The role of licA phase variation in the pathogenesis of invasive disease by Haemophilus influenzae type b

• Published in: FEMS immunology and medical microbiology Vol. 34; no. 3; pp. 221 - 230
• Main Authors: Humphries, Holly E., High, Nicola J.
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing Ltd 15.11.2002; Blackwell; Oxford University Press
• Subjects: Animals; Bactericidal activity; Bacteriology; Bacteriolysis - genetics; Biological and medical sciences; Chemical synthesis; Cloning, Molecular; Decoration; Electrophoresis, Polyacrylamide Gel - methods; Fundamental and applied biological sciences. Psychology; Gel electrophoresis; Gene Deletion; Gene expression; Gene Expression Regulation, Bacterial; H. influenzae; Haemophilus Infections - microbiology; Haemophilus Infections - pathology; Haemophilus influenzae type b - enzymology; Haemophilus influenzae type b - genetics; Haemophilus influenzae type b - pathogenicity; Immunoblotting; Kinases; Lipopolysaccharide, phase variation; Lipopolysaccharides; Lipopolysaccharides - analysis; Lipopolysaccharides - immunology; Microbiology; Models, Genetic; Mutagenesis, Site-Directed; Mutation; Pathogenesis; Pathogenicity, virulence, toxins, bacteriocins, pyrogens, host-bacteria relations, miscellaneous strains; Phase variations; Phosphorylcholine; Phosphorylcholine - metabolism; Phosphotransferases - genetics; Phosphotransferases - metabolism; Polymerase chain reaction; Rats; Sensitivity and Specificity; Sequence Analysis, DNA; Sequence Analysis, Protein; Sodium lauryl sulfate; Virulence; Virulence - genetics; Lipopolysaccharide, phase variation; Recombinant microorganism; Rat; Enzyme; Pathogenesis; Transferases; Virulence; Rodentia; Gene expression; Haemophilus influenzae; Pasteurellaceae; Vertebrata; Mammalia; Kinase; Lipopolysaccharide; Bacteria
• ISSN: 0928-8244; 1574-695X; 2049-632X
• DOI: 10.1111/j.1574-695X.2002.tb00628.x

Abstract LicA encodes the enzyme phosphorylcholine kinase which catalyses the incorporation of phosphorylcholine (ChoP) into H. influenzae LPS. Expression of this gene is subject to phase variation, resulting in the spontaneous loss, or gain of phosphorylcholine (ChoP)-decorated LPS structures. To investigate the role of this phenomenon in the pathogenesis of invasive disease an H. influenzae mutant was constructed which lacked the ability to phase vary licA. This was achieved by introducing an in-frame deletion of the 5′-CAAT-3′ repeats into licA using polymerase chain reaction. The resultant mutant, licAΔ5′-CAAT-3′, was unable to switch off expression of licA and constitutively expressed ChoP-decorated LPS structures, as judged by colony immunoblotting with Mabs 12D9 and HAS. This resulted in increased synthesis of high molecular mass LPS structures and the absence of non-ChoP-decorated LPS species as determined by T–SDS–PAGE analysis. Inability to switch off the expression of licA reduced the virulence of H. influenzae in an infant rat model of invasive disease and resulted in increased sensitivity to the bactericidal activity of serum in the presence of CRP. The ability to switch off the expression of licA through phase variation is therefore concluded to enhance the systemic survival of H. influenzae.