Title: Enhanced Fungal Inhibition with High-Dose Rose Bengal Photodynamic Antimiocrobial Therapy

Abstract

Purpose : Fungal keratitis, primarily caused by Aspergillus and Fusarium species, is the leading cause of unilateral blindness worldwide. When first-line treatments like polyenes and azoles fail, alternative approaches are needed. One such therapy is rose bengal (RB) photodynamic antimicrobial therapy (PDAT) which uses a dye activated by green light to generate antimicrobial singlet oxygen. Aspergillus and Curvularia species are resistant to the standard clinical dose of RB PDAT (0.1% RB, 5.4 J/cm²) and we hypothesize that increasing the RB concentration and light energy will increase the antifungal inhibition.

Methods : Four clinical isolates (Aspergillus glaucus (n =1), Aspergillus fumigatus (n = 1), and Curvularia species (n = 2)) were isolated and prepared in NaCl for a final concentration of 1x10E4. RB was prepared at 3 concentrations (0.1, 0.2, 0.5%) in NaCl. Organisms were mixed with either NaCl (control) or each concentration of RB, and plated in 1mL aliquots onto SabDex agar in triplicate. Plates were further separated into three groups based on light intensity (6, 12, 18 mW) and irradiated with a custom-made LED green (518nm) light for 15 minutes. A control group with no irradiation was per in all four species. At 72 hours, agar plates were photographed to evaluate fungal growth inside of a centered 47-mm irradiation zone.

Results : Growth inhibition to RB PDAT was strain dependent with A. fumigatus showing slightly greater inhibition to RB PDAT than A. glaucus. Inhibition zones for A. fumigatus were highest in the 0.5%RB, 18 mW condition with a small central clearing (~8mm). Both Curvularia species showed less inhibition to RB PDAT with some clearing but no clear inhibition zone. Increasing the concentration and light energy increased the growth inhibition, but not completely.

Conclusions : Growth inhibition to high concentration RB-PDAT was strain dependent; with greater inhibition in the high concentration RB groups, rather than the high fluence groups. However, these results must be considered together as higher RB levels have more light absorption, potentially limiting efficacy in deeper corneal layers. Balancing these factors, the combination of High concentration/High Fluence is likely the most efficacious for these recalcitrant fungal species. Nonetheless, further safety studies are crucial to assess the potential impacts of increased RB and green light exposure on the cornea.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.