Skip to main content
NCBI home page
Infection and Immunity logoLink to Infection and Immunity
. 1993 Nov;61(11):4582–4589. doi: 10.1128/iai.61.11.4582-4589.1993

Novel antigens expressed by Aeromonas salmonicida grown in vivo.

J C Thornton 1, R A Garduño 1, S J Carlos 1, W W Kay 1
PMCID: PMC281208  PMID: 8406855

Abstract

Virulent and avirulent Aeromonas salmonicida strains grown inside intraperitoneal implants in Rainbow trout (Oncorhynchus mykiss) were examined for unique antigen expression. Western blots (immunoblots), performed with immune rabbit serum raised against in vivo-grown cells, revealed several unique antigens. With the exception of lipopolysaccharide (LPS), these novel antigens were destroyed after proteinase K treatment. The majority of these antigens were not induced in vitro in response to either iron limitation or anaerobiosis. In addition, electron microscopy demonstrated the presence of a putative capsule on in vivo-grown cells. Purification and fractionation of this carbohydrate material from cells grown in carbon-rich synthetic media resulted in the isolation and separation of an antigenically distinct LPS not seen with cells grown in standard media. Antiserum raised against in vivo-grown cells recognized both this LPS and the typical LPS of A. salmonicida apparent in in vitro-grown cells. Antiserum raised against in vitro-grown cells recognized only the LPS expressed in vitro. Antiserum directed against in vivo-grown cells was approximately 10 times more sensitive than serum directed against in vitro-grown cells in detecting A. salmonicida in infected fish kidney tissue.

Full text

PDF
4582

Images in this article

4584Image
on p.4584
4584Image
on p.4584
4585Image
on p.4585
4585Image
on p.4585

Selected References

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

  1. Almeida R. A., Rosenbusch R. F. Capsulelike surface material of Mycoplasma dispar induced by in vitro growth in culture with bovine cells is antigenically related to similar structures expressed in vivo. Infect Immun. 1991 Sep;59(9):3119–3125. doi: 10.1128/iai.59.9.3119-3125.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ames G. F. Resolution of bacterial proteins by polyacrylamide gel electrophoresis on slabs. Membrane, soluble, and periplasmic fractions. J Biol Chem. 1974 Jan 25;249(2):634–644. [PubMed] [Google Scholar]
  3. Blake M. S., Johnston K. H., Russell-Jones G. J., Gotschlich E. C. A rapid, sensitive method for detection of alkaline phosphatase-conjugated anti-antibody on Western blots. Anal Biochem. 1984 Jan;136(1):175–179. doi: 10.1016/0003-2697(84)90320-8. [DOI] [PubMed] [Google Scholar]
  4. Brown M. R., Anwar H., Costerton J. W. Surface antigens in vivo: a mirror for vaccine development. Can J Microbiol. 1988 Apr;34(4):494–498. doi: 10.1139/m88-084. [DOI] [PubMed] [Google Scholar]
  5. Buchmeier N. A., Heffron F. Induction of Salmonella stress proteins upon infection of macrophages. Science. 1990 May 11;248(4956):730–732. doi: 10.1126/science.1970672. [DOI] [PubMed] [Google Scholar]
  6. Camprubi S., Smith M. A., Tomas J. M., Williams P. Modulation of surface antigen expression by Klebsiella pneumoniae in response to growth environment. Microb Pathog. 1992 Aug;13(2):145–155. doi: 10.1016/0882-4010(92)90074-x. [DOI] [PubMed] [Google Scholar]
  7. Chart H., Shaw D. H., Ishiguro E. E., Trust T. J. Structural and immunochemical homogeneity of Aeromonas salmonicida lipopolysaccharide. J Bacteriol. 1984 Apr;158(1):16–22. doi: 10.1128/jb.158.1.16-22.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chart H., Trust T. J. Acquisition of iron by Aeromonas salmonicida. J Bacteriol. 1983 Nov;156(2):758–764. doi: 10.1128/jb.156.2.758-764.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cochrane D. M., Brown M. R., Anwar H., Weller P. H., Lam K., Costerton J. W. Antibody response to Pseudomonas aeruginosa surface protein antigens in a rat model of chronic lung infection. J Med Microbiol. 1988 Dec;27(4):255–261. doi: 10.1099/00222615-27-4-255. [DOI] [PubMed] [Google Scholar]
  10. Crosa J. H. A plasmid associated with virulence in the marine fish pathogen Vibrio anguillarum specifies an iron-sequestering system. Nature. 1980 Apr 10;284(5756):566–568. doi: 10.1038/284566a0. [DOI] [PubMed] [Google Scholar]
  11. Evenberg D., Versluis R., Lugtenberg B. Biochemical and immunological characterization of the cell surface of the fish pathogenic bacterium Aeromonas salmonicida. Biochim Biophys Acta. 1985 May 14;815(2):233–244. doi: 10.1016/0005-2736(85)90294-9. [DOI] [PubMed] [Google Scholar]
  12. Garduño R. A., Thornton J. C., Kay W. W. Aeromonas salmonicida grown in vivo. Infect Immun. 1993 Sep;61(9):3854–3862. doi: 10.1128/iai.61.9.3854-3862.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Garrote A., Bonet R., Merino S., Simon-Pujol M. D., Congregado F. Occurrence of a capsule in Aeromonas salmonicida. FEMS Microbiol Lett. 1992 Aug 15;74(2-3):127–131. doi: 10.1016/0378-1097(92)90417-m. [DOI] [PubMed] [Google Scholar]
  14. Griffiths E., Stevenson P., Thorpe R., Chart H. Naturally occurring antibodies in human sera that react with the iron-regulated outer membrane proteins of Escherichia coli. Infect Immun. 1985 Mar;47(3):808–813. doi: 10.1128/iai.47.3.808-813.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Johne B., Jarp J., Haaheim L. R. Staphylococcus aureus exopolysaccharide in vivo demonstrated by immunomagnetic separation and electron microscopy. J Clin Microbiol. 1989 Jul;27(7):1631–1635. doi: 10.1128/jcm.27.7.1631-1635.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. KEPPIE J., SMITH H., HARRIS-SMITH P. W. The chemical basis of the virulence of Bacillus anthracis. III. The role of the terminal bacteraemia in death of guinea-pigs from anthrax. Br J Exp Pathol. 1955 Jun;36(3):315–322. [PMC free article] [PubMed] [Google Scholar]
  17. Karakawa W. W., Sutton A., Schneerson R., Karpas A., Vann W. F. Capsular antibodies induce type-specific phagocytosis of capsulated Staphylococcus aureus by human polymorphonuclear leukocytes. Infect Immun. 1988 May;56(5):1090–1095. doi: 10.1128/iai.56.5.1090-1095.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kay W. W., Buckley J. T., Ishiguro E. E., Phipps B. M., Monette J. P., Trust T. J. Purification and disposition of a surface protein associated with virulence of Aeromonas salmonicida. J Bacteriol. 1981 Sep;147(3):1077–1084. doi: 10.1128/jb.147.3.1077-1084.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Krivan H. C., Franklin D. P., Wang W., Laux D. C., Cohen P. S. Phosphatidylserine found in intestinal mucus serves as a sole source of carbon and nitrogen for salmonellae and Escherichia coli. Infect Immun. 1992 Sep;60(9):3943–3946. doi: 10.1128/iai.60.9.3943-3946.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. 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]
  21. Mandrell R. E. Further antigenic similarities of Neisseria gonorrhoeae lipooligosaccharides and human glycosphingolipids. Infect Immun. 1992 Jul;60(7):3017–3020. doi: 10.1128/iai.60.7.3017-3020.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Michel C. Furunculosis of salmonids: vaccination attempts in rainbow trout (Salmo gairdneri) by formalin-killed germs. Ann Rech Vet. 1979;10(1):33–40. [PubMed] [Google Scholar]
  23. Morck D. W., Ellis B. D., Domingue P. A., Olson M. E., Costerton J. W. In vivo expression of iron regulated outer-membrane proteins in Pasteurella haemolytica-A1. Microb Pathog. 1991 Nov;11(5):373–378. doi: 10.1016/0882-4010(91)90023-4. [DOI] [PubMed] [Google Scholar]
  24. Panigrahi P., Losonsky G., DeTolla L. J., Morris J. G., Jr Human immune response to Campylobacter jejuni proteins expressed in vivo. Infect Immun. 1992 Nov;60(11):4938–4944. doi: 10.1128/iai.60.11.4938-4944.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. SMITH H., KEPPIE J., STANLEY J. L. The chemical basis of the virulence of Bacillus anthracis. I. Properties of bacteria grown in vivo and preparation of extracts. Br J Exp Pathol. 1953 Oct;34(5):477–485. [PMC free article] [PubMed] [Google Scholar]
  26. Sciortino C. V., Finkelstein R. A. Vibrio cholerae expresses iron-regulated outer membrane proteins in vivo. Infect Immun. 1983 Dec;42(3):990–996. doi: 10.1128/iai.42.3.990-996.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Shand G. H., Anwar H., Kadurugamuwa J., Brown M. R., Silverman S. H., Melling J. In vivo evidence that bacteria in urinary tract infection grow under iron-restricted conditions. Infect Immun. 1985 Apr;48(1):35–39. doi: 10.1128/iai.48.1.35-39.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Smith D. G., Wilcox M. H., Williams P., Finch R. G., Denyer S. P. Characterization of cell envelope proteins of Staphylococcus epidermidis cultured in human peritoneal dialysate. Infect Immun. 1991 Feb;59(2):617–624. doi: 10.1128/iai.59.2.617-624.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Smith H. Pathogenicity and the microbe in vivo. The 1989 Fred Griffith Review Lecture. J Gen Microbiol. 1990 Mar;136(3):377–393. doi: 10.1099/00221287-136-3-377. [DOI] [PubMed] [Google Scholar]
  30. Thornton J. C., Garduño R. A., Newman S. G., Kay W. W. Surface-disorganized, attenuated mutants of Aeromonas salmonicida as furunculosis live vaccines. Microb Pathog. 1991 Aug;11(2):85–99. doi: 10.1016/0882-4010(91)90002-r. [DOI] [PubMed] [Google Scholar]
  31. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Tsai C. M., Frasch C. E. A sensitive silver stain for detecting lipopolysaccharides in polyacrylamide gels. Anal Biochem. 1982 Jan 1;119(1):115–119. doi: 10.1016/0003-2697(82)90673-x. [DOI] [PubMed] [Google Scholar]
  33. Whitfield C., Richards J. C., Perry M. B., Clarke B. R., MacLean L. L. Expression of two structurally distinct D-galactan O antigens in the lipopolysaccharide of Klebsiella pneumoniae serotype O1. J Bacteriol. 1991 Feb;173(4):1420–1431. doi: 10.1128/jb.173.4.1420-1431.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Williams P. Role of the cell envelope in bacterial adaptation to growth in vivo in infections. Biochimie. 1988 Aug;70(8):987–1011. doi: 10.1016/0300-9084(88)90263-5. [DOI] [PubMed] [Google Scholar]

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

RESOURCES