taylor D. N. Safety, immunogenicity, and efficacy of live attenuated Vibrio cholerae O139 vaccine prototype. Lancet. 1995 Apr 15;345(8955):949–952. doi: 10.1016/s0140-6736(95)90698-3. [DOI] [PubMed] [Google Scholar]
  11. DiRita V. J., Mekalanos J. J. Periplasmic interaction between two membrane regulatory proteins, ToxR and ToxS, results in signal transduction and transcriptional activation. Cell. 1991 Jan 11;64(1):29–37. doi: 10.1016/0092-8674(91)90206-e. [DOI] [PubMed] [Google Scholar]
  12. DiRita V. J., Parsot C., Jander G., Mekalanos J. J. Regulatory cascade controls virulence in Vibrio cholerae. Proc Natl Acad Sci U S A. 1991 Jun 15;88(12):5403–5407. doi: 10.1073/pnas.88.12.5403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Everiss K. D., Hughes K. J., Kovach M. E., Peterson K. M. The Vibrio cholerae acfB colonization determinant encodes an inner membrane protein that is related to a family of signal-transducing proteins. Infect Immun. 1994 Aug;62(8):3289–3298. doi: 10.1128/iai.62.8.3289-3298.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Finkelstein R. A., Boesman-Finkelstein M., Chang Y., Häse C. C. Vibrio cholerae hemagglutinin/protease, colonial variation, virulence, and detachment. Infect Immun. 1992 Feb;60(2):472–478. doi: 10.1128/iai.60.2.472-478.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Freter R., Jones G. W. Adhesive properties of Vibrio cholerae: nature of the interaction with intact mucosal surfaces. Infect Immun. 1976 Jul;14(1):246–256. doi: 10.1128/iai.14.1.246-256.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Freter R., O'Brien P. C., Macsai M. S. Role of chemotaxis in the association of motile bacteria with intestinal mucosa: in vivo studies. Infect Immun. 1981 Oct;34(1):234–240. doi: 10.1128/iai.34.1.234-240.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Freter R., O'Brien P. C. Role of chemotaxis in the association of motile bacteria with intestinal mucosa: chemotactic responses of Vibrio cholerae and description of motile nonchemotactic mutants. Infect Immun. 1981 Oct;34(1):215–221. doi: 10.1128/iai.34.1.215-221.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gardel C. L., Mekalanos J. J. Regulation of cholera toxin by temperature, pH, and osmolarity. Methods Enzymol. 1994;235:517–526. doi: 10.1016/0076-6879(94)35167-8. [DOI] [PubMed] [Google Scholar]
  19. Gish W., States D. J. Identification of protein coding regions by database similarity search. Nat Genet. 1993 Mar;3(3):266–272. doi: 10.1038/ng0393-266. [DOI] [PubMed] [Google Scholar]
  20. Grundy F. J., Waters D. A., Takova T. Y., Henkin T. M. Identification of genes involved in utilization of acetate and acetoin in Bacillus subtilis. Mol Microbiol. 1993 Oct;10(2):259–271. doi: 10.1111/j.1365-2958.1993.tb01952.x. [DOI] [PubMed] [Google Scholar]
  21. Guentzel M. N., Berry L. J. Motility as a virulence factor for Vibrio cholerae. Infect Immun. 1975 May;11(5):890–897. doi: 10.1128/iai.11.5.890-897.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Harkey C. W., Everiss K. D., Peterson K. M. The Vibrio cholerae toxin-coregulated-pilus gene tcpI encodes a homolog of methyl-accepting chemotaxis proteins. Infect Immun. 1994 Jul;62(7):2669–2678. doi: 10.1128/iai.62.7.2669-2678.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Herrington D. A., Hall R. H., Losonsky G., Mekalanos J. J., Taylor R. K., Levine M. M. Toxin, toxin-coregulated pili, and the toxR regulon are essential for Vibrio cholerae pathogenesis in humans. J Exp Med. 1988 Oct 1;168(4):1487–1492. doi: 10.1084/jem.168.4.1487. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Holmgren J., Svennerholm A. M. Mechanisms of disease and immunity in cholera: a review. J Infect Dis. 1977 Aug;136 (Suppl):S105–S112. doi: 10.1093/infdis/136.supplement.s105. [DOI] [PubMed] [Google Scholar]
  25. Hueck C., Kraus A., Hillen W. Sequences of ccpA and two downstream Bacillus megaterium genes with homology to the motAB operon from Bacillus subtilis. Gene. 1994 May 27;143(1):147–148. doi: 10.1016/0378-1119(94)90621-1. [DOI] [PubMed] [Google Scholar]
  26. Häse C. C., Finkelstein R. A. Cloning and nucleotide sequence of the Vibrio cholerae hemagglutinin/protease (HA/protease) gene and construction of an HA/protease-negative strain. J Bacteriol. 1991 Jun;173(11):3311–3317. doi: 10.1128/jb.173.11.3311-3317.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Jones G. W., Freter R. Adhesive properties of Vibrio cholerae: nature of the interaction with isolated rabbit brush border membranes and human erythrocytes. Infect Immun. 1976 Jul;14(1):240–245. doi: 10.1128/iai.14.1.240-245.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Jonson G., Sanchez J., Svennerholm A. M. Expression and detection of different biotype-associated cell-bound haemagglutinins of Vibrio cholerae O1. J Gen Microbiol. 1989 Jan;135(1):111–120. doi: 10.1099/00221287-135-1-111. [DOI] [PubMed] [Google Scholar]
  29. Jonson G., Svennerholm A. M., Holmgren J. Vibrio cholerae expresses cell surface antigens during intestinal infection which are not expressed during in vitro culture. Infect Immun. 1989 Jun;57(6):1809–1815. doi: 10.1128/iai.57.6.1809-1815.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Kenner J. R., Coster T. S., Taylor D. N., Trofa A. F., Barrera-Oro M., Hyman T., Adams J. M., Beattie D. T., Killeen K. P., Spriggs D. R. Peru-15, an improved live attenuated oral vaccine candidate for Vibrio cholerae O1. J Infect Dis. 1995 Oct;172(4):1126–1129. doi: 10.1093/infdis/172.4.1126. [DOI] [PubMed] [Google Scholar]
  31. Kroos L., Kaiser D. Construction of Tn5 lac, a transposon that fuses lacZ expression to exogenous promoters, and its introduction into Myxococcus xanthus. Proc Natl Acad Sci U S A. 1984 Sep;81(18):5816–5820. doi: 10.1073/pnas.81.18.5816. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Lee C. A., Falkow S. The ability of Salmonella to enter mammalian cells is affected by bacterial growth state. Proc Natl Acad Sci U S A. 1990 Jun;87(11):4304–4308. doi: 10.1073/pnas.87.11.4304. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Mekalanos J. J., Collier R. J., Romig W. R. Enzymic activity of cholera toxin. II. Relationships to proteolytic processing, disulfide bond reduction, and subunit composition. J Biol Chem. 1979 Jul 10;254(13):5855–5861. [PubMed] [Google Scholar]
  34. Mekalanos J. J. Live attenuated vaccine vectors. Int J Technol Assess Health Care. 1994 Winter;10(1):131–142. doi: 10.1017/s0266462300014057. [DOI] [PubMed] [Google Scholar]
  35. Mekalanos J. J., Swartz D. J., Pearson G. D., Harford N., Groyne F., de Wilde M. Cholera toxin genes: nucleotide sequence, deletion analysis and vaccine development. Nature. 1983 Dec 8;306(5943):551–557. doi: 10.1038/306551a0. [DOI] [PubMed] [Google Scholar]
  36. Miller S. I., Kukral A. M., Mekalanos J. J. A two-component regulatory system (phoP phoQ) controls Salmonella typhimurium virulence. Proc Natl Acad Sci U S A. 1989 Jul;86(13):5054–5058. doi: 10.1073/pnas.86.13.5054. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Miller V. L., DiRita V. J., Mekalanos J. J. Identification of toxS, a regulatory gene whose product enhances toxR-mediated activation of the cholera toxin promoter. J Bacteriol. 1989 Mar;171(3):1288–1293. doi: 10.1128/jb.171.3.1288-1293.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Miller V. L., Mekalanos J. J. 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. 1988 Jun;170(6):2575–2583. doi: 10.1128/jb.170.6.2575-2583.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Miller V. L., Taylor R. K., Mekalanos J. J. Cholera toxin transcriptional activator toxR is a transmembrane DNA binding protein. Cell. 1987 Jan 30;48(2):271–279. doi: 10.1016/0092-8674(87)90430-2. [DOI] [PubMed] [Google Scholar]
  40. Mostow P., Richardson K. High-frequency spontaneous mutation of classical Vibrio cholerae to a nonmotile phenotype. Infect Immun. 1990 Nov;58(11):3633–3639. doi: 10.1128/iai.58.11.3633-3639.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Ottemann K. M., DiRita V. J., Mekalanos J. J. ToxR proteins with substitutions in residues conserved with OmpR fail to activate transcription from the cholera toxin promoter. J Bacteriol. 1992 Nov;174(21):6807–6814. doi: 10.1128/jb.174.21.6807-6814.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Peterson K. M., Mekalanos J. J. Characterization of the Vibrio cholerae ToxR regulon: identification of novel genes involved in intestinal colonization. Infect Immun. 1988 Nov;56(11):2822–2829. doi: 10.1128/iai.56.11.2822-2829.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Richardson K. Roles of motility and flagellar structure in pathogenicity of Vibrio cholerae: analysis of motility mutants in three animal models. Infect Immun. 1991 Aug;59(8):2727–2736. doi: 10.1128/iai.59.8.2727-2736.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Stader J., Matsumura P., Vacante D., Dean G. E., Macnab R. M. Nucleotide sequence of the Escherichia coli motB gene and site-limited incorporation of its product into the cytoplasmic membrane. J Bacteriol. 1986 Apr;166(1):244–252. doi: 10.1128/jb.166.1.244-252.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Stibitz S., Aaronson W., Monack D., Falkow S. Phase variation in Bordetella pertussis by frameshift mutation in a gene for a novel two-component system. Nature. 1989 Mar 16;338(6212):266–269. doi: 10.1038/338266a0. [DOI] [PubMed] [Google Scholar]
  46. Taylor R. K., Miller V. L., Furlong D. B., Mekalanos J. J. Use of phoA gene fusions to identify a pilus colonization factor coordinately regulated with cholera toxin. Proc Natl Acad Sci U S A. 1987 May;84(9):2833–2837. doi: 10.1073/pnas.84.9.2833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Teppema J. S., Guinée P. A., Ibrahim A. A., Pâques M., Ruitenberg E. J. In vivo adherence and colonization of Vibrio cholerae strains that differ in hemagglutinating activity and motility. Infect Immun. 1987 Sep;55(9):2093–2102. doi: 10.1128/iai.55.9.2093-2102.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Waldor M. K., Colwell R., Mekalanos J. J. The Vibrio cholerae O139 serogroup antigen includes an O-antigen capsule and lipopolysaccharide virulence determinants. Proc Natl Acad Sci U S A. 1994 Nov 22;91(24):11388–11392. doi: 10.1073/pnas.91.24.11388. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Yamamoto T., Kamano T., Uchimura M., Iwanaga M., Yokota T. Vibrio cholerae O1 adherence to villi and lymphoid follicle epithelium: in vitro model using formalin-treated human small intestine and correlation between adherence and cell-associated hemagglutinin levels. Infect Immun. 1988 Dec;56(12):3241–3250. doi: 10.1128/iai.56.12.3241-3250.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Yancey R. J., Willis D. L., Berry L. J. Flagella-induced immunity against experimental cholera in adult rabbits. Infect Immun. 1979 Jul;25(1):220–228. doi: 10.1128/iai.25.1.220-228.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]

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