IMMUNOLOGICAL CHARACTERIZATION OF THE LIPOOLIGOSACCHARIDE-B BAND OF BORDETELLA-PERTUSSIS

Two structurally and immunologically different components of Bordetella pertussis endotoxin can be visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining: a major A band and a faster-migrating minor B band. Certain mutant strains of B. pertussis express only the B band, while the wild-type strains produce both lipooligosaccharides (LOS). Two monoclonal antibodies (MAbs) directed against the minor LOS B band were generated, allowing the study of this surface molecule on different strains of Bordetella. These two MAbs, designated BL-8 and BL-9, reacted strongly with phenol-water-purified LOS obtained from a B. pertussis LOS B mutant strain. Sodium periodate treatment of the purified LOS prevented binding of the MAbs, indicating the carbohydrate nature of the epitope(s). Western immunoblotting experiments revealed that the epitope(s) recognized by these MAbs is conserved on all B. pertussis and Bordetella bronchiseptica Vir- (avirulent) variant strains tested but is not present on Bordetella parapertussis and B. bronchiseptica Vir+ (virulent) wild-type strains. Further studies showed that although present in the lipopolysaccharide B band expressed by Vir- strains, the epitope(s) recognized by the MAbs is not accessible on the surface of intact B. bronchiseptica cells. For B. pertussis, the density and accessibility of this epitope(s) are dependent on the virulence-associated or LOS phenotype expressed by the strain. Our data demonstrate that the expression and accessibility of the epitope(s) are significantly greater on the LOS B variant strains and LOS AB Vir- strains compared with fresh B. pertussis clinical isolates. For these latter strains, which are Vir+, this epitope(s) was barely detectable on the surface of intact bacteria, despite Western blot analyses that revealed specific reactions between the MAbs and the LOS B band. The two LOS B-specific MAbs had no bacteriolytic activity against a LOS AB wild-type strain, while the control MAb BL-2, which is specific for the B. pertussis LOS A band, significantly reduced the number of living bacteria in the same assay. Moderate lytic activity against a mutant strain expressing only the LOS B band was observed for MAb BL-8 but not for MAb BL-9 or BL-2. These data demonstrate that the type, amount, and surface exposure of the LOS are related to the phenotype expressed by a specific B. pertussis strain. In addition, the LOS B MAbs also reveal the antigenic conservation of carbohydrate epitopes among B. pertussis and B. bronchiseptica strains.