Skip to main content
PMC home page
Infection and Immunity logoLink to Infection and Immunity
. 1990 Aug;58(8):2420–2428. doi: 10.1128/iai.58.8.2420-2428.1990

Oral immunization of mice with a live recombinant Yersinia enterocolitica O:9 strain that produces the cholera toxin B subunit.

M P Sory 1, P Hermand 1, J P Vaerman 1, G R Cornelis 1
PMCID: PMC258836  PMID: 2370100

Abstract

The 70-kilobase pYV plasmid of Yersinia enterocolitica encodes a set of proteins called Yops that are produced during infection. To use Y. enterocolitica as a live carrier to present the cholera toxin B (CT-B) subunit to the immune system, we constructed an operon fusion between ctxB and the yop51 gene. This operon fusion was either cloned on an RSF1010-derived plasmid or integrated into the pYV plasmid itself. In Y. enterocolitica, both constructions directed the synthesis of free CT-B only under conditions of Yops production, i.e., at 37 degrees C in a medium deprived of Ca2+. Bacteria containing both types of recombinant plasmids were given orally to mice. A serum antibody response against CT-B was detected in both cases. A secretory immunoglobulin A activity specific to CT-B was also observed in the intestinal secretions. According to immunoblot analysis, the serum antibody response was only directed against the polymeric form of the B subunit. The ctxB gene was also inserted in frame within yop51, giving a chimeric Yop51-CT-B protein that was secreted into the surrounding medium. In this case, however, no antibody response was observed after oral inoculation of mice. This lack of response probably results from the inability of the hybrid protein to assemble into the polymeric form of the B subunit.

Full text

PDF
2421

Images in this article

2424Image
on p.2424
2425Image
on p.2425
2425Image
on p.2425

Selected References

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

  1. Attwood S. E., Cafferkey M. T., Keane F. B. Yersinia infections in surgical practice. Br J Surg. 1989 May;76(5):499–504. doi: 10.1002/bjs.1800760523. [DOI] [PubMed] [Google Scholar]
  2. Bakour R., Balligand G., Laroche Y., Cornelis G., Wauters G. A simple adult-mouse test for tissue invasiveness in Yersinia enterocolitica strains of low experimental virulence. J Med Microbiol. 1985 Apr;19(2):237–246. doi: 10.1099/00222615-19-2-237. [DOI] [PubMed] [Google Scholar]
  3. Biot T., Cornelis G. R. The replication, partition and yop regulation of the pYV plasmids are highly conserved in Yersinia enterocolitica and Y. pseudotuberculosis. J Gen Microbiol. 1988 Jun;134(6):1525–1534. doi: 10.1099/00221287-134-6-1525. [DOI] [PubMed] [Google Scholar]
  4. Bowe F., O'Gaora P., Maskell D., Cafferkey M., Dougan G. Virulence, persistence, and immunogenicity of Yersinia enterocolitica O:8 aroA mutants. Infect Immun. 1989 Oct;57(10):3234–3236. doi: 10.1128/iai.57.10.3234-3236.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brown A., Hormaeche C. E., Demarco de Hormaeche R., Winther M., Dougan G., Maskell D. J., Stocker B. A. An attenuated aroA Salmonella typhimurium vaccine elicits humoral and cellular immunity to cloned beta-galactosidase in mice. J Infect Dis. 1987 Jan;155(1):86–92. doi: 10.1093/infdis/155.1.86. [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. Clements J. D., Lyon F. L., Lowe K. L., Farrand A. L., el-Morshidy S. Oral immunization of mice with attenuated Salmonella enteritidis containing a recombinant plasmid which codes for production of the B subunit of heat-labile Escherichia coli enterotoxin. Infect Immun. 1986 Sep;53(3):685–692. doi: 10.1128/iai.53.3.685-692.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cornelis G. R., Biot T., Lambert de Rouvroit C., Michiels T., Mulder B., Sluiters C., Sory M. P., Van Bouchaute M., Vanooteghem J. C. The Yersinia yop regulon. Mol Microbiol. 1989 Oct;3(10):1455–1459. doi: 10.1111/j.1365-2958.1989.tb00129.x. [DOI] [PubMed] [Google Scholar]
  9. Cornelis G., Colson C. Restriction of DNA in Yersinia enterocolitica detected by recipient ability for a derepressed R factor from Escherichia coli. J Gen Microbiol. 1975 Apr;87(2):285–291. doi: 10.1099/00221287-87-2-285. [DOI] [PubMed] [Google Scholar]
  10. Cornelis G., Laroche Y., Balligand G., Sory M. P., Wauters G. Yersinia enterocolitica, a primary model for bacterial invasiveness. Rev Infect Dis. 1987 Jan-Feb;9(1):64–87. doi: 10.1093/clinids/9.1.64. [DOI] [PubMed] [Google Scholar]
  11. Cornelis G., Sluiters C., de Rouvroit C. L., Michiels T. Homology between virF, the transcriptional activator of the Yersinia virulence regulon, and AraC, the Escherichia coli arabinose operon regulator. J Bacteriol. 1989 Jan;171(1):254–262. doi: 10.1128/jb.171.1.254-262.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Cornelis G., Sory M. P., Laroche Y., Derclaye I. Genetic analysis of the plasmid region controlling virulence in Yersinia enterocolitica 0:9 by Mini-Mu insertions and lac gene fusions. Microb Pathog. 1986 Aug;1(4):349–359. doi: 10.1016/0882-4010(86)90067-7. [DOI] [PubMed] [Google Scholar]
  13. Cornelis G., Vanootegem J. C., Sluiters C. Transcription of the yop regulon from Y. enterocolitica requires trans acting pYV and chromosomal genes. Microb Pathog. 1987 May;2(5):367–379. doi: 10.1016/0882-4010(87)90078-7. [DOI] [PubMed] [Google Scholar]
  14. Curtiss R., 3rd, Kelly S. M., Gulig P. A., Nakayama K. Selective delivery of antigens by recombinant bacteria. Curr Top Microbiol Immunol. 1989;146:35–49. doi: 10.1007/978-3-642-74529-4_4. [DOI] [PubMed] [Google Scholar]
  15. Curtiss R., 3rd, Kelly S. M. Salmonella typhimurium deletion mutants lacking adenylate cyclase and cyclic AMP receptor protein are avirulent and immunogenic. Infect Immun. 1987 Dec;55(12):3035–3043. doi: 10.1128/iai.55.12.3035-3043.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Derbyshire K. M., Hatfull G., Willetts N. Mobilization of the non-conjugative plasmid RSF1010: a genetic and DNA sequence analysis of the mobilization region. Mol Gen Genet. 1987 Jan;206(1):161–168. doi: 10.1007/BF00326552. [DOI] [PubMed] [Google Scholar]
  17. Dorman C. J., Chatfield S., Higgins C. F., Hayward C., Dougan G. Characterization of porin and ompR mutants of a virulent strain of Salmonella typhimurium: ompR mutants are attenuated in vivo. Infect Immun. 1989 Jul;57(7):2136–2140. doi: 10.1128/iai.57.7.2136-2140.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Dougan G. Molecular characterization of bacterial virulence factors and the consequences for vaccine design. The 1988 Fleming lecture. J Gen Microbiol. 1989 Jun;135(6):1397–1406. doi: 10.1099/00221287-135-6-1397. [DOI] [PubMed] [Google Scholar]
  19. Elson C. O., Ealding W., Lefkowitz J. A lavage technique allowing repeated measurement of IgA antibody in mouse intestinal secretions. J Immunol Methods. 1984 Feb 24;67(1):101–108. doi: 10.1016/0022-1759(84)90089-9. [DOI] [PubMed] [Google Scholar]
  20. Forsberg A., Wolf-Watz H. The virulence protein Yop5 of Yersinia pseudotuberculosis is regulated at transcriptional level by plasmid-plB1-encoded trans-acting elements controlled by temperature and calcium. Mol Microbiol. 1988 Jan;2(1):121–133. [PubMed] [Google Scholar]
  21. Galán J. E., Curtiss R., 3rd Virulence and vaccine potential of phoP mutants of Salmonella typhimurium. Microb Pathog. 1989 Jun;6(6):433–443. doi: 10.1016/0882-4010(89)90085-5. [DOI] [PubMed] [Google Scholar]
  22. Gennaro M. L., Greenaway P. J., Broadbent D. A. The expression of biologically active cholera toxin in Escherichia coli. Nucleic Acids Res. 1982 Aug 25;10(16):4883–4890. doi: 10.1093/nar/10.16.4883. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Goetzl E. J., Metzger H. Affinity labeling of a mouse myeloma protein which binds nitrophenyl ligands. Kinetics of labeling and isolation of a labeled peptide. Biochemistry. 1970 Mar 3;9(5):1267–1278. doi: 10.1021/bi00807a031. [DOI] [PubMed] [Google Scholar]
  24. Heesemann J., Algermissen B., Laufs R. Genetically manipulated virulence of Yersinia enterocolitica. Infect Immun. 1984 Oct;46(1):105–110. doi: 10.1128/iai.46.1.105-110.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Hoiseth S. K., Stocker B. A. Aromatic-dependent Salmonella typhimurium are non-virulent and effective as live vaccines. Nature. 1981 May 21;291(5812):238–239. doi: 10.1038/291238a0. [DOI] [PubMed] [Google Scholar]
  26. Hone D., Attridge S., van den Bosch L., Hackett J. A chromosomal integration system for stabilization of heterologous genes in Salmonella based vaccine strains. Microb Pathog. 1988 Dec;5(6):407–418. doi: 10.1016/0882-4010(88)90002-2. [DOI] [PubMed] [Google Scholar]
  27. Isberg R. R. Mammalian cell adhesion functions and cellular penetration of enteropathogenic Yersinia species. Mol Microbiol. 1989 Oct;3(10):1449–1453. doi: 10.1111/j.1365-2958.1989.tb00128.x. [DOI] [PubMed] [Google Scholar]
  28. Marsh J. L., Erfle M., Wykes E. J. The pIC plasmid and phage vectors with versatile cloning sites for recombinant selection by insertional inactivation. Gene. 1984 Dec;32(3):481–485. doi: 10.1016/0378-1119(84)90022-2. [DOI] [PubMed] [Google Scholar]
  29. 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]
  30. Michiels T., Cornelis G. Nucleotide sequence and transcription analysis of yop51 from Yersinia enterocolitica W22703. Microb Pathog. 1988 Dec;5(6):449–459. doi: 10.1016/0882-4010(88)90006-x. [DOI] [PubMed] [Google Scholar]
  31. Miller V. L., Falkow S. Evidence for two genetic loci in Yersinia enterocolitica that can promote invasion of epithelial cells. Infect Immun. 1988 May;56(5):1242–1248. doi: 10.1128/iai.56.5.1242-1248.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. 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]
  33. Mulder B., Michiels T., Simonet M., Sory M. P., Cornelis G. Identification of additional virulence determinants on the pYV plasmid of Yersinia enterocolitica W227. Infect Immun. 1989 Aug;57(8):2534–2541. doi: 10.1128/iai.57.8.2534-2541.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Nugent M. E., Bone D. H., Datta N. A transposon, Tn732, encoding gentamicin/tobramycin resistance. Nature. 1979 Nov 22;282(5737):422–423. doi: 10.1038/282422a0. [DOI] [PubMed] [Google Scholar]
  35. Pearson G. D., Mekalanos J. J. Molecular cloning of Vibrio cholerae enterotoxin genes in Escherichia coli K-12. Proc Natl Acad Sci U S A. 1982 May;79(9):2976–2980. doi: 10.1073/pnas.79.9.2976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Pierre P., Langendries A., Vaerman J. P. Cholera toxin neutralization: a comparison of purified serum IgG and biliary secretory IgA antibodies. Immunol Lett. 1988 May;18(1):51–55. doi: 10.1016/0165-2478(88)90069-7. [DOI] [PubMed] [Google Scholar]
  37. Poirier T. P., Kehoe M. A., Beachey E. H. Protective immunity evoked by oral administration of attenuated aroA Salmonella typhimurium expressing cloned streptococcal M protein. J Exp Med. 1988 Jul 1;168(1):25–32. doi: 10.1084/jem.168.1.25. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Sory M. P., Cornelis G. Yersinia enterocolitica O:9 as a potential live oral carrier for protective antigens. Microb Pathog. 1988 Jun;4(6):431–442. doi: 10.1016/0882-4010(88)90028-9. [DOI] [PubMed] [Google Scholar]
  39. Takao T., Tominaga N., Yoshimura S., Shimonishi Y., Hara S., Inoue T., Miyama A. Isolation, primary structure and synthesis of heat-stable enterotoxin produced by Yersinia enterocolitica. Eur J Biochem. 1985 Oct 1;152(1):199–206. doi: 10.1111/j.1432-1033.1985.tb09183.x. [DOI] [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES