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Identification of two distinct types of flagellar cap proteins, FliD, in Pseudomonas aeruginosa
S K Arora1, N Dasgupta, S Lory
1Department of Medicine/Infectious Diseases, University of Florida, Gainesville, Florida 32610, USA.
Abstract:
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 should be recognized by the host immune system, analogous to the recognition of dominant epitopes located in the exposed parts of the flagellin polypeptide 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 induced during a previous infection. Since similar mechanisms may direct the selection of FliD variants, we examined the extent of sequence heterogeneity among various FliD sequences among a selected group of P. aeruginosa. The results of PCR and nucleotide sequencing of the fliD region of eight different P. aeruginosa strains (laboratory strains PAK, PAO1, and PA103; clinical strains 1244, CS2, and CS32; cystic fibrosis strains CS29 and MDR) suggested that there were two distinct types of FliD in P. aeruginosa, which we named 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 (PAO1) further confirmed the existence of two distinct antigenic types of FliD proteins, with no cross-reactivity between the two serotypes. Further Western immunoblot analysis of the same strains using polyclonal FliC antibody showed that the strains with A-type FliD possessed a-type FliC and those with B-type FliD had b-type FliC. Similar Western blot analyses of 50 more P. aeruginosa strains obtained from varied sources revealed that all strains contained 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 coinherited. This limited diversity of FliC and FliD serotypes seems to be a unique feature of flagellar proteins. A chromosomal mutant having an insertion in the fliD gene of P. aeruginosa PAO1 was constructed. The motility defect of this mutant and a previously constructed PAK fliD mutant was better complemented with the fliD gene of the homologous types.
Insights
Pseudomonas aeruginosa exhibits two main types of flagellar cap protein (FliD), designated A and B. These FliD types are consistently linked with specific flagellin (FliC) types, suggesting coinheritance and limited diversity in these bacterial structures.
Area of Science:
- Microbiology
- Immunology
- Bacterial Pathogenesis
Background:
- Pseudomonas aeruginosa utilizes flagellar proteins, including flagellar cap (FliD) and flagellin (FliC), for host interaction and immune evasion.
- The FliD protein mediates bacterial binding to host mucin, and its exposed nature suggests potential immunogenicity.
- P. aeruginosa exhibits flagellar diversity, likely as a mechanism to evade host antibody responses.
Purpose of the Study:
- To investigate the sequence heterogeneity and antigenic diversity of the FliD protein in P. aeruginosa.
- To determine if FliD diversity correlates with flagellin (FliC) diversity.
- To assess the functional implications of FliD types in bacterial motility and complementation.
Main Methods:
- PCR and nucleotide sequencing were used to analyze the fliD gene region in various P. aeruginosa strains.
- Western blotting and Western immunoblot analysis were employed to assess FliD and FliC protein types and their antigenic relationships.
- Complementation studies were performed using fliD mutants and homologous fliD genes.
Main Results:
- Two distinct types of FliD (A and B) were identified in P. aeruginosa strains.
- A strong correlation was observed between FliD type and FliC type, indicating coinheritance.
- No cross-reactivity was found between antibodies raised against A-type and B-type FliD, confirming distinct antigenic types.
- Complementation of motility defects was most effective with homologous FliD genes.
Conclusions:
- P. aeruginosa possesses only two major serotypes of FliD, which are coinherited with corresponding FliC serotypes.
- This limited diversity in flagellar proteins FliC and FliD is a notable characteristic of P. aeruginosa.
- Understanding FliD and FliC diversity is crucial for developing effective immune strategies against P. aeruginosa infections.