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The purpose is to assess the efficacy of rose bengal photodynamic antimicrobial therapy (PDAT) using different irradiation energy levels and photosensitizer concentrations for the inhibition of fungal keratitis isolates. Seven different fungi (Aspergillus fumigatus, Candida albicans, Curvularia lunata, Fusarium keratoplasticum, Fusarium solani, Paecilomyces variotii, and Pseudallescheria boydii) were isolated from patients with confirmed infectious keratitis. Experiments were performed in triplicate with suspensions of each fungus exposed to different PDAT parameters including a control, green light exposure of 5.4 J/cm 2 , 2.7 J/cm 2 (continuous and pulsed), and 1.8 J/cm 2 and rose bengal concentrations of 0.1%, 0.05%, and 0.01%. Plates were photographed 72 h after experimentation, and analysis was performed to assess fungal growth inhibition. PDAT using 5.4 J/cm 2 of irradiation and 0.1% rose bengal completely inhibited growth of five of the seven fungal species. Candida albicans and Fusarium keratoplasticum were the most susceptible organisms, with growth inhibited with the lowest fluence and minimum rose bengal concentration. Fusarium solani, Pseudallescheria boydii, and Paecilomyces variotii were inhibited by lower light exposures and photosensitizer concentrations. Aspergillus fumigatus and Curvularia lunata were not inhibited by any PDAT parameters tested. Continuous and pulsed irradiation using 2.7 J/cm 2 produced similar results. Rose bengal PDAT successfully inhibits the in vitro growth of five fungi known to cause infectious keratitis. Differences in growth inhibition of the various fungi to multiple PDAT parameters suggest that susceptibilities to PDAT are unique among fungal species. These findings support modifying PDAT parameters based on the infectious etiology., (© 2024. The Author(s), under exclusive licence to Springer-Verlag London Ltd., part of Springer Nature.)

Purpose: To evaluate the intermediate-term clinical outcomes of Rose Bengal Photodynamic Antimicrobial Therapy (RB-PDAT) for infectious keratitis; secondarily, to evaluate the surgical outcomes of individuals who underwent optical keratoplasty after RB-PDAT., Design: Retrospective cohort study., Method: A retrospective chart review was performed of 31 eyes from 30 consecutive individuals with infectious keratitis refractory to standard medical therapy who underwent RB-PDAT at the Bascom Palmer Eye Institute between January 2016 and July 2020. Data collected included demographics, risk factors for infectious keratitis, microbiological diagnosis, best spectacle-corrected visual acuity (BCVA), clinical outcomes after RB-PDAT, and complication rates post-keratoplasty. RB-PDAT was performed as described in previous studies. Graft survival was evaluated using Kaplan-Meier curves with log-ranks in individuals who underwent keratoplasty after RB-PDAT., Results: The mean age of the study population was 53 ± 18.0 years. In all, 70% were female; 53.3% self-identified as non-Hispanic White and 43.3% as Hispanic. Mean follow-up time was 28.0 ± 14.4 months. Risk factors included contact lens use (80.6%), history of infectious keratitis (19.3%), and ocular surface disease (16.1%). Cultures were positive for Acanthamoeba (51.6%), Fusarium (12.9%), and Pseudomonas (6.5%). Of the individuals with Acanthamoeba infection, 22.5% were treated with concomitant Miltefosine. Clinical resolution was achieved in 77.4% of patients on average 2.72 ± 1.85 months after RB-PDAT, with 22.5% requiring therapeutic penetrating keratoplasties and 54.8% subsequently requiring optical penetrating keratoplasties. At 2 years, the overall probability of graft survival was 78.7%, and the graft failure rate was 21.3%., Conclusion: RB-PDAT is a potential adjunct therapy for infectious keratitis that may reduce the need for a therapeutic penetrating keratoplasty. Patients who undergo keratoplasty after RB-PDAT may have a higher probability of graft survival at 1 year postoperatively., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Purpose: To examine and compare the efficacy of in vitro growth inhibition using rose bengal and riboflavin photodynamic antimicrobial therapy (PDAT) for Nocardia keratitis isolates., Methods: Nocardia asteroides complex, Nocardia amikacinitolerans, and Nocardia farcinica species were isolated from patients with confirmed Nocardia keratitis. Isolates were tested against three experimental groups: (1) no photosensitizer/no irradiation, (2) photosensitizer/no irradiation, and (3) photosensitizer/irradiation. Each isolate was prepared in suspension to a concentration of 1.5 × 108 CFU/mL. Bacterial suspensions were mixed with water or prepared 0.1% photosensitizer solution for a final bacterial concentration of 1.5 × 107 CFU/mL. Aliquots of 1 mL were plated on 5% sheep blood agar. Rose bengal and riboflavin PDAT plates were irradiated for 15 minutes with a 525- or 375-nm custom 6-mW/cm2 powered light source for a total fluence of 5.4 J/cm2. All experimental groups were repeated in triplicate. Plates were incubated in a 35°C non-CO2 incubator for 96 hours and photographed. Percent inhibition was evaluated using LabVIEW-based software., Results: All strains of Nocardia tested with 0.1% rose bengal and irradiated for 15 minutes demonstrated statistically significant inhibition of growth (P < 0.05). No other experimental groups displayed any bacterial inhibition., Conclusions: Rose bengal is superior to riboflavin PDAT against selected Nocardia isolates. In vivo testing is warranted to investigate the utility of rose bengal PDAT for severe Nocardia keratitis., Translational Relevance: In vitro results for three clinical strains of Nocardia support the possible use of rose bengal PDAT as a complementary treatment of Nocardia keratitis.

Competing Interests: Conflicts of interest Guillermo Amescua is part holder of the patent for the design of the light-source used in the treatment of this patent. The other authors have no conflicts of interest to disclose.

Purpose: To report clinical outcomes of rose bengal photodynamic antimicrobial therapy (RB-PDAT) as an adjunct treatment for severe, progressive infectious keratitis., Design: Consecutive interventional case series., Methods: Patients with progressive infectious keratitis unresponsive to standard medical therapy underwent RB-PDAT at the Bascom Palmer Eye Institute from January 2016 through March 2018. RB-PDAT was performed by applying a solution of rose bengal (0.1% or 0.2% RB in balanced salt solution) to the de-epithelialized cornea for 30 minutes, followed by irradiation with a 6 mW/cm 2 custom-made green LED source for 15 minutes (5.4 J/cm 2 )., Results: The current study included 18 patients (7 male and 11 female) ranging from 17 to 83 years old. Acanthamoeba was the most frequent microbe (10/17; 59%), followed by Fusarium spp. (4/17; 24%), Pseudomonas aeruginosa (2/17; 12%), and Curvularia spp. (1/17; 6%); 1 patient had no confirmed microbiologic diagnosis. Main clinical risk factor for keratitis included contact lens wear (79%). The average area of epithelial defect prior to first RB-PDAT was 32 ± 27 mm 2 and average stromal depth hyperreflectivity measured with anterior segment optical coherence tomography was 269 ± 75 μm. Successful RB-PDAT (avoidance of therapeutic keratoplasty) was achieved in 72% of the cases, with an average time to clinical resolution (decreased pain and inflammation with re-epithelialization and infiltrate resolution) of 46.9 ± 26.4 days after RB-PDAT. Time of follow-up after RB-PDAT was 13.3 ± 5.7 months., Conclusion: RB-PDAT can be considered as an adjunct therapy for cases of severe, progressive infectious keratitis before performing a therapeutic keratoplasty., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Rose Bengal Photodynamic Antimicrobial Therapy (RB-PDAT) is a novel potential treatment for progressive infectious keratitis. The principle behind this therapy is using Rose Bengal as a photosensitizer that can be activated by green light and results in the production of oxygen free radicals which in turn eradicate the microorganism. Given RB-PDAT's mechanism of action and the potential cytotoxic effects, concerns regarding the safety of this technique have arisen. The purpose of this study was to evaluate the effect of RB-PDAT on keratocytes, while focusing on the safety profile that the photo-chemical reaction has on the limbal stem cell (LSC) niche and endothelial cell layer of the treated cornea. To perform RB-PDAT, Rose Bengal solution (0.1% RB in BSS) was applied to the right cornea of rabbits for 30 min and then irradiated by a custom-made green LED light source (525 nm, 6 mW/cm 2 ) for 15 min (5.4 J/cm 2 ). Three rabbits were sacrificed and enucleated after 24 h for evaluation. TUNEL assay and immunohistochemistry for endothelium and limbal stem cell viability were performed on whole mounts and frozen sections in treated and control eyes. LSC of both eyes were isolated and cultured to perform MTT viability and proliferation, and scratch wound healing assays under time-lapse microscopy. Interestingly, while Rose Bengal dye penetration was superficial, yet associated cellular apoptosis was evidenced in up to 1/3 of the stromal thickness on frozen sections. TUNEL assay on whole mounts showed no endothelial cell death following treatment. Immunohistochemistry on frozen sections of LSC displayed no structural difference between treated and non-treated eyes. There was no difference in LSC proliferation rates and scratch wound healing assay demonstrated adequate cell migration from treated and non-treated eyes. The current study suggests that even though penetration of the RB dye has been shown to be limited, oxidative stress produced by RB-PDAT can reach deeper into the corneal stroma. Nevertheless, our results show that performing RB-PDAT is safe on the corneal endothelium and has no effect on LSC viability or function., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Purpose: To evaluate the efficacy of rose bengal PDAT for the management of a patient with multidrug-resistant Fusarium keratoplasticum keratitis unresponsive to standard clinical treatment., Methods: This case report presents a clinical case of F. keratoplasticum keratitis not responsive to standard medical care. In vitro studies from patients culture isolated responded to rose bengal PDAT. Patient received two treatments with rose bengal 0.1% and exposure to green light with a total energy of 2.7 J/cm., Results: In vitro results demonstrated the efficacy of rose bengal PDAT a multidrug-resistant F. keratoplasticum species. There was complete fungal inhibition in our irradiation zone on the agar plates. In the clinical case, the patient was successfully treated with 2 sessions of rose bengal PDAT, and at 8-month follow-up, there was neither recurrence of infection nor adverse effects to report., Conclusions: Rose bengal PDAT is a novel treatment that may be considered in cases of aggressive infectious keratitis. Further studies are needed to understand the mechanisms of PDAT in vivo.