A Possible Phenomenon of Persistence in Pseudomonas aeruginosa Treated with Methylene Blue and Red Light

Planktonic Pseudomonas aeruginosa cells harvested in stationary phase were exposed to red light in the presence of methylene blue to study the potential occurrence of persistence in bacterial populations submitted to photodynamic antimicrobial therapy. Survival curves revealed the existence of small subpopulations of cells exhibiting increased ability to tolerate the treatment. These subpopulations were detected even using high concentrations of photosensitizer, whether added in a single step or following a fractionated scheme, and when the irradiation medium was modified to delay the photodecomposition of methylene blue. When cells grown from survivors to the treatment were cultured and exposed to red light and dye, their responses were similar to that of the original strain. These results exclude exhaustion of the photosensitizer and selection of resistant mutants as explanations for the features of the survival curves. Cells able to tolerate the treatment were found even when radiation was imparted at a high-dose rate. They exhibit a response typical of persisters, which tolerate antimicrobial agents due to transient and reversible changes in their phenotype, suggesting that persistence is a factor to consider upon evaluating the efficacy of photodynamic antimicrobial therapy.