Molecular analysis of Haemophilus influenzae hemagglutinating pili.

Abstract

Haemophilus influenzae colonizes the upper respiratory tract of humans and is the etiologic agent of various mucosal and invasive diseases. These organisms express surface structures called pili that adhere to host cells and promote colonization. They are defined in vitro by their ability to hemagglutinate human erythrocytes. Pili are composed predominantly of a subunit called pilin. Detailed molecular analysis of the pilus structure may provide information leading to the development of a vaccine capable of blocking microbial attachment. The gene encoding pilin (hifA) was cloned and the deduced primary structure had homology to various E. coli pilus subunits. Further genetic analysis of DNA surrounding the pilin gene revealed additional genes involved in pilus expression. One gene, hifC, was found to express a putative outer membrane protein most likely required for the assembly of pilus subunits into pili on the cell surface. Amino acid homology studies suggested that two other genes, hifD and hifE, encoded pilus structural components. Antisera were made against HifD and HifE proteins expressed from the cloned genes and these antisera were used to demonstrate that both proteins were pilus components localized at the tip of the pilus structure. The hifD and hifE genes and corresponding proteins of many H. influenzae stains were compared and these comparisons revealed that the proteins were genetically and immunologically conserved among organisms responsible for invasive diseases but variable among organisms causing surface localized diseases. These studies demonstrate that H. influenzae pili are heteropolymeric structures and provide the molecular detail necessary to define the adhesive nature of pili and to identify conserved regions of pili useful for vaccine development.