PROTEIN-D, THE GLYCEROPHOSPHODIESTER PHOSPHODIESTERASE FROM HAEMOPHILUS-INFLUENZAE WITH AFFINITY FOR HUMAN-IMMUNOGLOBULIN-D, INFLUENCES VIRULENCE IN A RAT OTITIS MODEL

A mutant lacking the ability to express the surface-exposed lipoprotei n protein D was constructed by linker insertion and deletion mutagenes is of a cloned DNA insert containing the protein D structural gene fro m a nontypeable Haemophilus influenzae strain (NTHi). An isogenic NTHi mutant was isolated after transformation of genetically competent bac teria. The transformant was unreactive to a protein D-specific monoclo nal antibody in a colony immunoassay. In addition, the mutant lacked t he ability to synthesize detectable levels of protein D by protein sta ining, immunoblot methods, glycerophosphodiester phosphodiesterase act ivity, and binding studies of radiolabelled immunoglobulin D. The isog enic protein D-deficient mutant was compared with its parental strain for its ability to induce experimental otitis media in rats challenged with bacteria. An approximately 100-times-higher concentration of the mutant compared with that of the wild-type strain was required in ord er to cause otitis among all rats challenged with that given dose. The protein D mutant exhibited a generation time that was equal to that o f the wild-type strain in complex broth medium. No difference in lipop olysaccharide expression was found between the mutant and the parental strain. These results suggest that protein D may influence the pathog enesis of NTHi in the upper respiratory tract.