Skip to main content
PMC home page
Applied and Environmental Microbiology logoLink to Applied and Environmental Microbiology
. 1997 Jun;63(6):2372–2377. doi: 10.1128/aem.63.6.2372-2377.1997

Effect of bile on Vibrio parahaemolyticus.

J L Pace 1, T J Chai 1, H A Rossi 1, X Jiang 1
PMCID: PMC168531  PMID: 9172358

Abstract

Many enteric pathogens are thought to enter a viable but nonculturable state when deprived of nutrients. Virulent strains of the enteric pathogen Vibrio parahaemolyticus are rarely isolated from their low-nutrient aquatic environments, possibly due to their nonculturability. Host factors such as bile may trigger release from dormancy and increase virulence in these strains. In this study, the addition of bile or the bile acid deoxycholic acid to estuarine water-cultured bacteria led to an increase in the direct viable count and colony counts among the virulent strains. This effect was not demonstrated in the nonvirulent strains, and it was reversed by extraction of bile acids with cholestyramine. Bile-treated V. parahaemolyticus had lower levels of intracellular calcium than untreated cells, and this effect coincided with an increase in the number of metabolically active cells. Chelation of intracellular calcium with BAPTA/AM (R. Y. Tsien, Biochemistry 19:2396-2402, 1980) produced similar results. Addition of bile to V. parahaemolyticus cultures in laboratory medium enhanced factors associated with virulence such as Congo red binding, bacterial capsule size, and adherence to epithelial cells. These results suggest that a bile acid-containing environment such as that found in the human host favors growth of virulent strains of V. parahaemolyticus and that bile acids enhance the expression of virulence factors. These effects seem to be mediated by a decrease in intracellular calcium.

Full Text

The Full Text of this article is available as a PDF (174.4 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. AMES B. N., DUBIN D. T. The role of polyamines in the neutralization of bacteriophage deoxyribonucleic acid. J Biol Chem. 1960 Mar;235:769–775. [PubMed] [Google Scholar]
  2. Andrews G. P., Maurelli A. T. mxiA of Shigella flexneri 2a, which facilitates export of invasion plasmid antigens, encodes a homolog of the low-calcium-response protein, LcrD, of Yersinia pestis. Infect Immun. 1992 Aug;60(8):3287–3295. doi: 10.1128/iai.60.8.3287-3295.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. BLIGH E. G., DYER W. J. A rapid method of total lipid extraction and purification. Can J Biochem Physiol. 1959 Aug;37(8):911–917. doi: 10.1139/o59-099. [DOI] [PubMed] [Google Scholar]
  4. Borgström B. Bile salts--their physiological functions in the gastrointestinal tract. Acta Med Scand. 1974 Jul-Aug;196(1-2):1–10. doi: 10.1111/j.0954-6820.1974.tb00958.x. [DOI] [PubMed] [Google Scholar]
  5. Brubaker R. R. The Vwa+ virulence factor of yersiniae: the molecular basis of the attendant nutritional requirement for Ca++. Rev Infect Dis. 1983 Sep-Oct;5 (Suppl 4):S748–S758. doi: 10.1093/clinids/5.supplement_4.s748. [DOI] [PubMed] [Google Scholar]
  6. Bölin I., Wolf-Watz H. The plasmid-encoded Yop2b protein of Yersinia pseudotuberculosis is a virulence determinant regulated by calcium and temperature at the level of transcription. Mol Microbiol. 1988 Mar;2(2):237–245. doi: 10.1111/j.1365-2958.1988.tb00025.x. [DOI] [PubMed] [Google Scholar]
  7. Carruthers M. M. In vitro adherence of Kanagawa-positive Vibrio parahaemolyticus to epithelial cells. J Infect Dis. 1977 Oct;136(4):588–592. doi: 10.1093/infdis/136.4.588. [DOI] [PubMed] [Google Scholar]
  8. Chai T. J. Characteristics of Escherichia coli grown in bay water as compared with rich medium. Appl Environ Microbiol. 1983 Apr;45(4):1316–1323. doi: 10.1128/aem.45.4.1316-1323.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chakrabarti M. K., Sinha A. K., Biswas T. Adherence of Vibrio parahaemolyticus to rabbit intestinal epithelial cells in vitro. FEMS Microbiol Lett. 1991 Nov 1;68(1):113–117. doi: 10.1016/0378-1097(91)90405-y. [DOI] [PubMed] [Google Scholar]
  10. Costerton J. W., Irvin R. T., Cheng K. J. The role of bacterial surface structures in pathogenesis. Crit Rev Microbiol. 1981;8(4):303–338. doi: 10.3109/10408418109085082. [DOI] [PubMed] [Google Scholar]
  11. D'Mello A., Yotis W. W. The action of sodium deoxycholate on Escherichia coli. Appl Environ Microbiol. 1987 Aug;53(8):1944–1946. doi: 10.1128/aem.53.8.1944-1946.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Daskaleros P. A., Payne S. M. Congo red binding phenotype is associated with hemin binding and increased infectivity of Shigella flexneri in the HeLa cell model. Infect Immun. 1987 Jun;55(6):1393–1398. doi: 10.1128/iai.55.6.1393-1398.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dazzo F. B., Brill W. J. Bacterial polysaccharide which binds Rhizobium trifolii to clover root hairs. J Bacteriol. 1979 Mar;137(3):1362–1373. doi: 10.1128/jb.137.3.1362-1373.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fitz-James P. C. Formation of protoplasts from resting spores. J Bacteriol. 1971 Mar;105(3):1119–1136. doi: 10.1128/jb.105.3.1119-1136.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Forsberg A., Rosqvist R. In vivo expression of virulence genes of Yersinia pseudotuberculosis. Infect Agents Dis. 1993 Aug;2(4):275–278. [PubMed] [Google Scholar]
  16. Galán J. E., Curtiss R., 3rd Cloning and molecular characterization of genes whose products allow Salmonella typhimurium to penetrate tissue culture cells. Proc Natl Acad Sci U S A. 1989 Aug;86(16):6383–6387. doi: 10.1073/pnas.86.16.6383. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gangola P., Rosen B. P. Fura-2 measurements of intracellular [Ca2+] in Escherichia coli. Prog Clin Biol Res. 1988;252:215–220. [PubMed] [Google Scholar]
  18. Gangola P., Rosen B. P. Maintenance of intracellular calcium in Escherichia coli. J Biol Chem. 1987 Sep 15;262(26):12570–12574. [PubMed] [Google Scholar]
  19. Gingras S. P., Howard L. V. Adherence of Vibrio parahaemolyticus to human epithelial cell lines. Appl Environ Microbiol. 1980 Feb;39(2):369–371. doi: 10.1128/aem.39.2.369-371.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Grimes D. J., Atwell R. W., Brayton P. R., Palmer L. M., Rollins D. M., Roszak D. B., Singleton F. L., Tamplin M. L., Colwell R. R. The fate of enteric pathogenic bacteria in estuarine and marine environments. Microbiol Sci. 1986 Nov;3(11):324–329. [PubMed] [Google Scholar]
  21. Grynkiewicz G., Poenie M., Tsien R. Y. A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem. 1985 Mar 25;260(6):3440–3450. [PubMed] [Google Scholar]
  22. HORAN J. M., DILUZIO N. R., ETTELDORF J. N. USE OF AN ANION EXCHANGE RESIN IN TREATMENT OF TWO SIBLINGS WITH FAMILIAL HYPERCHOLESTEROLEMIA. J Pediatr. 1964 Feb;64:201–209. doi: 10.1016/s0022-3476(64)80263-8. [DOI] [PubMed] [Google Scholar]
  23. Hofmann A. F., Small D. M. Detergent properties of bile salts: correlation with physiological function. Annu Rev Med. 1967;18:333–376. doi: 10.1146/annurev.me.18.020167.002001. [DOI] [PubMed] [Google Scholar]
  24. Janda J. M., Powers C., Bryant R. G., Abbott S. L. Current perspectives on the epidemiology and pathogenesis of clinically significant Vibrio spp. Clin Microbiol Rev. 1988 Jul;1(3):245–267. doi: 10.1128/cmr.1.3.245. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Jann K., Jann B. Capsules of Escherichia coli, expression and biological significance. Can J Microbiol. 1992 Jul;38(7):705–710. doi: 10.1139/m92-116. [DOI] [PubMed] [Google Scholar]
  26. Jones D. M., Sutcliffe E. M., Curry A. Recovery of viable but non-culturable Campylobacter jejuni. J Gen Microbiol. 1991 Oct;137(10):2477–2482. doi: 10.1099/00221287-137-10-2477. [DOI] [PubMed] [Google Scholar]
  27. Joseph S. W., Colwell R. R., Kaper J. B. Vibrio parahaemolyticus and related halophilic Vibrios. Crit Rev Microbiol. 1982;10(1):77–124. doi: 10.3109/10408418209113506. [DOI] [PubMed] [Google Scholar]
  28. Kjelleberg S., Albertson N., Flärdh K., Holmquist L., Jouper-Jaan A., Marouga R., Ostling J., Svenblad B., Weichart D. How do non-differentiating bacteria adapt to starvation? Antonie Van Leeuwenhoek. 1993;63(3-4):333–341. doi: 10.1007/BF00871228. [DOI] [PubMed] [Google Scholar]
  29. Kogure K., Simidu U., Taga N. A tentative direct microscopic method for counting living marine bacteria. Can J Microbiol. 1979 Mar;25(3):415–420. doi: 10.1139/m79-063. [DOI] [PubMed] [Google Scholar]
  30. Laoudj D., Andersen C. L., Bras A., Goldberg M., Jacq A., Holland I. B. EGTA induces the synthesis in Escherichia coli of three proteins that cross-react with calmodulin antibodies. Mol Microbiol. 1994 Aug;13(3):445–457. doi: 10.1111/j.1365-2958.1994.tb00439.x. [DOI] [PubMed] [Google Scholar]
  31. Lawlor K. M., Daskaleros P. A., Robinson R. E., Payne S. M. Virulence of iron transport mutants of Shigella flexneri and utilization of host iron compounds. Infect Immun. 1987 Mar;55(3):594–599. doi: 10.1128/iai.55.3.594-599.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Matin A., Auger E. A., Blum P. H., Schultz J. E. Genetic basis of starvation survival in nondifferentiating bacteria. Annu Rev Microbiol. 1989;43:293–316. doi: 10.1146/annurev.mi.43.100189.001453. [DOI] [PubMed] [Google Scholar]
  33. Maurelli A. T., Hromockyj A. E., Bernardini M. L. Environmental regulation of Shigella virulence. Curr Top Microbiol Immunol. 1992;180:95–116. doi: 10.1007/978-3-642-77238-2_5. [DOI] [PubMed] [Google Scholar]
  34. Mekalanos J. J. Environmental signals controlling expression of virulence determinants in bacteria. J Bacteriol. 1992 Jan;174(1):1–7. doi: 10.1128/jb.174.1.1-7.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Miller J. F., Mekalanos J. J., Falkow S. Coordinate regulation and sensory transduction in the control of bacterial virulence. Science. 1989 Feb 17;243(4893):916–922. doi: 10.1126/science.2537530. [DOI] [PubMed] [Google Scholar]
  36. Miyamoto Y., Kato T., Obara Y., Akiyama S., Takizawa K., Yamai S. In vitro hemolytic characteristic of Vibrio parahaemolyticus: its close correlation with human pathogenicity. J Bacteriol. 1969 Nov;100(2):1147–1149. doi: 10.1128/jb.100.2.1147-1149.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Nishibuchi M., Kaper J. B. Thermostable direct hemolysin gene of Vibrio parahaemolyticus: a virulence gene acquired by a marine bacterium. Infect Immun. 1995 Jun;63(6):2093–2099. doi: 10.1128/iai.63.6.2093-2099.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Norris V., Chen M., Goldberg M., Voskuil J., McGurk G., Holland I. B. Calcium in bacteria: a solution to which problem? Mol Microbiol. 1991 Apr;5(4):775–778. doi: 10.1111/j.1365-2958.1991.tb00748.x. [DOI] [PubMed] [Google Scholar]
  39. O'Hara M. B., Hageman J. H. Energy and calcium ion dependence of proteolysis during sporulation of Bacillus subtilis cells. J Bacteriol. 1990 Aug;172(8):4161–4170. doi: 10.1128/jb.172.8.4161-4170.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. POSTGATE J. R., HUNTER J. R. The survival of starved bacteria. J Gen Microbiol. 1962 Oct;29:233–263. doi: 10.1099/00221287-29-2-233. [DOI] [PubMed] [Google Scholar]
  41. Pace J., Chai T. J. Comparison of Vibrio parahaemolyticus grown in estuarine water and rich medium. Appl Environ Microbiol. 1989 Aug;55(8):1877–1887. doi: 10.1128/aem.55.8.1877-1887.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Payne S. M., Finkelstein R. A. Detection and differentiation of iron-responsive avirulent mutants on Congo red agar. Infect Immun. 1977 Oct;18(1):94–98. doi: 10.1128/iai.18.1.94-98.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Plésiat P., Nikaido H. Outer membranes of gram-negative bacteria are permeable to steroid probes. Mol Microbiol. 1992 May;6(10):1323–1333. doi: 10.1111/j.1365-2958.1992.tb00853.x. [DOI] [PubMed] [Google Scholar]
  44. Pope L. M., Reed K. E., Payne S. M. Increased protein secretion and adherence to HeLa cells by Shigella spp. following growth in the presence of bile salts. Infect Immun. 1995 Sep;63(9):3642–3648. doi: 10.1128/iai.63.9.3642-3648.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. RODE L. J., FOSTER J. W. Germination of bacterial spores with alkyl primary amines. J Bacteriol. 1961 May;81:768–779. doi: 10.1128/jb.81.5.768-779.1961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. RODE L. J., FOSTER J. W. The action of surfactants on bacterial spores. Arch Mikrobiol. 1960;36:67–94. doi: 10.1007/BF00405943. [DOI] [PubMed] [Google Scholar]
  47. Rajagopalan N., Lindenbaum S. The binding of Ca2+ to taurine and glycine-conjugated bile salt micelles. Biochim Biophys Acta. 1982 Apr 15;711(1):66–74. doi: 10.1016/0005-2760(82)90010-8. [DOI] [PubMed] [Google Scholar]
  48. Roszak D. B., Colwell R. R. Survival strategies of bacteria in the natural environment. Microbiol Rev. 1987 Sep;51(3):365–379. doi: 10.1128/mr.51.3.365-379.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Roszak D. B., Grimes D. J., Colwell R. R. Viable but nonrecoverable stage of Salmonella enteritidis in aquatic systems. Can J Microbiol. 1984 Mar;30(3):334–338. doi: 10.1139/m84-049. [DOI] [PubMed] [Google Scholar]
  50. Sanyal A. J., Shiffmann M. L., Hirsch J. I., Moore E. W. Premicellar taurocholate enhances ferrous iron uptake from all regions of rat small intestine. Gastroenterology. 1991 Aug;101(2):382–389. doi: 10.1016/0016-5085(91)90015-d. [DOI] [PubMed] [Google Scholar]
  51. Shi S. P., Chang C. C., Gould G. W., Chang T. Y. Comparison of phosphatidylethanolamine and phosphatidylcholine vesicles produced by treating cholate-phospholipid micelles with cholestyramine. Biochim Biophys Acta. 1989 Jul 10;982(2):187–195. doi: 10.1016/0005-2736(89)90054-0. [DOI] [PubMed] [Google Scholar]
  52. Straley S. C., Plano G. V., Skrzypek E., Haddix P. L., Fields K. A. Regulation by Ca2+ in the Yersinia low-Ca2+ response. Mol Microbiol. 1993 Jun;8(6):1005–1010. doi: 10.1111/j.1365-2958.1993.tb01644.x. [DOI] [PubMed] [Google Scholar]
  53. Sudo S. Z., Dworkin M. Comparative biology of prokaryotic resting cells. Adv Microb Physiol. 1973;9:153–224. doi: 10.1016/s0065-2911(08)60378-1. [DOI] [PubMed] [Google Scholar]
  54. Swevers L., Lambert J. G., De Loof A. Synthesis and metabolism of vertebrate-type steroids by tissues of insects: a critical evaluation. Experientia. 1991 Jul 15;47(7):687–698. doi: 10.1007/BF01958817. [DOI] [PubMed] [Google Scholar]
  55. Tisa L. S., Adler J. Cytoplasmic free-Ca2+ level rises with repellents and falls with attractants in Escherichia coli chemotaxis. Proc Natl Acad Sci U S A. 1995 Nov 7;92(23):10777–10781. doi: 10.1073/pnas.92.23.10777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Tsien R. Y. New calcium indicators and buffers with high selectivity against magnesium and protons: design, synthesis, and properties of prototype structures. Biochemistry. 1980 May 27;19(11):2396–2404. doi: 10.1021/bi00552a018. [DOI] [PubMed] [Google Scholar]
  57. Twedt R. M., Peeler J. T., Spaulding P. L. Effective ileal loop dose of Kanagawa-positive Vibrio parahaemolyticus. Appl Environ Microbiol. 1980 Dec;40(6):1012–1016. doi: 10.1128/aem.40.6.1012-1016.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. 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]
  59. Wilson G. R., Benoit T. G. Activation and germination of Bacillus thuringiensis spores in Manduca sexta larval gut fluid. J Invertebr Pathol. 1990 Sep;56(2):233–236. doi: 10.1016/0022-2011(90)90105-f. [DOI] [PubMed] [Google Scholar]
  60. Yamamoto T., Albert M. J., Sack R. B. Adherence to human small intestines of capsulated Vibrio cholerae O139. FEMS Microbiol Lett. 1994 Jun 1;119(1-2):229–235. doi: 10.1111/j.1574-6968.1994.tb06893.x. [DOI] [PubMed] [Google Scholar]
  61. Yamamoto T., Yokota T. Adherence targets of Vibrio parahaemolyticus in human small intestines. Infect Immun. 1989 Aug;57(8):2410–2419. doi: 10.1128/iai.57.8.2410-2419.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Applied and Environmental Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES