Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1996 Dec;178(24):7254–7259. doi: 10.1128/jb.178.24.7254-7259.1996

Cytometric detection of mycobacterial surface antigens: exposure of mannosyl epitopes and of the arabinan segment of arabinomannans.

V Ozanne 1, A Ortalo-Magne 1, A Vercellone 1, J J Fournie 1, M Daffe 1
PMCID: PMC178641  PMID: 8955410

Abstract

The physical arrangement of cell envelope components leads to the exposure of selected structural motifs which in turn may influence host-parasite interactions. To gain insight into the exposed epitopes, the present study describes a flow cytometric method designed to probe defined molecules on dispersed mycobacteria. The hydrophobic fluorophore N-hexadecanoyl aminofluorescein inserted in the mycobacterial cell envelope permitted focusing of fluorescence-activated cell sorter analysis on cells that were further labeled with defined monoclonal antibodies and fluorochrome-coupled streptavidin. The use of antibodies directed against the lipooligosaccharide of Mycobacterium tuberculosis demonstrated the specific detection of the antigen on the cell surface of a Canetti-like strain of M. tuberculosis, and not on those of mycobacterial strains that were devoid of the glycolipid. Thus, the method was applied to investigate the relative amounts of surface-exposed mannosylated compounds and D-arabinan-containing substances of different strains of the tubercle bacillus and a strain of the rapidly growing nonpathogenic species Mycobacterium smegmatis. Both M. tuberculosis and M. smegmatis are endowed with mannosyl and arabinan epitopes on their surfaces, although there are many differences in terms of exposed mannosyl epitopes between the various strains of the tubercle bacillus examined. These differences are correlated with the amounts of terminal mannosyl residues that cap the surface-exposed arabinomannans (A. Ortalo-Magné, A. B. Andersen, and M. Daffé, Microbiology 142:927-935, 1996) but not with the degrees of virulence of the strains. This novel approach could provide new insights into the distribution of defined surface-exposed antigens and thereby into the architecture of the cell envelopes.

Full Text

The Full Text of this article is available as a PDF (287.4 KB).

Selected References

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

  1. Anton V., Rougé P., Daffé M. Identification of the sugars involved in mycobacterial cell aggregation. FEMS Microbiol Lett. 1996 Nov 1;144(2-3):167–170. doi: 10.1111/j.1574-6968.1996.tb08525.x. [DOI] [PubMed] [Google Scholar]
  2. Bloom B. R., Murray C. J. Tuberculosis: commentary on a reemergent killer. Science. 1992 Aug 21;257(5073):1055–1064. doi: 10.1126/science.257.5073.1055. [DOI] [PubMed] [Google Scholar]
  3. Chatterjee D., Bozic C. M., McNeil M., Brennan P. J. Structural features of the arabinan component of the lipoarabinomannan of Mycobacterium tuberculosis. J Biol Chem. 1991 May 25;266(15):9652–9660. [PubMed] [Google Scholar]
  4. Chatterjee D., Lowell K., Rivoire B., McNeil M. R., Brennan P. J. Lipoarabinomannan of Mycobacterium tuberculosis. Capping with mannosyl residues in some strains. J Biol Chem. 1992 Mar 25;267(9):6234–6239. [PubMed] [Google Scholar]
  5. Cho S. N., Shin J. S., Daffe M., Chong Y., Kim S. K., Kim J. D. Production of monoclonal antibody to a phenolic glycolipid of Mycobacterium tuberculosis and its use in detection of the antigen in clinical isolates. J Clin Microbiol. 1992 Dec;30(12):3065–3069. doi: 10.1128/jcm.30.12.3065-3069.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Daffe M., McNeil M., Brennan P. J. Major structural features of the cell wall arabinogalactans of Mycobacterium, Rhodococcus, and Nocardia spp. Carbohydr Res. 1993 Nov 3;249(2):383–398. doi: 10.1016/0008-6215(93)84102-c. [DOI] [PubMed] [Google Scholar]
  7. Daffe M., McNeil M., Brennan P. J. Novel type-specific lipooligosaccharides from Mycobacterium tuberculosis. Biochemistry. 1991 Jan 15;30(2):378–388. doi: 10.1021/bi00216a011. [DOI] [PubMed] [Google Scholar]
  8. Daffé M., Lacave C., Lanéelle M. A., Lanéelle G. Structure of the major triglycosyl phenol-phthiocerol of Mycobacterium tuberculosis (strain Canetti). Eur J Biochem. 1987 Aug 17;167(1):155–160. doi: 10.1111/j.1432-1033.1987.tb13317.x. [DOI] [PubMed] [Google Scholar]
  9. Draper P., Rees R. J. Electron-transparent zone of mycobacteria may be a defence mechanism. Nature. 1970 Nov 28;228(5274):860–861. doi: 10.1038/228860a0. [DOI] [PubMed] [Google Scholar]
  10. Draper P. The structure of the mycobacterial cell envelope is not yet understood. Res Microbiol. 1991 May;142(4):420–422. doi: 10.1016/0923-2508(91)90113-o. [DOI] [PubMed] [Google Scholar]
  11. Fournié J. J., Mullins R. J., Basten A. Isolation and structural characteristics of a monoclonal antibody-defined cross-reactive phospholipid antigen from Mycobacterium tuberculosis and Mycobacterium leprae. J Biol Chem. 1991 Jan 15;266(2):1211–1219. [PubMed] [Google Scholar]
  12. Fournié J. J., Rivière M., Puzo G. Structural elucidation of the major phenolic glycolipid from Mycobacterium kansasii. I. Evidence for tetrasaccharide structure of the oligosaccharide moiety. J Biol Chem. 1987 Mar 5;262(7):3174–3179. [PubMed] [Google Scholar]
  13. Gaylord H., Brennan P. J., Young D. B., Buchanan T. M. Most Mycobacterium leprae carbohydrate-reactive monoclonal antibodies are directed to lipoarabinomannan. Infect Immun. 1987 Nov;55(11):2860–2863. doi: 10.1128/iai.55.11.2860-2863.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. HANKS J. H. Significance of capsular components of Mycobacterium leprae and other mycobacteria. Int J Lepr. 1961 Jan-Mar;29:74–83. [PubMed] [Google Scholar]
  15. Hunter S. W., Brennan P. J. Evidence for the presence of a phosphatidylinositol anchor on the lipoarabinomannan and lipomannan of Mycobacterium tuberculosis. J Biol Chem. 1990 Jun 5;265(16):9272–9279. [PubMed] [Google Scholar]
  16. Khoo K. H., Dell A., Morris H. R., Brennan P. J., Chatterjee D. Inositol phosphate capping of the nonreducing termini of lipoarabinomannan from rapidly growing strains of Mycobacterium. J Biol Chem. 1995 May 26;270(21):12380–12389. doi: 10.1074/jbc.270.21.12380. [DOI] [PubMed] [Google Scholar]
  17. Lemassu A., Lanéelle M. A., Daffé M. Revised structure of a trehalose-containing immunoreactive glycolipid of Mycobacterium tuberculosis. FEMS Microbiol Lett. 1991 Mar 1;62(2-3):171–175. doi: 10.1016/0378-1097(91)90153-2. [DOI] [PubMed] [Google Scholar]
  18. Lemassu A., Lévy-Frébault V. V., Lanéelle M. A., Daffé M. Lack of correlation between colony morphology and lipooligosaccharide content in the Mycobacterium tuberculosis complex. J Gen Microbiol. 1992 Jul;138(7):1535–1541. doi: 10.1099/00221287-138-7-1535. [DOI] [PubMed] [Google Scholar]
  19. Lemassu A., Ortalo-Magné A., Bardou F., Silve G., Laneélle M. A., Daffé M. Extracellular and surface-exposed polysaccharides of non-tuberculous mycobacteria. Microbiology. 1996 Jun;142(Pt 6):1513–1520. doi: 10.1099/13500872-142-6-1513. [DOI] [PubMed] [Google Scholar]
  20. McNeil M. R., Brennan P. J. Structure, function and biogenesis of the cell envelope of mycobacteria in relation to bacterial physiology, pathogenesis and drug resistance; some thoughts and possibilities arising from recent structural information. Res Microbiol. 1991 May;142(4):451–463. doi: 10.1016/0923-2508(91)90120-y. [DOI] [PubMed] [Google Scholar]
  21. McNeil M., Daffe M., Brennan P. J. Evidence for the nature of the link between the arabinogalactan and peptidoglycan of mycobacterial cell walls. J Biol Chem. 1990 Oct 25;265(30):18200–18206. [PubMed] [Google Scholar]
  22. McNeil M., Daffe M., Brennan P. J. Location of the mycolyl ester substituents in the cell walls of mycobacteria. J Biol Chem. 1991 Jul 15;266(20):13217–13223. [PubMed] [Google Scholar]
  23. Misaki A., Seto N., Azuma I. Structure and immunological properties of D-arabino-D-galactans isolated from cell walls of Mycobacterium species. J Biochem. 1974 Jul;76(1):15–27. doi: 10.1093/oxfordjournals.jbchem.a130540. [DOI] [PubMed] [Google Scholar]
  24. Ortalo-Magné A., Andersen A. B., Daffé M. The outermost capsular arabinomannans and other mannoconjugates of virulent and avirulent tubercle bacilli. Microbiology. 1996 Apr;142(Pt 4):927–935. doi: 10.1099/00221287-142-4-927. [DOI] [PubMed] [Google Scholar]
  25. Ortalo-Magné A., Dupont M. A., Lemassu A., Andersen A. B., Gounon P., Daffé M. Molecular composition of the outermost capsular material of the tubercle bacillus. Microbiology. 1995 Jul;141(Pt 7):1609–1620. doi: 10.1099/13500872-141-7-1609. [DOI] [PubMed] [Google Scholar]
  26. Ortalo-Magné A., Lemassu A., Lanéelle M. A., Bardou F., Silve G., Gounon P., Marchal G., Daffé M. Identification of the surface-exposed lipids on the cell envelopes of Mycobacterium tuberculosis and other mycobacterial species. J Bacteriol. 1996 Jan;178(2):456–461. doi: 10.1128/jb.178.2.456-461.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Prinzis S., Chatterjee D., Brennan P. J. Structure and antigenicity of lipoarabinomannan from Mycobacterium bovis BCG. J Gen Microbiol. 1993 Nov;139(11):2649–2658. doi: 10.1099/00221287-139-11-2649. [DOI] [PubMed] [Google Scholar]
  28. Schlesinger L. S. Macrophage phagocytosis of virulent but not attenuated strains of Mycobacterium tuberculosis is mediated by mannose receptors in addition to complement receptors. J Immunol. 1993 Apr 1;150(7):2920–2930. [PubMed] [Google Scholar]
  29. Venisse A., Berjeaud J. M., Chaurand P., Gilleron M., Puzo G. Structural features of lipoarabinomannan from Mycobacterium bovis BCG. Determination of molecular mass by laser desorption mass spectrometry. J Biol Chem. 1993 Jun 15;268(17):12401–12411. [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES