Identification of two distinct types of flagellar cap proteins, FliD, in Pseudomonas aeruginosa

Binding of Pseudomonas aeruginosa strain PAK to mucin has been shown to be mediated by the flagellar cap protein, product of the fliD gene. Since the flagellar cap is very likely an exposed structure, the FliD polypeptide sho uld be recognized by the host immune system, analogous to the recognition o f dominant epitopes located in the exposed parts of the flagellin poly-pept ide within the assembled flagellum. In P. aeruginosa, a number of distinct flagellin variants are made, and these variable sequences presumably allow the newly infected P, aeruginosa to escape recognition by the antibody indu ced during a previous infection. Since similar mechanisms mag direct the se lection of FliD variants, we examined the extent of sequence heterogeneity among various FliD sequences among a selected group of P. aeruginosa. The r esults of PCR and nucleotide sequencing of the fliD region of eight differe nt P. aeruginosa strains (laboratory strains PAK, PAO1, and PA103; clinical strains 1244, CS2, and CS32; cystic fibrosis strains CS29 and MDR) suggest ed that there were two distinct types of FliD in P. aeruginosa, which we na med A type and B type. The results of Western blotting using the polyclonal antibodies raised against the purified FliD of A type (PAK) or B type (PAO 1) further confirmed the existence of two distinct antigenic types of FliD proteins, with no cross-reactivity between the two serotypes, Further Weste rn immunoblot analysis of the same strains using polyclonal FliC antibody s howed that the strains with A-type FliD possessed a-type FliC and those wit h E-type FliD had b-type FliC. Similar Western blot analyses of 50 more P. aeruginosa strains obtained from varied sources revealed that all strains c ontained either A-type or B-type FliD, suggesting the existence of only two types of FliD in P. aeruginosa and indicating that fliC and fliD were coin herited. This limited diversity of FliC and FliD serotypes seems to be a un ique feature of flagellar proteins. A chromosomal mutant having an insertio n in the fliD gene of P. aeruginosa PAO1 was constructed. The motility defe ct of this mutant and a previously constructed PAK fliD mutant was better c omplemented with the fliD gene of the homologous types.