Susceptibility of bacterial species commonly found in abdominal abscesses to low‐dose photodynamic therapy: Effects of methylene blue concentration, fluence rate, and fluence

The objective of this study was to determine the effects of methylene blue (MB) concentration, laser fluence rate, and laser fluence on the efficacy of in vitro photodynamic therapy (PDT) for four bacteria commonly found in human abscesses. PDT experiments were performed with four of the most common bacteria found in abdominal abscesses: Escherichia coli, Enterococcus faecalis, Staphylococcus aureus, and Pseudomonas aeruginosa. MB concentration was varied from 50 to 300 μg/mL, and the laser fluence rate was varied from 1 to 4 mW/cm² at a fluence of 7.2 J/cm². Higher fluence rates and fluences were explored for P. aeruginosa. Primary outcomes were the reduction in colony-forming units (CFU) following PDT and measured MB uptake following drug incubation. Gram-positive bacteria (E. faecalis and S. aureus) were eradicated at all MB concentrations and laser fluence rates tested. Efficacy was reduced for E. coli but still resulted in >6 log₁₀ reduction in CFU when MB concentration was at least 100 μg/mL. P. aeruginosa required higher fluence (28.8 J/cm²) to achieve comparable efficacy, while increasing fluence rate did not have a significant effect on PDT efficacy. MB uptake was reduced in Gram-negative species compared to Gram-positive species, particularly P. aeruginosa, although uptake was not significantly correlated with CFU reduction. Gram-positive bacteria can be eradicated in vitro with low levels of MB (50 μg/mL), laser fluence (7.2 J/cm²), and laser fluence rate (1 mW/cm²). E. coli showed substantial cell killing (>6 log₁₀ CFU reduction) with these same parameters. Low MB uptake and PDT efficacy in P. aeruginosa could be overcome by increasing the laser fluence, while increasing fluence rate did not have an effect.