In Vitro Analysis of Interactions Between Staphylococcus aureus and Pseudomonas aeruginosa During Biofilm Formation

Staphylococcus aureus and Pseudomonas aeruginosa are classified as ESKAPE pathogens that present a significant challenge to treatment due to their increased resistance to a considerable number of antimicrobial agents. Background/Objective: Biofilms exacerbate treatment challenges by providing enhanced antimicrobial and environmental protection. Mixed-species biofilms further complicate treatment options through numerous complex interspecies interactions, leading to potentially severe adverse clinical outcomes.

This study assessed the interaction between clinical S. aureus and P. aeruginosa isolates during biofilm formation using microplate biofilm formation assays, scanning electron microscopy, and confocal microscopy.

We identified a competitive relationship between P. aeruginosa and S. aureus, where both pathogens exhibited a reduction in biofilm formation during mixed-species biofilms compared with monocultures, although P. aeruginosa outcompeted S. aureus. Furthermore, we found that the cell-free conditioned media (CFCM) of P. aeruginosa significantly reduced the S. aureus biofilms. Using fractioned CFCM, we identified that the anti-staphylococcal activity of the >10 kDa fraction was almost identical to the non-fractioned CFCM. Our confocal microscopy results suggest that P. aeruginosa CFCM depolarize S. aureus membranes and reduces the biofilm burden.

These findings contribute to our understanding of the mechanisms underlying the interactions between these pathogens, suggesting that there is an antagonistic relationship between S. aureus and P. aeruginosa in a biofilm setting.