Susceptibility of phenotypic variants of Haemophilus influenzae type b to serum bactericidal activity: relation to

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Purification and Visualization of Lipopolysaccharide from Gram-negative Bacteria by Hot Aqueous-phenol Extraction
05:31

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

Published on: May 28, 2012

J R Gilsdorf, P Ferrieri

    The Journal of Infectious Diseases
    |February 1, 1986
    PubMed

    Insights

    Passaging Haemophilus influenzae type b in rats created a variant with altered colonial morphology and serum resistance. This change was linked to differences in lipopolysaccharide, not outer membrane proteins or capsule amount.

    Area of Science:

    • Microbiology
    • Immunology
    • Bacterial Pathogenesis

    Background:

    • Haemophilus influenzae type b (Hib) is a significant human pathogen.
    • Bacterial resistance to complement-mediated serum killing is a key virulence factor.
    • Understanding mechanisms of serum resistance is crucial for developing effective therapies.

    Purpose of the Study:

    • To investigate phenotypic and genotypic changes in Haemophilus influenzae type b after serial passage in a rat model.
    • To determine the role of lipopolysaccharide (LPS) and outer membrane proteins (OMPs) in altered serum resistance.

    Main Methods:

    • Serial passage of Hib strain Fuju in rats.
    • Colonial morphology assessment on Levinthal agar.
    • Serum bactericidal assays using normal and immune rat serum.
    • SDS-PAGE analysis of outer membrane proteins.
    • Electrophoretic analysis and monoclonal antibody reactivity of lipopolysaccharides.

    Main Results:

    • Rat-passaged Hib (rat3 Fuju) exhibited opaque colonies and resistance to normal rat serum, unlike the transparent, serum-sensitive parent strain (Fuju).
    • Serum resistance was not attributed to changes in capsular polysaccharide elaboration or major OMP profiles.
    • Differences in LPS electrophoretic mobility and reactivity with Hib LPS-specific monoclonal antibodies were observed between strains.
    • Serum killing of the sensitive strain involved the classic complement pathway.

    Conclusions:

    • Serial passage of Hib in rats induces changes in colonial morphology and serum resistance.
    • Altered serum resistance in the variant strain is associated with modifications in lipopolysaccharide structure.
    • These findings highlight the role of LPS in bacterial adaptation and immune evasion.

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