Photodynamic fungicidal efficacy of hypericin and dimethyl methylene blue against azole-resistant Candida albicans strains

Antimicrobial photodynamic therapy (aPDT) is an emerging alternative to treat infections based on the use of photosensitisers (PSs) and visible light. To investigate the fungicidal effect of PDT against azole-resistant Candida albicans strains using two PSs with a different mechanism of action, hypericin (HYP) and 1,9-dimethyl methylene blue (DMMB), comparing their efficacy and the reactive oxygen species (ROS) species involved in their cytotoxicity. Azole-resistant and the azole-susceptible C. albicans strains were used. Solutions of 0.5 and 4 McFarland inoculum of each Candida strain were treated with different concentrations of each PS, and exposed to two light-emitting diode light fluences (18 and 37 J cm^-2). Mechanistic insight was gained using several ROS quenchers. The minimal fungicidal concentration of HYP for ≥3 log₁₀ CFU reduction (0.5 McFarland) was 0.62 μmol l^-1 for most strains, whereas for DMMB it ranged between 1.25 and 2.5 μmol l^-1. Increasing the fluence to 37 J cm^-2 allowed to reduce the DMMB concentration. Higher concentrations of both PSs were required to reach a 6 log₁₀ reduction (4 McFarland). H₂O₂ was the main phototoxic species involved in the fungicidal effect of HYP-aPDT whereas ¹O₂ was more important for DMMB-based treatments. aPDT with either HYP or DMMB is effective in killing of C. albicans strains independent of their azole resistance pattern. HYP was more efficient at low fungal concentration and DMMB at higher concentrations.