P. aeruginosa flow-cell biofilms are enhanced by repeated phage treatments but can be eradicated by phage-ciprofloxacin combination.

Phage therapy has shown promising results in the treatment of Pseudomonas aeruginosa biofilm infections in animal studies and case reports. The aim of this study was to quantify effects of phage treatments on P. aeruginosa biofilm production and structure. Confocal scanning microscopy was used to follow the interaction between a cocktail of three virulent phages and P. aeruginosa flow-cell biofilms. The role of (i) biofilm age, (ii) repeated phage treatments, (iii) alginate production and (iv) the combination with sub-MIC levels of ciprofloxacin was investigated. Single phage treatment in the early biofilm stages significantly reduced P. aeruginosa PAO1 biovolume (85%-98% reduction). Repeated phage treatments increased the biovolume from 18.25 (untreated biofilm) to 22.24 and 31.07 µm3/µm2 for biofilms treated with phages twice and thrice, respectively. Alginate protected against the phage treatment as the live biovolume remained unaffected by the phage treatment in the mucoid biofilm (20.11 µm3/µm2 in untreated and 21.74 µm3/µm2 in phage-treated biofilm) but decreased in the PAO1 biofilm from 27.35 to 0.89 µm3/µm2. We show that the combination of phages with antibiotics at sub-MIC levels caused a ∼6 log units reduction in the abundance of P. aeruginosa cells in biofilms and that phage treatment increased the size of microcolonies in flow-cell system.