This study was designed to investigate the molecular mechanisms of Staphylococcus aureus adherence to human airway epithelium. Using a humanized bronchial xenograft model in the nude mouse and primary cultures of human airway epithelial cells (HAEC), we showed that S. aureus adhered mainly to undifferentiated HAEC whereas weak adherence (11- to 20-fold lower) to differentiated HAEC was observed (P < 0.01). A fibronectin (FN)-binding protein (FnBP)-deficient strain of S. aureus had a fivefold-lower adherence level to undifferentiated HAEC than did the parental strain (P < 0.005), suggesting that S. aureus FN-binding capacity is involved in the adherence to HAEC. We also showed that 97% of 32 S. aureus clinical strains, isolated from the airway secretions of cystic fibrosis patients (n = 18) and patients with nosocomial pneumonia (n = 14), possessed the two fnb genes. The strains from pneumonia patients had a significantly (P < 0.05) higher FN-binding capacity than did the strains from CF patients. This result was confirmed by the expression of FnBPs, investigated by Western ligand affinity blotting. Our results suggest a major role of FnBPs in the colonization of the airways by S. aureus and point to the importance of the adhesin regulatory pathways in the staphylococcal infectious process.