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Purification and Visualization of Lipopolysaccharide from Gram-negative Bacteria by Hot Aqueous-phenol Extraction
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Purification and Visualization of Lipopolysaccharide from Gram-negative Bacteria by Hot Aqueous-phenol Extraction

Published on: May 28, 2012

Release of lipopolysaccharide during Bordetella pertussis growth

D Hozbor1, M E Rodriguez, A Samo

  • 1Centro de Investigación y Desarrollo en Fermentaciones Industriales (CINDEFI), La Plata, Argentina.

Research in Microbiology
|March 1, 1993
PubMed

Insights

Adding (2,6-O-dimethyl)-beta-cyclodextrin (Me beta CD) to Bordetella pertussis cultures increased lipopolysaccharide (LPS) release into the medium. This suggests Me beta CD may bind to pertussis cells, affecting endotoxin levels.

Area of Science:

  • Microbiology
  • Immunology
  • Biochemistry

Background:

  • Lipopolysaccharide (LPS), also known as endotoxin, is a major component of the outer membrane of Gram-negative bacteria like Bordetella pertussis.
  • LPS plays a crucial role in the pathogenesis of pertussis and is a key target for vaccine development.
  • Cyclodextrins, particularly modified versions like (2,6-O-dimethyl)-beta-cyclodextrin (Me beta CD), are known to interact with bacterial components and can influence antigen production.

Purpose of the Study:

  • To investigate the effect of Me beta CD addition on LPS release during Bordetella pertussis growth.
  • To determine if Me beta CD influences the yield and pattern of LPS extracted from B. pertussis cells.
  • To assess the impact of Me beta CD on free LPS levels in the culture supernatant.

Main Methods:

  • Bordetella pertussis grown in synthetic Stainer-Scholte (SS) liquid medium with and without Me beta CD (3 mg/ml).
  • Analysis of LPS profiles using SDS-PAGE (sodium dodecyl sulphate/polyacrylamide gel electrophoresis).
  • Quantification of LPS content in whole cells and culture supernatants at different growth phases.

Main Results:

  • SDS-PAGE profiles of LPS from cells grown with and without Me beta CD showed similar patterns.
  • LPS yield from whole cells was lower in the presence of Me beta CD compared to SS medium alone.
  • Higher levels of free LPS were detected in the supernatants of cultures grown with Me beta CD, particularly in the stationary phase.

Conclusions:

  • The addition of Me beta CD to B. pertussis cultures enhances the release of LPS into the supernatant.
  • Me beta CD may interact with Bordetella cells, leading to altered LPS localization and release.
  • These findings have implications for understanding endotoxin dynamics in B. pertussis cultures and potentially for vaccine antigen preparation.

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