Antigenic diversity of Haemophilus somnus lipooligosaccharide: Phase-variable accessibility of the phosphorylcholine epitope

The lipooligosaccharide (LOS) of Haemophilus somnus undergoes antigenic pha se variation, which may facilitate evasion from the bovine host immune resp onse and/or colonization and dissemination. However, LOS antigenic diversit y in H, somnus has not been adequately investigated. In this study, monoclo nal antibodies (MAbs) specific to various LOS epitopes were used to investi gate antigenic variation and stability in LOS from H, somnus strains and ph ase variants. Clinical isolates of H. somnus exhibited intrastrain, as well as interstrain, antigenic heterogeneity in LOS when probed with MAbs to ou ter core oligosaccharide epitopes in an enzyme-linked immunosorbent assay ( ELISA), However, epitopes reactive with MAbs directed predominately to the inner core heptose region were highly conserved. At least one epitope, whic h was expressed in few strains, was identified. One LOS component affected by phase variation was identified as phosphorylcholine (PCho), which is lin ked to the primary glucose residue. Inhibition ELISA, immunoblotting, and e lectrospray-mass spectrometry were used to confirm that MAb 5F5.9 recognize d PCho, LOS reactivity with MAb 5F5.9 was associated with loss of most of t he outer core oligosaccharide, indicating that reactivity with PCho was aff ected by phase variation of the glycose residues in this region. Our result s indicate that outer core epitopes of H. somnus LOS exhibit a high degree of random, phase-variable antigenic heterogeneity and that such heterogenei ty must be considered in the design of vaccines and diagnostic tests.