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Journal of Medical Microbiology logo Journal of Medical Microbiology

Volume 42, Issue 2

A re-appraisal of the biological activity of bacteroides LPS No Access

Abstract

Surmmary

Lipopolysaccharides (LPS) were extracted from seven strains by three different techniques: The phenol-water (PW), phenol-chloroform-petroleum (PCP) and Triton-Mg methods. The strains selected included two different strains, one of which was grown in two different media. Yields varied between the strains, growth media and extraction technique, but generally the highest yield by weight was from the PCP method and the lowest from the PW method. The PW method was selected for the greatest amounts of carbohydrate and KDO, and the PCP method for the least. Phosphorus levels were more uniform among all extraction methods. Protein contamination was found in all LPS extracts, with extremely low levels in PW-LPS and the highest levels in material extracted by the PCP and Triton-Mg techniques. No protein contamination could be detected after proteinase K treatment. After silver staining LPS PAGE profiles showed ladder patterns characteristic of smooth LPS for and the control O18:K strains, whereas the other strains showed mainly rough and low M material only. The PCP method did not select for high M material in the strains; otherwise the LPS profiles for all extraction methods were identical. The biological activities of native and sodium salt form LPS were investigated on a weight for weight basis and compared to that of O18:K PW-LPS. Amongst the LPS from strains, those prepared by the PW method were found to have a significantly higher activity in a galactosamine mouse lethality model, in induction of TNF and the amoebocyte lysate (LAL) assay, than LPS extracted by the PCP or Triton-Mg methods. LPS from strains extracted by the PCP method had consistently low activity in all assays. Comparing PW-LPS from strains with that from O18:K in the galactosamine mouse model, the O18:K LPS was . 5000-fold more active than the most active bacteroides LPS. However, in the LAL assay native PW-LPS from both the strains, and had higher activities (up to 30-fold) than O18:K LPS, with the PW-LPS from the other spp. being up to 15-fold less active than the O18:K PW-LPS. In the TNF induction assay, O18:K PW-LPS was 4–50-fold more active than bacteroides PW-LPS. In the LAL assay and galactosamine mouse model, native LPS had more activity (. two-fold) than sodium salt form LPS. There was no clear difference in activity between native and sodium salt form LPS in the TNF induction assay. The results for the LAL and TNF induction assay were re-evaluated relative to KDO concentration. In the TNF induction assay, previously low activities seen on a weight for weight basis were due in part to less KDO being present. However, LAL activity for PCP-LPS was still low after re-evaluation relative to KDO concentration. The molecular basis for the differences in biological activity of bacteroides LPS in relation to extraction methods and chemical composition is not yet understood.

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© J. MED. MICROBIOL. 1995
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1995-02-01
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References

  1. Duerden B. I., Drasar B. S. (eds) Anaerobes in human disease. London: Edward Arnold; 1991
     [Google Scholar]
  2. Shah H. N., Collins M. D. Proposal to restrict the genus Bacter oides to Bacteroides fragilis and closely related species. Int J Syst Bacteriol 1989; 39:85–87
     [Google Scholar]
  3. Patrick S. The virulence of Bacteroides fragilis. Rev Med Microbiol 1993; 4:40–49
     [Google Scholar]
  4. Kasper D. L., Weintraub A., Lindberg A. A., Lonngren J. Capsular polysaccharides and lipopolysaccharides from two Bacteroides fragilis reference strains: chemical and immunochemical characterization. J Bacteriol 1983; 153:991–997
     [Google Scholar]
  5. Poxton I. R., Brown R. Immunochemistry of the surface carbo hydrate antigens of Bacteroides fragilis and definition of a common antigen. J Gen Microbiol 1986; 132:2475–2481
     [Google Scholar]
  6. Weintraub A., Larsson B. E., Lindberg A. A. Chemical and immunochemical analyses of Bacteroides fragilis lipopolysaccharides. Infect Immun 1985; 49:197–201
     [Google Scholar]
  7. Lindberg A. A., Weintraub A., Zähringer U., Rietschel E. T. Structure-activity relationships in lipopolysaccharides of Bacteroides fragilis. Rev Infect Dis 1990; 12: Suppl 2S133–S141
     [Google Scholar]
  8. Maskell J. P. The resolution of bacteroides lipopolysaccharides by polyacrylamide gel electrophoresis. J Med Microbiol 1991; 34:253–257
     [Google Scholar]
  9. Beckmann I., van Eijk H. G., Meisel-Mikokijczyk F., Wallenburg H. C. Detection of 2-keto-3-deoxyoctonate in endotoxins isolated from six reference strains of the Bacteroides fragilis group. Int J Biochem 1989; 21:661–666
     [Google Scholar]
  10. Runcie C., Ramsay G. Intraabdominal infection: pulmonary failure. World. J Surg 1990; 14:196–203
     [Google Scholar]
  11. Bone R. C. Toward an epidemiology and natural history of SIRS (systemic inflammatory response syndrome). JAMA 1992; 268:3452–3455
     [Google Scholar]
  12. Van Tassell R. L., Wilkins T. D. Isolation of auxotrophs of Bacteroides fragilis. Can J Microbiol 1978; 24:1619–1621
     [Google Scholar]
  13. Westphal O., Lüderitz O. Chemische Erforschung von Lipopoly-sacchariden gramnegativer Bakterien. Angew Chem 1954; 66:407–417
     [Google Scholar]
  14. Galanos C., Lüderitz O., Westphal O. A new method for the extraction of R lipopolysaccharides. Eur J Biochem 1969; 9:245–249
     [Google Scholar]
  15. Uchida K., Mizushima S. A simple method for isolation of lipopolysaccharides from Pseudomonas aeruginosa and some other bacterial strains. Agric Biol Chem 1987; 51:3107–3114
     [Google Scholar]
  16. Hancock I. C., Poxton I. R. (eds) Bacterial cell surface techniques Chichester: UK, Wiley; 1988
     [Google Scholar]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 1970; 227:680–685
     [Google Scholar]
  18. Tsai C.-M., Frasch C. E. A sensitive silver stain for detecting lipopolysaccharides in polyacrylamide gels. Anal Biochem 1982; 119:115–119
     [Google Scholar]
  19. Dubois M., Gilles K. A., Hamilton J. K., Rebers P. A., Smith F. Colorimetric method for determination of sugars and related substances. Anal Chem 1956; 28:350–356
     [Google Scholar]
  20. Chen P. S., Toribara T. Y., Warner H. Microdetermination of phosphorus. Anal Chem 1956; 28:1756–1758
     [Google Scholar]
  21. Lowry O. H., Rosebrough N. J., Farr A. L., Randall R. J. Protein measurement with the Folin phenol reagent. J Biol Chem 1951; 193:265–275
     [Google Scholar]
  22. Galanos C., Lüderitz O. Lipopolysaccharide: properties of an amphipathic molecule. In Rietschel E. T. (ed) Handbook of endotoxin vol 1 Amsterdam: Elsevier; 198446–58
     [Google Scholar]
  23. Loppnow H., Brade H., Dürrbaum I. IL-1 induction-capacity of defined lipopolysaccharide partial structures. J Immunol 1988; 142:3229–3238
     [Google Scholar]
  24. Fujiwara T., Nishihara T., Koga T., Hamada S. Serological properties and immunobiological activities of lipopolysaccharides from black-pigmented and related oral Bacteroides species. J Gen Microbiol 1988; 134:2867–2876
     [Google Scholar]
  25. Allan E., Poxton I. R. The influence of growth medium on serum sensitivity of Bacteroides. J Med Microbiol 1994; 41:45–50
     [Google Scholar]
  26. Lutton D. A., Patrick S., Crockard A. D. Flow cytometric analysis of within-strain variation in polysaccharide expression by Bacteroides fragilis by use of murine monoclonal antibodies. J Med Microbiol 1991; 35:229–237
     [Google Scholar]
/content/journal/jmm/10.1099/00222615-42-2-102
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A re-appraisal of the biological activity of bacteroides LPS
J. Med. Microbiol. 42, 102 (1995); https://doi.org/10.1099/00222615-42-2-102
/content/journal/jmm/10.1099/00222615-42-2-102
/content/journal/jmm/10.1099/00222615-42-2-102
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