Host‐Specific Interactions Between <i>Staphylococcus aureus</i> and <i>Pseudomonas aeruginosa</i> Impair Epithelial Repair in Chronic Rhinosinusitis

BACKGROUND: Chronic rhinosinusitis (CRS) is frequently associated with polymicrobial biofilms involving Staphylococcus aureus and Pseudomonas aeruginosa. Interactions between these organisms are thought to influence disease severity, but the epithelial effects of exoproteins derived from patient-matched cocultures remain poorly defined. METHODS: Clinical isolates of S. aureus and P. aeruginosa (n = 3 each) co-isolated from three CRS patients were cultured in a Transwell system as same-patient or cross-patient pairs. Cell-free exoproteins were applied to primary human nasal epithelial cells. Epithelial repair was assessed using a scratch assay, while cytotoxicity, oxidative stress, and inflammatory responses were evaluated by lactate dehydrogenase release, intracellular reactive oxygen species measurement, and interleukin-6 secretion. Exoprotein profiles were further characterized using data-independent acquisition proteomics. RESULTS: Exoproteins derived from same-patient cocultures consistently impaired epithelial wound closure compared with P. aeruginosa monocultures, although the timing of inhibition varied among patients. These exoproteins also induced greater epithelial cytotoxicity, elevated intracellular reactive oxygen species levels, and increased interleukin-6 secretion compared with monocultures or cross-patient cocultures. In contrast, cross-patient pairings produced epithelial responses similar to monoculture conditions. Proteomic analysis indicated that patient origin was a major determinant of exoproteome organization, with same-patient cocultures showing increased abundance of proteins associated with redox balance and metabolic regulation. CONCLUSIONS: Patient-matched S. aureus-P. aeruginosa interactions were associated with increased epithelial stress and inflammatory responses, together with differences in the exoproteomic profile. These findings suggest that host-specific bacterial interactions may contribute to virulence and possibly recalcitrance in CRS.