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Functional characterization of type IV pili expressed on diarrhea-associated isolates of Aeromonas species
S M Kirov1, L A O'Donovan, K Sanderson
1Discipline of Pathology, University of Tasmania, Hobart, Tasmania 7001, Australia. S.M.Kirov@utas.edu.au
Abstract:
Our past work has shown that long, flexible type IV pili (single or in bundles) are the predominant pili expressed on fecal isolates of diarrhea-associated species of Aeromonas (Aeromonas veronii biovar sobria and A. caviae). They represent a family of type IV pili which we have designated Bfp (for bundle-forming pili). Reports from Japan suggest that Bfp are intestinal colonization factors. This study presents compelling evidence to support this conclusion. Aeromonas bacteria and/or Bfp purified from a strain of A. veronii biovar sobria were shown to adhere to epithelial and intestinal cell lines, freshly isolated human enterocytes, and fresh and fixed human and rabbit intestinal tissues, as determined by light and electron microscopy and immunohistochemical detection. Removal of Bfp by mechanical means decreased adhesion to cell lines by up to 80%. Purified Bfp blocked adhesion of the test strain to intestinal cells in a dose-dependent manner. Adhesion was also blocked by the Fab fraction of anti-Bfp immunoglobulin G. Moreover, ultrastructural studies (ruthenium red staining and transmission and scanning electron microscopy) demonstrated for the first time that Aeromonas adhesion to human enterocytes is pilus mediated and suggested that Bfp may also promote colonization by forming bacterium-to-bacterium linkages. Bfp-positive isolates examined for type IV pilus-mediated twitching motility in agar and slide culture assays developed for Pseudomonas aeruginosa did not, however, exhibit this function.
Insights
Bundle-forming pili (Bfp) from Aeromonas bacteria are key intestinal colonization factors. These pili mediate bacterial adhesion to human enterocytes, crucial for infection.
Area of Science:
- Microbiology
- Cell Biology
- Infectious Diseases
Background:
- Diarrhea-associated Aeromonas species predominantly express long, flexible type IV pili, termed bundle-forming pili (Bfp).
- Previous reports suggest Bfp function as intestinal colonization factors.
Purpose of the Study:
- To provide compelling evidence that Bfp are intestinal colonization factors.
- To elucidate the role of Bfp in Aeromonas adhesion to host intestinal cells.
Main Methods:
- Adhesion assays using Aeromonas bacteria and purified Bfp with various cell lines and intestinal tissues.
- Light and electron microscopy, immunohistochemical detection.
- Mechanical removal of Bfp, dose-dependent inhibition assays with purified Bfp and anti-Bfp antibodies (Fab fraction).
- Ultrastructural studies including ruthenium red staining and transmission/scanning electron microscopy.
Main Results:
- Aeromonas bacteria and purified Bfp demonstrated significant adherence to epithelial and intestinal cell lines, human enterocytes, and human/rabbit intestinal tissues.
- Mechanical removal of Bfp reduced bacterial adhesion by up to 80%.
- Purified Bfp and anti-Bfp antibodies dose-dependently inhibited bacterial adhesion.
- Ultrastructural studies confirmed pilus-mediated adhesion and suggested Bfp's role in bacterium-to-bacterium linkages.
- Bfp-positive isolates did not exhibit type IV pilus-mediated twitching motility.
Conclusions:
- Bundle-forming pili (Bfp) are critical for the intestinal colonization of diarrhea-associated Aeromonas species.
- Bfp mediate bacterial adhesion to human enterocytes, representing a primary mechanism for colonization.
- Bfp may also contribute to colonization through bacterium-to-bacterium interactions.