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 lipoprotein protein D was constructed by linker insertion and deletion mutagenesis of a cloned DNA insert containing the protein D structural gene from a nontypeable Haemophilus influenzae strain (NTHi). An isogenic NTHi mutant was isolated after transformation of genetically competent bacteria. The transformant was unreactive to a protein D-specific monoclonal antibody in a colony immunoassay. In addition, the mutant lacked the ability to synthesize detectable levels of protein D by protein staining, immunoblot methods, glycerophosphodiester phosphodiesterase activity, and binding studies of radiolabelled immunoglobulin D. The isogenic 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 order to cause otitis among all rats challenged with that given dose. The protein D mutant exhibited a generation time that was equal to that of the wild-type strain in complex broth medium. No difference in lipopolysaccharide expression was found between the mutant and the parental strain. These results suggest that protein D may influence the pathogenesis of NTHi in the upper respiratory tract.
Janson
H
H
Department of Medical Microbiology, Lund University, University Hospital, Malmö, Sweden.
Melhus
A
A
Hermansson
A
A
Forsgren
A
A
D000818:Animals; D001426:Bacterial Proteins; D001483:Base Sequence; D002352:Carrier Proteins / Q000502:physiology*; D017931:DNA Primers; D005798:Genes, Bacterial; D006192:Haemophilus Infections / Q000382:microbiology*; D006193:Haemophilus influenzae / Q000201:enzymology / Q000235:genetics / Q000472:pathogenicity*; D007072:Immunoglobulin D / Q000276:immunology*; D008074:Lipoproteins / Q000502:physiology*; D008297:Male; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D010033:Otitis Media / Q000382:microbiology*; D010727:Phosphoric Diester Hydrolases / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D017384:Sequence Deletion
D001426:Bacterial Proteins; D002352:Carrier Proteins; D017931:DNA Primers; D007072:Immunoglobulin D; D008074:Lipoproteins; D010727:Phosphoric Diester Hydrolases; C067989:glpQ protein, Haemophilus influenzae; C038133:glycerophosphodiester phosphodiesterase