Structural analysis of the lipopolysaccharide (LPS) of nontypeable Haemophilus influenzae strain 1003 has been achieved by the application of high-field NMR techniques, ESI-MS. capillary electrophoresis coupled to ESI-MS. composition and linkage analyses on O-deacylated LPS and core oligosaccharide material. It was found that the LPS contains the common structural element of H. influenzae, L-alpha-D-Hepp-(1 --> 2)-[PEtn --> 6]-L-alpha-D-Hepp-(1 --> 3)-[beta-D-Glcp-(1 --> 4)]-L-alpha-D-Hepp-(1 --> 5)-[PP Etn --> 4]-alpha-Kdop-(2 --> 6)-Lipid A. in which the beta-D-Glcp residue is substituted by phosphocholine at O-6 and an acetyl group at O-4. A second acetyl group is located at O-3 of the distal heptose residue (HepIII). HepIII is chain elongated at O-2 by either a beta-D-Glcp residue (major), lactose or sialyllactose (minor, i.e. alpha-Neu5Ac-(2 --> 3)-beta-D-Galp-(1 --> 4)-beta-D-Glcp), where a third minor acetylation site was identified at the glucose residue. Disialylated species were also detected. In addition. a minor substitution of ester-linked glycine at HepIII and Kdo was observed.
Nontypeable Haemophilus influenzae (NTHi) is an important cause of otitis media and respiratory tract infections. Its lipopolysaccharide (LPS) molecule, an outer membrane component, is a major virulence factor of NTHi. The LPS molecule may also be an efficient target for antibodies which might be protective against NTHi disease. The present thesis describes the structural analyses of the LPS from a number of NTHi clinical isolates. These isolates have been selected to be representative of the genetic diversity in the natural population of NTHi. The studies revealed extended inter- and intrastrain variability in LPS expression but also the presence of a conserved structural element. Several novel structural features were found, including epitopes not previously observed in Haemophilus influenzae LPS. The studies were performed using nuclear magnetic resonance spectroscopy, electrospray ionization mass spectrometry and different chemical degradations of the LPS as the principal methods.