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Evidence that surface fibrils expressed by Haemophilus influenzae type b promote attachment to human epithelial cells
1Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri.
Abstract:
Haemophilus influenzae type b is a Gram-negative bacillus that initiates infection by colonizing the upper respiratory tract. Previous studies have established that H. influenzae haemagglutinating pili possess adhesive properties and influence the process of colonization. Additional studies suggest the presence of a second H. influenzae adhesin distinct from haemagglutinating pili. In the present study, we examined a non-piliated H. influenzae type b strain by transmission electron microscopy and visualized occasional short, thin, surface fibrils. Subsequently, we isolated a spontaneous mutant that lacked surface fibrils and was deficient in adherence to cultured human epithelial cells. Using a cloning strategy that exploited this mutant, we isolated a fragment of DNA that promotes in vitro adherence to human epithelial cells when expressed in laboratory strains of Escherichia coli. Mutagenesis of this fragment in a series of H. influenzae type b strains resulted in loss of expression of surface fibrils and a marked decrease in attachment. Furthermore, restoration of surface fibril expression was associated with reacquisition of wild-type levels of adherence. Our results are consistent with the conclusion that H. influenzae type b surface fibrils have adhesive capacity. We speculate that these organelles facilitate colonization of the human respiratory tract.
Insights
Haemophilus influenzae type b uses surface fibrils, not just pili, for adherence to human cells. These fibrils are crucial for bacterial colonization in the respiratory tract.
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Cellular Adhesion
Background:
- Haemophilus influenzae type b (Hib) is a Gram-negative bacterium.
- Hib colonizes the upper respiratory tract, initiating infection.
- Previous research identified haemagglutinating pili as adhesins, but suggested other adhesins exist.
Purpose of the Study:
- To investigate a potential second adhesin in Hib distinct from haemagglutinating pili.
- To characterize the role of surface fibrils in Hib adherence and colonization.
Main Methods:
- Transmission electron microscopy to visualize bacterial structures.
- Isolation of a mutant deficient in surface fibrils and adherence.
- DNA cloning to identify adherence-associated genes.
- Mutagenesis studies to confirm gene function.
- In vitro adherence assays using human epithelial cells.
Main Results:
- Observed short, thin surface fibrils on a non-piliated Hib strain.
- A mutant lacking fibrils showed significantly reduced adherence to epithelial cells.
- A cloned DNA fragment encoding surface fibrils enhanced adherence in E. coli.
- Mutagenesis of this fragment in Hib reduced fibril expression and adherence.
Conclusions:
- Hib surface fibrils possess adhesive properties.
- These fibrils likely contribute to the colonization of the human respiratory tract.
- Surface fibrils represent a novel adhesin mechanism for Hib.