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

Objectives The objective of this study was to determine the effects of methylene blue (MB) concentration, laser fluence rate, and laser fluence on efficacy of in vitro photodynamic therapy (PDT) for four bacteria commonly found in human abscesses. Materials and Methods PDT experiments were performed with four of the most common bacteria found in abdominal abscesses: Escherichia coli ( E. coli ), Enterococcus faecalis ( E. faecalis ), Staphylococcus aureus ( S. aureus ), and Pseudomonas aeruginosa ( P. aeruginosa ). MB concentration was varied from 50-300 µg/mL, and laser fluence rate was varied from 1-4 mW/cm 2 at a fluence of 7.2 J/cm 2 . Higher fluence rates and fluences were explored for P. aeruginosa . Primary outcomes were reduction in colony forming units (CFU) following PDT, and measured MB uptake following drug incubation. Results 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 log 10 reduction in CFU when MB concentration was at least 100 µg/mL. P. aeruginosa required higher fluence (28.8 J/cm 2 ) 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, particularly P. aeruginosa , although uptake was not significantly correlated with CFU reduction. Conclusions Gram-positive bacteria can be eradicated in vitro with low levels of MB (50 µg/mL), laser fluence (7.2 J/cm 2 ), and laser fluence rate (1 mW/cm 2 ). E. coli showed substantial cell killing (>6 log 10 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.