Abstract
The O-polysaccharide (OPS) of smooth classical Brucella species is a critical virulence determinant, essential serodiagnostic antigen and a crucial component in two of the three vaccines described by the WOAH. Yet, despite its significance, the OPS structure has only been determined for a few (n=7) strains. For other strains the structure and approximate proportion of α1-2 or α1-3 glycosidic linkages is inferred from monoclonal antibody binding profiles; this determines the A or M dominance of strains. The genetic basis for linkage type and proportion has not been identified so cannot be inferred from sequence data or PCR. It is possible that linkage type may have a functional role for Brucella. The ratio and positioning of α1-2 or α1-3 may also play an increasing role in diagnosis and vaccine design given the advent of synthetic oligosaccharide antigens. To better characterise the OPS structure and define relative abundance of the α1-3 glycosidic linkages, we evaluated smooth type strains of B. abortus (n=10), B. melitensis (n=4) and B. suis (n=5) (including vaccine strains B. abortus S19 and B. melitensis Rev.1) and Y. enterocolitica O:9, using 1H Nuclear Magnetic Resonance (NMR). Assignments of the anomeric proton were made at chemical shifts of 5.05 ppm for α1-3 and 5.17 ppm for α1-2 linkage types. Relative abundance of linkage types was determinedas a ratio of peak heights at these chemical shifts. We compared this ratio with monoclonal antibody (Mab) binding data (determined by iELISA). Structural analysis confirmed that α1-3 glycosidic linkages are present in all evaluated Brucella stains (other than B. suis biovar 2), in line with the abundance suggested by MAb binding profiles. A finding from this work that may have a more immediate application is that the OPS from the attenuated B. abortus vaccine strain S19 showed the same glycosidic linkage composition profile as the OPS from diagnostic antigen strains B. abortus S99 and S1119-3. Strain S19 may be a less virulent and safer alternative for use as a diagnostic antigen. SDS-PAGE demonstrated that S19 has similar length OPS to S19 and S1119-3, and this may also be an important diagnostic property.
