A double-masked, sham-controlled trial of rose bengal photodynamic therapy for the treatment of fungal and acanthameoba keratitis: Rose Bengal Electromagnetic Activation with Green Light for Infection Reduction (REAGIR) Study

A double-masked, sham-controlled trial of rose bengal photodynamic therapy for the treatment of fungal and acanthameoba keratitis: Rose Bengal Electromagnetic Activation with Green Light for Infection Reduction (REAGIR) Study | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article A double-masked, sham-controlled trial of rose bengal photodynamic therapy for the treatment of fungal and acanthameoba keratitis: Rose Bengal Electromagnetic Activation with Green Light for Infection Reduction (REAGIR) Study Venkatesh Prajna, Lalitha Prajna, Sankalp Sharma, Denise de Freitas, and 7 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4165312/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 28 Aug, 2024 Read the published version in Trials → Version 1 posted 5 You are reading this latest preprint version Abstract Background: Infectious keratitis secondary to fungus or acanthamoeba often has a poor outcome despite receiving the best available medical therapy. In vitro Rose Bengal Photodynamic therapy (RB-PDT) appears to be effective against fungal and acanthamoeba isolates. 22,23 In one published series RB-PDT reduced the need for therapeutic penetrating keratoplasty in severe bacterial, fungal, and acanthameoba keratitis not responsive to medical therapy. Methods: This international, randomized, sham and placebo controlled 2-arm clinical trial, randomizes patients with smear positive fungal and acanthameoba and smear negative corneal ulcers in a 1:1 fashion to one of two treatment arms: 1) Topical antimicrobial plus sham RB-PDT or 2) Topical antimicrobial plus RB-PDT Discussion: We anticipate that RB-PDT will improve best spectacle corrected visual acuity and also reduce complications such as corneal perforation and the need for therapeutic penetrating keratoplasty. This study will comply with the NIH Data Sharing Policy and Policy on the Dissemination of NIH-Funded Clinical Trial Information and the Clinical Trials Registration and Results Information Submission rule. Our results will be disseminated via clinicaltrials.gov website, meetings, and journal publications. Our data will also be available upon reasonable request. Trial Registration: NCT, NCT05110001, Registered November 5, 2021. https://www.clinicaltrials.gov/study/NCT05110001 Infectious keratitis cornea RB-PDT Figures Figure 1 Key Messages Fungal or acanthamoeba keratitis often has a poor outcome despite receiving the best available medical therapy. Rose Bengal Photodynamic Therapy (RB-PDT) may directly kill infectious organisms and strengthen the cornea, reducing the risk of perforation or need for therapeutic penetrating keratoplasty. This protocol describes the first large, NIH-funded, randomized clinical trial to study this therapy in patients and will determine whether RB-PDT is a beneficial adjunctive therapy for infectious keratitis. Introduction The photochemical reaction produced during cross-linking (CXL) may benefit patients with infectious corneal ulcers through direct anti-microbial effects as well as increased resistance of corneal tissue to enzymatic degradation. 1 – 3 Activation of a photosensitizer such as riboflavin with exposure to a specific wavelength of light results in release of reactive oxygen species and promotes chemical covalent bond formation between adjacent collagen molecules. CXL with riboflavin, is currently used as a treatment for corneal ectatic disorders such as keratoconus and post-LASIK ectasia and has been shown to stiffen the cornea and allow it to retain its normal shape. 4 – 7 Recently another similar treatment has been proposed that uses rose bengal (RB) as the photosensitizer and green light (532 nm) and is termed Rose Bengal Photodynamic Therapy (RB-PDT). 8 RB-PDT appears to have similar effects on corneal biomechanical properties, is safe for limbal stem cells and endothelium and demonstrates less toxicity to keratocytes in vitro than traditional CXL. 9 – 14 Reactive oxygen species are thought to have an antiseptic effect against a broad range of pathogens. 15 In vitro studies suggest that CXL is effective against common bacterial pathogens, including drug resistant organisms such as Pseudomonas and MRSA. 16 , 17 In vitro studies have demonstrated limited benefit of CXL for fungal or acanthamoeba keratitis, and one randomized clinical trial also did not show a benefit of adjuvant CXL in filamentous fungal keratitis patients. 18 CXL for infectious keratitis is also identified in the literature as Photoactivated Chromophore for Infectious Keratitis (PACK-CXL). 19 – 21 In vitro RB-PDT appears to be much more effective against fungal and acanthamoeba isolates. 22 , 23 Infectious keratitis secondary to fungus or acanthamoeba often has a poor outcome despite receiving the best available medical therapy. For example, the NIH-funded MUTT II randomized controlled trial (RCT) of severe filamentous fungal keratitis demonstrated a 50% rate of full thickness corneal perforation or need for TPK despite maximal medical therapy, including topical natamycin, topical voriconazole, and oral voriconazole. 24 , 25 Clinical studies have suggested a benefit of RB-PDT in infectious keratitis not responsive to medical therapy. 26 One published series demonstrated a 72% reduction in the need for TPK in severe non-responsive bacterial, fungal, and acanthameoba keratitis after RB-PDT. 27 Here, we propose a randomized clinical trial to investigate adjunctive RB-PDT in the treatment of fungal, acanthameoba, and smear and culture negative keratitis. Methods/design Study design The Rose Bengal Electromagnetic Activation with Green Light for Infection Reduction (REAGIR) Study is an international, randomized, outcome masked, sham-controlled 2-arm clinical trial (Full protocol available as online Supplement ). The purpose of this study is to determine differences in 6-month visual acuity between standard therapy with antimicrobials with sham RB-PDT and standard therapy with antimicrobials plus RB-PDT. Patients presenting to one of the Aravind Eye Hospitals in India, or the University Hospital in São Paulo with smear and/or culture-positive fungal, acanthameoba or smear and culture negative keratitis and moderate vision loss, defined as Snellen visual acuity of 20/40 or worse and corneal thickness of greater than or equal to 350 µm as measured on AS-OCT will be included. Figure 1 provides a schematic outline of the study. Those who agree to participate will be randomized in a 1:1 fashion to one of two treatment arms: Group 1, Sham RB-PDT: anti-microbial* plus sham RB-PDT Group 2, RB-PDT: anti-microbial* plus RB-PDT * anti-microbials include moxifloxacin 0.5% for smear and culture negative keratitis, cationoic antiseptic such as chlorhexidine gluconate 0.02% or polyhexamethylene biguanide 0.3% for acanthamoeba keratitis and polyene macrolide such as natamycin 5% or amphotericin b 0.12% for fungal keratitis. Objective and hypothesis The objective of this study is to determine if RB-PDT is a beneficial adjuvant in the treatment of filamentous fungal, acanthamoeba, or smear and culture negative keratitis. We anticipate that RB-PDT will result in better best spectacle corrected visual acuity (BSCVA) at 6 months compared with antimicrobial alone. Study oversight An independent Data and Safety Monitoring Committee (DSMC) oversees the data collection and safety of the study. The DSMC members have expertise in ophthalmology with cornea subspecialty training, biostatistics, and ethics. Interim reports for the DSMC are prepared by the Data Coordinating Center at the F.I. Proctor Foundation (Proctor) at UCSF. These reports include (a) recruitment overall, and by study site, (b) compliance, and (c) retention. The reports also list study outcomes, including 6-month BSCVA and microbiological outcomes, and all adverse outcomes, including mortality and perforations or need for therapeutic penetrating keratoplasty (TPK). The DSMC meets annually in person and biannually via teleconference to monitor study progress and safety. There are also ad hoc meetings as needed. Study investigators conduct site visits at least biannually. The principal investigators notify the DSMC, study sites, and institutional review boards of any changes to study protocols or any deviations from the trial protocols. Setting Participants will be enrolled at four sites in India and one site in Brazil. The study has obtained institutional review board approval at each facility and government approval in each country as well as at the University of California, San Francisco (IRB# 18-26045). In India, participants will be enrolled at Aravind Eye Hospitals in Madurai, Coimbatore and Pondicherry (IRB# 2020009CLI ICMR# 011-26589492). In Brazil, participants will be enrolled at the University Hospital of São Paulo (IRB# 482/21 CONEP# 5.900.981). Inclusion and exclusion criteria Inclusion criteria include age greater than 18, presence of and ulcer that is either smear and culture negative, or is smear or culture positive for filamentous fungus or acanthameoba. They must have Snellen visual acuity of 20/40 or worse with a central corneal thickness greater than or equal to 350 microns as measured by Anterior Segment Optical Coherence Tomography (AS-OCT). Exclusion criteria include evidence of concurrent viral keratitis, impending or frank corneal perforation, involvement of the sclera, non-infectious or autoimmune keratitis, history of recent intraocular surgery or prior corneal transplant, and fellow eye visual acuity worse than 20/200. The investigator will confirm their ability to understand the study and willingness to participate. Randomization Each study eye is randomly assigned to the treatment group. Block randomization stratified by study site using randomly permuted block lengths was performed using a computer program (Statistical package R; Version 2.12; R Foundation for Statistical Computing, Vienna, Austria) by the Data Coordinating Center. Once an eye is enrolled in the study, the study coordinator will assign the study participant’s eye an ID (alpha-numeric code) and topical antimicrobial will begin every hour for 2 days and then every 2 hours while awake until resolution of the epithelial defect. The study coordinator will organize the procedure in the operating room within 48 hours. Once the study participant has been assigned a study participant ID and randomized to treatment group, they will be included in the intent to treat analysis. Intervention & Masking Study participants will undergo RB-PDT or sham RB-PDT within 48 hours of enrollment. Those randomized to the RB-PDT arm will receive a 30-minute loading dose of topical 0.2% rose bengal drops applied in 5-minute intervals to the de-epithelialized cornea. Full penetration through the cornea with anterior segment flare will be confirmed prior to CXL procedure. This will be followed by exposure to continuous 6mW/cm 2 custom-made green light LED source for 15 minutes (total of 5.4 J/cm2). During irradiation patients will continue to receive topical rose bengal at 5-minute intervals. Sham RB-PDT simulates this experience however a green light will be shined adjacent to the patient, careful to avoid exposure to the cornea and the cornea will be covered with a corneal light shield. In place of rose bengal we will use either saline drops. Rose bengal will not be used in the sham procedure due to concern that photochemical activation of the Rose Bengal may occur with exposure to ambient light and therefore produce some treatment effect. All study participants will have repeat corneal cultures 30 minutes after the RB-PDT or sham RB-PDT procedure. Due to the nature of the surgical intervention, the surgeon and technician performing cross-linking will not be masked. The patient, physician performing repeat scraping and clinical follow up, microbiologist, and refractionist performing the BSCVA will be masked to treatment arm. Data collection and management Data collection is the responsibility of the clinical trial staff at the site under the supervision of the site investigator. The investigator is responsible for ensuring the accuracy, completeness, legibility, and timeliness of the data reported. Table 1 outlines the schedule of enrolment, interventions, and assessments. Table 1 Schedule of enrolment, interventions and assessments for the Rose Bengal Electromagnetic Activation with Green Light for Infection Reduction Trial Visit 1 Day 0 Visit 2 Day 1 Visit 3 Day 2 Visit 4 Day 3 Visit 5 3-Week Follow-Up Visit 6 3-Month Follow-Up Visit 7 6-Month Follow-Up Visit 8 12-Month Follow-Up Enrolment Consent & Authorization X Baseline Form X Clinical Drawing X X X X X X VFQ X X Follow-up Form X X X X X Final Form X Interventions CXL/Sham CXL X Study medication # X Assessments IOP X X X X X X Pain Scale X X X AS-OCT X X X X X Confocal Microscopy X X X X X Pentacam Topography X X X X X Clinical Photography* X X X X X Slit Lamp Examination X X X X X X BSCVA/ETDRS/MRx X X X X X Pinhole Visual Acuity X Culture/Smear X X Total Visit Time 2 hours 2 hours 3 hours 0.5 hours 1 hour 1 hour 1 hour 1 hour *Clinical photographs also taken upon adverse events # Difluprednate versus placebo starting at 24 hours Clinical data (including adverse events (AEs), concomitant medications, and expected adverse reactions data) and clinical laboratory data will be entered into Research Electronic Data Capture (REDCap), a 21 CFR Part 11-compliant data capture system provided by the Data Coordinating Center at UCSF. These data will be kept confidential. The data system includes password protection and internal quality checks, such as automatic range checks, to identify data that appear inconsistent, incomplete, or inaccurate. Primary outcome measurement and statistical analyses Visual Acuity . The primary outcome will be 6-month Best Spectacle Corrected Visual Acuity (BSCVA) BSCVA will be measured in a masked fashion using the EDTRS chart with the patient seated 4-meters away and the room lights will be set between 50 to 100 foot-candles. We will use multiple linear regression models to evaluate BSCVA measured with covariates for treatment arm, study site (randomization strata), and baseline pinhole visual acuity. Secondary outcome measures and statistical analyses Visual Acuity at additional time points . As secondary analyses we will also look at 3-week, 3-month, and 12-month BSCVA. We will use multiple linear regression models to evaluate BSCVA measured with covariates for treatment arm, study site (randomization strata), and baseline pinhole visual acuity. A number of subgroup analyses will be performed including organism subtype, infiltrate and/or scar location, and prior antimicrobial use. Microbiological cure. Studies have suggested that in addition to providing an initial diagnosis, repeated culture can be used to assess response to treatment and is highly correlated with clinical outcomes such as visual acuity. 28 – 31 We will re-culture all study participants at Day 2 to assess the effect of RB-PDT on rate of microbiological cure. We hypothesize that those in the RB-PDT group will have a higher rate of microbiological cure on Day 2 cultures than those randomized to sham RB-PDT. We propose the primary analysis to be a Fisher’s exact test comparing the proportion of positivity at follow-up between initially culture positive individuals who were assigned to RB-PDT versus initially culture positive individuals assigned to sham RB-PDT. Additionally, we will report the results for initially culture negative individuals as a supplementary analysis in a logistic regression with assignment, indicators for site (randomization strata), and initial culture results as covariates. Scar/infiltrate. Infiltrate and/or scar size will be measured at the slit lamp by a masked physician by taking the geometric mean of the longest diameter and longest perpendicular to that diameter in millimeters. Hypopyon height will also be recorded in millimeters at the slit lamp. The analysis for scar and/or infiltrate size will follow the templates for visual acuity given above. Multiple linear regression models will be used to evaluate 12-month scar size by treatment arm while correcting for baseline measurements. Corneal thinning and scarring will be evaluated similarly using Anterior Segment Optical Coherence Tomography (AS-OCT) correcting for baseline values. Visual Function Questionnaire (VFQ). VFQ will be compared between arms controlling for Day 1 VFQ. NEI-VFQ in the US, the Brazilian version of the NEI-VFQ will be used in Sao Paulo and the Indian-VFQ (IND-VFQ) in India. This will be conducted using linear regression with baseline and assignment variables. Pentacam Scheimpflug Tomography . Pentacam is a rotating Scheimpflug camera, which provides 3 dimensional images of the cornea. In addition to topographic maps with keratometric readings of the anterior and posterior cornea, Pentacam reports on the total corneal power, corneal thickness maps, higher order aberrations, and densitometry. Statistical analysis will be similar to that describe above, linear mixed effects regression using treatment assignment and baseline values as covariates, using the same template as we did for BSCVA. Adverse events and statistical analyses TPK/Perforation . A Cox proportional hazards model will estimate the hazard of perforation, defined as perforation (flat anterior chamber with presence of iris plugging a defect in the cornea or seidel positivity) or the need for TPK while correcting for baseline infiltrate depth. Interim analysis Interim reports for the DSMC are prepared by the Data Coordinating Center. These reports include (a) recruitment overall, and by study site, (b) compliance, and (c) retention. The reports will also list study outcomes, including 6-month BSCVA and microbiological outcomes, and all adverse outcomes, including mortality and perforations. All adverse events are tabulated and reported. Statistical comparisons will be conducted using Fisher’s exact test, but with the caution that failure to find a statistically significant difference cannot be used to infer a lack of risk difference, since the study is not powered to examine rare outcomes. Procedures for reporting both adverse events and serious adverse events, including notification of the Medical Monitor, were reviewed by the DSMC prior to opening enrollment. We will categorize adverse events, severe adverse events, and events of interest following recommended best practices for clinical trial monitoring and reporting. 11 Sample size calculation The trial’s sample size calculation was based on the primary outcome, 6-month BSCVA. We informed the calculation with measurements from the first Steroids for Corneal Ulcers Trial (SCUT), among patients enrolled with between 20/60 and 20/400 vision. The SCUT trial measured BSCVA at baseline, 3-months, and 12-months. We conservatively used the 12-month outcome measure for the calculations since there was no 6-month measurement. The standard deviation of BSCVA at 12 months was 0.293. Since the primary analysis will adjust for baseline BSCVA, we used an estimate of the residual standard deviation, which is 𝑆𝐷𝑟=𝑆𝐷1−𝑟2‾‾‾‾‾‾√SDr = SD1−r2 where r is the correlation between the baseline measure and primary endpoint. In SCUT, the correlation between baseline and 12-month BSCVA among patients with between 20/60 and 20/400 vision at enrollment was 0.216. We thus assumed a residual standard deviation of 0.2931 − 0.2162‾‾‾‾‾‾‾‾‾‾√=0.2860.2931 − 0.2162 = 0.286 Assuming a significance level of 0.05, allowing for approximately 15% loss to follow up, we estimate that we will have 90% power to detect a 1.1-line difference (logMAR 0.11) between groups with 165 study participants per arm (330 total). For the same sample size and under the same assumptions, the detectable difference at 80% power is 1.0-lines (logMAR 0.10). These calculations were based on the standard power formula for the T-test (using an estimated residual standard deviation). Dissemination plan This study will comply with the NIH Data Sharing Policy and Policy on the Dissemination of NIH-Funded Clinical Trial Information and the Clinical Trials Registration and Results Information Submission rule. As such, this trial is registered at ClinicalTrials.gov (NCT05110001), and results from this trial will be submitted and published on ClinicalTrials.gov. In addition, every attempt will be made to publish results in peer-reviewed journals and to present these data at national and international meetings. Consistent with the collaborative nature of the proposed research, the PI anticipates sharing all data generated by the study with collaborators. Analytic datasets that will be developed through the project will comply with the NIH Data Sharing Policy. The analytical datasets from this project will include patient-level data generated from the study visits. Data from the trial will be made available upon reasonable request. Discussion Although bacterial corneal ulcers are more common in the US, fungal and acanthameoba keratitis (AK) presents a therapeutic challenge to clinicians because of poor outcomes and few treatment options. 32 – 38 In the tropics, fungal infection can account for upwards of 50% of corneal ulcers. 32 , 37 , 39 In the United States fungal keratitis ranges from 35% of corneal ulcers in South Florida 40 to 4% in temperate climates such as Los Angeles. 41 These infections can occur after trauma, with contact lens wear, or after refractive surgery. 42 , 43 An outbreak of Fusarium keratitis among contact lens wearers was related to the ReNu Moistureloc™ contact lens solution, which was subsequently removed from the market. 44 The best treatment strategies for fungal keratitis have not been well characterized. Topical natamycin, a polyene, is the only antifungal agent approved by the Food and Drug Administration (FDA) for treatment of fungal keratitis. The Mycotic Ulcer Treatment Trials (MUTT) I and II, were two NEI-funded randomized double-masked clinical trials that found topical natamycin to be superior to topical voriconazole and no additional benefit of adjuvant oral voriconazole. Two recent randomized clinical trials also failed to demonstrate a benefit of adjuvant intrastromal voriconazole or adjuvant UVX in the treatment fungal keratitis. 45 , 46 However, natamycin is fungistatic and has limited penetration into the corneal layers. 47 Furthermore, outcomes of fungal keratitis with topical natamycin are extremely poor as demonstrated in MUTT II where approximately 50% of patients had full thickness corneal perforation or required TPK despite topical natamycin, topical voriconazole, and adjuvant oral voriconazole. Although much less common, acanthameoba keratitis (AK) may have the most prolonged and severe course of any corneal infection. AK is typically related to contact lens use and the incidence of these infections varies from as low as 1% to 4–8% of culture positive microbial keratitis cases in countries where contact lens use is common. 48 Topical biguanides such as chlorhexidine 0.02% and Polyhexamethylene biguanide (PHMB) 0.02% are thought to be the most effective available medical therapy. However, large series suggest that only 60% of patients achieve complete cure with medical therapy alone by one year and that almost 50% end up with a poor outcome, defined as requiring TPK or having visual acuity less than 20/80. 49 Furthermore, these medications are highly toxic and cause permanent damage to delicate ocular structures such as limbal stem cells and trabecular meshwork. 50 Corneal cross-linking (CXL) is a novel prospective therapy that may simultaneously reduce both ocular pathogens and inflammatory cells and strengthen the cornea. 1 – 3 CXL with riboflavin (UVX) and Rose Bengal with green light (RB-PDT) are both effective in vitro against common bacterial ocular pathogens, such as Pseudomonas aeruginosa and Streptococcus pneumoniae . 17 , 51 However, UVX appears to have much less effect on fungal and acanthamoeba organisms in vitro and one randomized clinical trial also did not show a benefit of adjuvant UVX in the treatment of filamentous fungal keratitis patients. 18 In vitro RB-PDT, appears to be much more effective against fungal and acanthamoeba isolates. 22 , 23 Rose Bengal (RB) is one of the most commonly used dyes in the diagnosis of ocular surface disease. 52 Rose Bengal is an effective photosensitizer, readily converting triplet oxygen ( 3 O 2 ) to produce high singlet oxygen ( 1 O 2 ) yields with exposure to green light. 53 Although RB dye penetration is to approximately 100µm into the stroma, subsequent free radical formation occurs up to 1/3 of the corneal stromal depth. 26 , 54 The ability of RB to continue free radical formation is self-limited after photo-irradiation has ceased. 55 Multiple In vitro and ex vivo studies have suggested that RGX may be safer than UVX. Wound healing studies found more corneal haze and slower wound healing after UVX compared with RGX. 9 Rabbit studies have demonstrated the safety of RGX on limbal stem cells and endothelium and found anterior stromal keratocyte damage in RGX comparable to epithelial debridement alone. 13 , 14 By contrast, UVX causes an immediate decrease in the sub-epithelial nerve plexus and loss of keratocytes in the anterior one-third of the corneal stroma, although this recovers after a few months. 56 , 57 Smear and culture negative ulcers represent another therapeutic challenge for clinicians. Up to 60% of corneal cultures are smear and culture negative. 58 When these patients do not improve with topical antibiotics alone, clinicians must decide what alternative medical therapy to introduce. There is little guidance in the literature on how to manage these patients. These cases are challenging to study since they represent different underlying etiologies and one medical therapy is unlikely to address all of them. RB-PDT is unique in its potential to address bacterial, fungal and parasitic infections making it a particularly attractive novel therapy. Limitations to our study include the fact that while our study design is practical in terms of resources, the organisms have different clinical courses will likely respond differently to RB-PDT. It is true that these cases are typically not studied together since one medical therapy is unlikely to address all of the underlying etiologies. RB-PDT is unique in its potential to address bacterial, fungal and parasitic infections, making it possible to enroll all of these ulcers in one study. This also makes RB-PDT a particularly appealing therapy for smear and culture negative cases, which are common and are a therapeutic challenge for clinicians. 58 Here, we propose pre-specified subgroup analyses for acanthameoba, fungal, and smear/culture negative, to analyze the effects of RB-PDT on each type of ulcer. Conclusion Here we explore a novel adjuvant therapy for the treatment of fungal, acanthameoba and smear and culture negative keratitis. Reducing the global burden of vision loss from corneal opacification will likely require a multidisciplinary approach including corneal ulcer prevention, novel antimicrobial agents and adjuvant therapies such as RB-PDT. Trial status Protocol version 3.0 last edited March 15 th 2023. Recruitment began in January 2022 and is expected to last until approximately January 2025. Declarations Declarations Ethics Approval and Consent to Participate Single IRB approval for US sites was obtained at the University of California, San Francisco (#1826045). Institutional IRB approval was obtained at the Aravind Eye Hospital, Madurai (2020009CLI). Council on Medical Research (ICMR) approval was also obtained for the study (011-26589492). Competing Interests There are no conflicts of interest to report. Funding This work was supported by grants UG1 EY028518 (Lietman/Rose-Nussbaumer) and K23 EY025025 (Rose-Nussbaumer) from the National Eye Institute an institutional P30 grant from the National Eye Institute (Stanford) and Research to Prevent Blindness (UCSF & Stanford University). Authors’ contributions Tom Lietman, Jennifer Rose-Nussbaumer, NV Prajna and N Radhakrishnan have contributed to the design of the study and study implementation and these collaborators make up the steering committee. Tom Lietman is the head of the data coordinating center. Jennifer Rose-Nussbaumer is the head of the clinical coordinating center. P Lalitha and AL Höfling-Lima are contributing to study implementation. Benjamin Arnold is the biostatistician and as such has directed the statistical analysis plan and will have access to the final dataset. Acknowledgements Our study team would like to thank Drs. Jean-Marie Parel and Guillermo Amescua from the Ophthalmic Biophysics Center at the Bascom Palmer Eye Institute for donation of RB-PDT equipment and their generous support with their technical expertise. Availability of Data and Material Every attempt will be made to publish results in peer-reviewed journals and to present these data at national and international meetings. Consistent with the collaborative nature of the proposed research, the PI anticipates sharing all data generated by the study with collaborators. Analytic datasets that will be developed through the project will comply with the NIH Data Sharing Policy. The analytical datasets from this project will include patient-level data generated from the study visits. Data from the trial will be made available upon reasonable request. References Alio JL, Abbouda A, Valle DD, et al. Corneal cross linking and infectious keratitis: a systematic review with a meta-analysis of reported cases. Journal of ophthalmic inflammation and infection. 2013;3(1):47. Sand D, She R, Shulman IA, et al. Microbial keratitis in los angeles: the doheny eye institute and the los angeles county hospital experience. Ophthalmology. 2015;122(5):918-924. Papaioannou L, Miligkos M, Papathanassiou M. Corneal Collagen Cross-Linking for Infectious Keratitis: A Systematic Review and Meta-Analysis. Cornea. 2016;35(1):62-71. Keating A, Pineda R, 2nd, Colby K. Corneal cross linking for keratoconus. Seminars in ophthalmology. 2010;25(5-6):249-255. Lamy R, Netto CF, Reis RG, et al. Effects of corneal cross-linking on contrast sensitivity, visual acuity, and corneal topography in patients with keratoconus. Cornea. 2013;32(5):591-596. Raiskup-Wolf F, Hoyer A, Spoerl E, et al. Collagen crosslinking with riboflavin and ultraviolet-A light in keratoconus: long-term results. Journal of cataract and refractive surgery. 2008;34(5):796-801. Vinciguerra P, Albe E, Trazza S, et al. Intraoperative and postoperative effects of corneal collagen cross-linking on progressive keratoconus. Archives of ophthalmology. 2009;127(10):1258-1265. Zhu H, Alt C, Webb RH, et al. Corneal Crosslinking With Rose Bengal and Green Light: Efficacy and Safety Evaluation. Cornea. 2016;35(9):1234-1241. Lorenzo-Martin E, Gallego-Munoz P, Ibares-Frias L, et al. Rose Bengal and Green Light Versus Riboflavin-UVA Cross-Linking: Corneal Wound Repair Response. Invest Ophthalmol Vis Sci. 2018;59(12):4821-4830. Cherfan D, Verter EE, Melki S, et al. Collagen cross-linking using rose bengal and green light to increase corneal stiffness. Invest Ophthalmol Vis Sci. 2013;54(5):3426-3433. Bekesi N, Gallego-Munoz P, Ibares-Frias L, et al. Biomechanical Changes After In Vivo Collagen Cross-Linking With Rose Bengal-Green Light and Riboflavin-UVA. Invest Ophthalmol Vis Sci. 2017;58(3):1612-1620. Wang T, Zhu L, Zhu J, et al. Subacute effects of rose Bengal/Green light cross linking on rabbit thin corneal stability and safety. Lasers Surg Med. 2018;50(4):324-332. Naranjo A, Pelaez D, Arrieta E, et al. Cellular and molecular assessment of rose bengal photodynamic antimicrobial therapy on keratocytes, corneal endothelium and limbal stem cell niche. Exp Eye Res. 2019;188:107808. Gallego-Munoz P, Ibares-Frias L, Lorenzo E, et al. Corneal Wound Repair After Rose Bengal and Green Light Crosslinking: Clinical and Histologic Study. Invest Ophthalmol Vis Sci. 2017;58(9):3471-3480. Vatansever F, de Melo WC, Avci P, et al. Antimicrobial strategies centered around reactive oxygen species--bactericidal antibiotics, photodynamic therapy, and beyond. FEMS Microbiol Rev. 2013;37(6):955-989. Halili F, Arboleda A, Durkee H, et al. Rose Bengal- and Riboflavin-Mediated Photodynamic Therapy to Inhibit Methicillin-Resistant Staphylococcus aureus Keratitis Isolates. Am J Ophthalmol. 2016;166:194-202. Durkee H, Arboleda A, Aguilar MC, et al. Rose bengal photodynamic antimicrobial therapy to inhibit Pseudomonas aeruginosa keratitis isolates. Lasers Med Sci. 2019. Prajna NV, Radhakrishnan N, Lalitha P, et al. Cross-Linking-Assisted Infection Reduction: A Randomized Clinical Trial Evaluating the Effect of Adjuvant Cross-Linking on Outcomes in Fungal Keratitis. Ophthalmology. 2019. Ozbek-Uzman S, Yalniz-Akkaya Z, Burcu A. Corneal Collagen Cross-Linking With Photoactivated Chromophore for Infectious Keratitis After Penetrating Keratoplasty. Cornea. 2020;39(3):283-289. Kasetsuwan N, Reinprayoon U, Satitpitakul V. Photoactivated Chromophore for Moderate to Severe Infectious Keratitis as an Adjunct Therapy: A Randomized Controlled Trial. Am J Ophthalmol. 2016;165:94-99. Said DG, Elalfy MS, Gatzioufas Z, et al. Collagen cross-linking with photoactivated riboflavin (PACK-CXL) for the treatment of advanced infectious keratitis with corneal melting. Ophthalmology. 2014;121(7):1377-1382. Atalay HT, Dogruman-Al F, Sarzhanov F, et al. Effect of Riboflavin/Rose Bengal-Mediated PACK-CXL on Acanthamoeba Trophozoites and Cysts in Vitro. Curr Eye Res. 2018;43(11):1322-1325. Arboleda A, Miller D, Cabot F, et al. Assessment of rose bengal versus riboflavin photodynamic therapy for inhibition of fungal keratitis isolates. Am J Ophthalmol. 2014;158(1):64-70.e62. Prajna NV, Krishnan T, Rajaraman R, et al. Effect of Oral Voriconazole on Fungal Keratitis in the Mycotic Ulcer Treatment Trial II (MUTT II): A Randomized Clinical Trial. JAMA Ophthalmol. 2016;134(12):1365-1372. Prajna NV, Krishnan T, Rajaraman R, et al. Predictors of Corneal Perforation or Need for Therapeutic Keratoplasty in Severe Fungal Keratitis: A Secondary Analysis of the Mycotic Ulcer Treatment Trial II. JAMA Ophthalmol. 2017;135(9):987-991. Arboleda A, Miller D, Cabot F, et al. Assessment of rose bengal versus riboflavin photodynamic therapy for inhibition of fungal keratitis isolates. Am J Ophthalmol. 2014;158(1):64-70 e62. Naranjo A, Arboleda A, Martinez JD, et al. Rose Bengal Photodynamic Antimicrobial Therapy (RB-PDAT) for Patients with Progressive Infectious Keratitis: A Pilot Clinical Study. Am J Ophthalmol. 2019. Ray KJ, Lalitha P, Prajna NV, et al. The Utility of Repeat Culture in Fungal Corneal Ulcer Management: A Secondary Analysis of the MUTT I Randomized Clinical Trial. Am J Ophthalmol. 2017. Bhadange Y, Das S, Kasav MK, et al. Comparison of culture-negative and culture-positive microbial keratitis: cause of culture negativity, clinical features and final outcome. The British journal of ophthalmology. 2015;99(11):1498-1502. McLeod SD, Kolahdouz-Isfahani A, Rostamian K, et al. The Role of Smears, Cultures, and Antibiotic Sensitivity Testing in the Management of Suspected Infectious Keratitis. Ophthalmology. 1996;103(1):23-28. Vemuganti GK, Garg P, Gopinathan U, et al. Evaluation of agent and host factors in progression of mycotic keratitis: A histologic and microbiologic study of 167 corneal buttons. Ophthalmology. 2002;109(8):1538-1546. Gopinathan U, Garg P, Fernandes M, et al. The epidemiological features and laboratory results of fungal keratitis: a 10-year review at a referral eye care center in South India. Cornea. 2002;21(6):555-559. FlorCruz NV, Evans JR. Medical interventions for fungal keratitis. The Cochrane database of systematic reviews. 2015(4):CD004241. Feilmeier MR, Sivaraman KR, Oliva M, et al. Etiologic diagnosis of corneal ulceration at a tertiary eye center in Kathmandu, Nepal. Cornea. 2010;29(12):1380-1385. Dunlop AA, Wright ED, Howlader SA, et al. Suppurative corneal ulceration in Bangladesh. A study of 142 cases examining the microbiological diagnosis, clinical and epidemiological features of bacterial and fungal keratitis. Australian and New Zealand journal of ophthalmology. 1994;22(2):105-110. Wong TY, Ng TP, Fong KS, et al. Risk factors and clinical outcomes between fungal and bacterial keratitis: a comparative study. The CLAO journal : official publication of the Contact Lens Association of Ophthalmologists, Inc. 1997;23(4):275-281. Srinivasan M, Gonzales CA, George C, et al. Epidemiology and aetiological diagnosis of corneal ulceration in Madurai, south India. The British journal of ophthalmology. 1997;81(11):965-971. Whitcher JP, Srinivasan M. Corneal ulceration in the developing world--a silent epidemic. The British journal of ophthalmology. 1997;81(8):622-623. Deorukhkar S, Katiyar R, Saini S. Epidemiological features and laboratory results of bacterial and fungal keratitis: a five-year study at a rural tertiary-care hospital in western Maharashtra, India. Singapore medical journal. 2012;53(4):264-267. Liesegang TJ, Forster RK. Spectrum of microbial keratitis in South Florida. American journal of ophthalmology. 1980;90(1):38-47. McLeod SD, Kolahdouz-Isfahani A, Rostamian K, et al. The role of smears, cultures, and antibiotic sensitivity testing in the management of suspected infectious keratitis. Ophthalmology. 1996;103(1):23-28. Verma S, Tuft SJ. Fusarium solani keratitis following LASIK for myopia. The British journal of ophthalmology. 2002;86(10):1190-1191. Bernal MD, Acharya NR, Lietman TM, et al. Outbreak of Fusarium keratitis in soft contact lens wearers in San Francisco. Archives of ophthalmology. 2006;124(7):1051-1053. Yildiz EH, Abdalla YF, Elsahn AF, et al. Update on fungal keratitis from 1999 to 2008. Cornea. 2010;29(12):1406-1411. Prajna NV, Radhakrishnan N, Lalitha P, et al. Cross-Linking-Assisted Infection Reduction: A Randomized Clinical Trial Evaluating the Effect of Adjuvant Cross-Linking on Outcomes in Fungal Keratitis. Ophthalmology. 2020;127(2):159-166. Narayana S, Krishnan T, Ramakrishnan S, et al. Mycotic Antimicrobial Localized Injection: A Randomized Clinical Trial Evaluating Intrastromal Injection of Voriconazole. Ophthalmology. 2019;126(8):1084-1089. O'Day DM, Head WS, Robinson RD, et al. Corneal penetration of topical amphotericin B and natamycin. Current eye research. 1986;5(11):877-882. Dart JK, Saw VP, Kilvington S. Acanthamoeba keratitis: diagnosis and treatment update 2009. Am J Ophthalmol. 2009;148(4):487-499.e482. Papa V, Rama P, Radford C, et al. Acanthamoeba keratitis therapy: time to cure and visual outcome analysis for different antiamoebic therapies in 227 cases. Br J Ophthalmol. 2019. Carrijo-Carvalho LC, Sant'ana VP, Foronda AS, et al. Therapeutic agents and biocides for ocular infections by free-living amoebae of Acanthamoeba genus. Surv Ophthalmol. 2017;62(2):203-218. Martins SA, Combs JC, Noguera G, et al. Antimicrobial efficacy of riboflavin/UVA combination (365 nm) in vitro for bacterial and fungal isolates: a potential new treatment for infectious keratitis. Investigative ophthalmology & visual science. 2008;49(8):3402-3408. Khan-Lim D, Berry M. Still confused about rose bengal? Curr Eye Res. 2004;29(4-5):311-317. Wachter E, Dees C, Harkins J, et al. Topical rose bengal: pre-clinical evaluation of pharmacokinetics and safety. Lasers Surg Med. 2003;32(2):101-110. Amescua G, Arboleda A, Nikpoor N, et al. Rose Bengal Photodynamic Antimicrobial Therapy: A Novel Treatment for Resistant Fusarium Keratitis. Cornea. 2017;36(9):1141-1144. Tonogai Y, Ito Y, Iwaida M, Tati M, et al. Studies on the toxicity of coal-tar dyes. III. Reason of acute toxicity to fish caused by coal-tar dyes and their industrial effluents. J Toxicol Sci. 1980;5(1):23-33. Mazzotta C, Hafezi F, Kymionis G, et al. In Vivo Confocal Microscopy after Corneal Collagen Crosslinking. Ocul Surf. 2015;13(4):298-314. Sharma N, Suri K, Sehra SV, et al. Collagen cross-linking in keratoconus in Asian eyes: visual, refractive and confocal microscopy outcomes in a prospective randomized controlled trial. Int Ophthalmol. 2015;35(6):827-832. Peng MY, Cevallos V, McLeod SD, et al. Bacterial Keratitis: Isolated Organisms and Antibiotic Resistance Patterns in San Francisco. Cornea. 2018;37(1):84-87. 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19:12:49","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-4165312/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-4165312/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s13063-024-08376-3","type":"published","date":"2024-08-28T15:57:23+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":59523820,"identity":"8f411dd2-4612-44c6-b9b7-8b9f6dda96f6","added_by":"auto","created_at":"2024-07-02 20:42:21","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":125048,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSchema of Rose Bengal Electromagnetic Activation with Green Light Protocol\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"Fig1.REAGIRSCHEMA.png","url":"https://assets-eu.researchsquare.com/files/rs-4165312/v1/d344d3ecea866a559ee8eb14.png"},{"id":63820882,"identity":"19857732-e362-42d9-a4b8-c8ce22c481f4","added_by":"auto","created_at":"2024-09-02 16:09:49","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":881663,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-4165312/v1/0d79b760-97e4-4e29-a328-68b75a47863c.pdf"},{"id":59523822,"identity":"714f0025-0ad2-483a-b077-c8bdee1ed6df","added_by":"auto","created_at":"2024-07-02 20:42:21","extension":"doc","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":125952,"visible":true,"origin":"","legend":"","description":"","filename":"SPIRITFillablechecklist.doc","url":"https://assets-eu.researchsquare.com/files/rs-4165312/v1/4f770e8fe5fa030242ddbbe9.doc"},{"id":59523821,"identity":"6fd43deb-f11a-4c54-a4dc-b4545f627bfc","added_by":"auto","created_at":"2024-07-02 20:42:21","extension":"docx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":454791,"visible":true,"origin":"","legend":"","description":"","filename":"SCUTIIClinicalProtocolv3.0.docx","url":"https://assets-eu.researchsquare.com/files/rs-4165312/v1/f217cf39f2939b831cb5e392.docx"}],"financialInterests":"","formattedTitle":"A double-masked, sham-controlled trial of rose bengal photodynamic therapy for the treatment of fungal and acanthameoba keratitis: Rose Bengal Electromagnetic Activation with Green Light for Infection Reduction (REAGIR) Study","fulltext":[{"header":"Key Messages","content":"\u003cp\u003eFungal or acanthamoeba keratitis often has a poor outcome despite receiving the best available medical therapy. Rose Bengal Photodynamic Therapy (RB-PDT) may directly kill infectious organisms and strengthen the cornea, reducing the risk of perforation or need for therapeutic penetrating keratoplasty. This protocol describes the first large, NIH-funded, randomized clinical trial to study this therapy in patients and will determine whether RB-PDT is a beneficial adjunctive therapy for infectious keratitis.\u003c/p\u003e"},{"header":"Introduction","content":"\u003cp\u003eThe photochemical reaction produced during cross-linking (CXL) may benefit patients with infectious corneal ulcers through direct anti-microbial effects as well as increased resistance of corneal tissue to enzymatic degradation.\u003csup\u003e\u003cspan additionalcitationids=\"CR2\" citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e Activation of a photosensitizer such as riboflavin with exposure to a specific wavelength of light results in release of reactive oxygen species and promotes chemical covalent bond formation between adjacent collagen molecules. CXL with riboflavin, is currently used as a treatment for corneal ectatic disorders such as keratoconus and post-LASIK ectasia and has been shown to stiffen the cornea and allow it to retain its normal shape.\u003csup\u003e\u003cspan additionalcitationids=\"CR5 CR6\" citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e Recently another similar treatment has been proposed that uses rose bengal (RB) as the photosensitizer and green light (532 nm) and is termed Rose Bengal Photodynamic Therapy (RB-PDT).\u003csup\u003e\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e RB-PDT appears to have similar effects on corneal biomechanical properties, is safe for limbal stem cells and endothelium and demonstrates less toxicity to keratocytes \u003cem\u003ein vitro\u003c/em\u003e than traditional CXL.\u003csup\u003e\u003cspan additionalcitationids=\"CR10 CR11 CR12 CR13\" citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eReactive oxygen species are thought to have an antiseptic effect against a broad range of pathogens.\u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e \u003cem\u003eIn vitro\u003c/em\u003e studies suggest that CXL is effective against common bacterial pathogens, including drug resistant organisms such as Pseudomonas and MRSA.\u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e,\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e \u003cem\u003eIn vitro\u003c/em\u003e studies have demonstrated limited benefit of CXL for fungal or acanthamoeba keratitis, and one randomized clinical trial also did not show a benefit of adjuvant CXL in filamentous fungal keratitis patients.\u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e CXL for infectious keratitis is also identified in the literature as Photoactivated Chromophore for Infectious Keratitis (PACK-CXL).\u003csup\u003e\u003cspan additionalcitationids=\"CR20\" citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e \u003cem\u003eIn vitro\u003c/em\u003e RB-PDT appears to be much more effective against fungal and acanthamoeba isolates.\u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e,\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eInfectious keratitis secondary to fungus or acanthamoeba often has a poor outcome despite receiving the best available medical therapy. For example, the NIH-funded MUTT II randomized controlled trial (RCT) of severe filamentous fungal keratitis demonstrated a 50% rate of full thickness corneal perforation or need for TPK despite maximal medical therapy, including topical natamycin, topical voriconazole, and oral voriconazole.\u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e,\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e\u003c/sup\u003e Clinical studies have suggested a benefit of RB-PDT in infectious keratitis not responsive to medical therapy.\u003csup\u003e\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e One published series demonstrated a 72% reduction in the need for TPK in severe non-responsive bacterial, fungal, and acanthameoba keratitis after RB-PDT.\u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e Here, we propose a randomized clinical trial to investigate adjunctive RB-PDT in the treatment of fungal, acanthameoba, and smear and culture negative keratitis.\u003c/p\u003e"},{"header":"Methods/design","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eStudy design\u003c/h2\u003e \u003cp\u003eThe Rose Bengal Electromagnetic Activation with Green Light for Infection Reduction (REAGIR) Study is an international, randomized, outcome masked, sham-controlled 2-arm clinical trial (Full protocol available as online \u003cb\u003eSupplement\u003c/b\u003e). The purpose of this study is to determine differences in 6-month visual acuity between standard therapy with antimicrobials with sham RB-PDT and standard therapy with antimicrobials plus RB-PDT. Patients presenting to one of the Aravind Eye Hospitals in India, or the University Hospital in S\u0026atilde;o Paulo with smear and/or culture-positive fungal, acanthameoba or smear and culture negative keratitis and moderate vision loss, defined as Snellen visual acuity of 20/40 or worse and corneal thickness of greater than or equal to 350 \u0026micro;m as measured on AS-OCT will be included. Figure\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e provides a schematic outline of the study.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThose who agree to participate will be randomized in a 1:1 fashion to one of two treatment arms:\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003eGroup 1, Sham RB-PDT: anti-microbial* plus sham RB-PDT\u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003eGroup 2, RB-PDT: anti-microbial* plus RB-PDT\u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003e* anti-microbials include moxifloxacin 0.5% for smear and culture negative keratitis, cationoic antiseptic such as chlorhexidine gluconate 0.02% or polyhexamethylene biguanide 0.3% for acanthamoeba keratitis and polyene macrolide such as natamycin 5% or amphotericin b 0.12% for fungal keratitis.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003eObjective and hypothesis\u003c/h2\u003e \u003cp\u003eThe objective of this study is to determine if RB-PDT is a beneficial adjuvant in the treatment of filamentous fungal, acanthamoeba, or smear and culture negative keratitis. We anticipate that RB-PDT will result in better best spectacle corrected visual acuity (BSCVA) at 6 months compared with antimicrobial alone.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003eStudy oversight\u003c/h2\u003e \u003cp\u003eAn independent Data and Safety Monitoring Committee (DSMC) oversees the data collection and safety of the study. The DSMC members have expertise in ophthalmology with cornea subspecialty training, biostatistics, and ethics. Interim reports for the DSMC are prepared by the Data Coordinating Center at the F.I. Proctor Foundation (Proctor) at UCSF. These reports include (a) recruitment overall, and by study site, (b) compliance, and (c) retention. The reports also list study outcomes, including 6-month BSCVA and microbiological outcomes, and all adverse outcomes, including mortality and perforations or need for therapeutic penetrating keratoplasty (TPK). The DSMC meets annually in person and biannually via teleconference to monitor study progress and safety. There are also \u003cem\u003ead hoc\u003c/em\u003e meetings as needed. Study investigators conduct site visits at least biannually. The principal investigators notify the DSMC, study sites, and institutional review boards of any changes to study protocols or any deviations from the trial protocols.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eSetting\u003c/h2\u003e \u003cp\u003eParticipants will be enrolled at four sites in India and one site in Brazil. The study has obtained institutional review board approval at each facility and government approval in each country as well as at the University of California, San Francisco (IRB# 18-26045). In India, participants will be enrolled at Aravind Eye Hospitals in Madurai, Coimbatore and Pondicherry (IRB# 2020009CLI ICMR# 011-26589492). In Brazil, participants will be enrolled at the University Hospital of S\u0026atilde;o Paulo (IRB# 482/21 CONEP# 5.900.981).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003eInclusion and exclusion criteria\u003c/h2\u003e \u003cp\u003eInclusion criteria include age greater than 18, presence of and ulcer that is either smear and culture negative, or is smear or culture positive for filamentous fungus or acanthameoba. They must have Snellen visual acuity of 20/40 or worse with a central corneal thickness greater than or equal to 350 microns as measured by Anterior Segment Optical Coherence Tomography (AS-OCT). Exclusion criteria include evidence of concurrent viral keratitis, impending or frank corneal perforation, involvement of the sclera, non-infectious or autoimmune keratitis, history of recent intraocular surgery or prior corneal transplant, and fellow eye visual acuity worse than 20/200. The investigator will confirm their ability to understand the study and willingness to participate.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eRandomization\u003c/h2\u003e \u003cp\u003eEach study eye is randomly assigned to the treatment group. Block randomization stratified by study site using randomly permuted block lengths was performed using a computer program (Statistical package R; Version 2.12; R Foundation for Statistical Computing, Vienna, Austria) by the Data Coordinating Center. Once an eye is enrolled in the study, the study coordinator will assign the study participant\u0026rsquo;s eye an ID (alpha-numeric code) and topical antimicrobial will begin every hour for 2 days and then every 2 hours while awake until resolution of the epithelial defect. The study coordinator will organize the procedure in the operating room within 48 hours. Once the study participant has been assigned a study participant ID and randomized to treatment group, they will be included in the intent to treat analysis.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003eIntervention \u0026amp; Masking\u003c/h2\u003e \u003cp\u003eStudy participants will undergo RB-PDT or sham RB-PDT within 48 hours of enrollment. Those randomized to the RB-PDT arm will receive a 30-minute loading dose of topical 0.2% rose bengal drops applied in 5-minute intervals to the de-epithelialized cornea. Full penetration through the cornea with anterior segment flare will be confirmed prior to CXL procedure. This will be followed by exposure to continuous 6mW/cm\u003csup\u003e2\u003c/sup\u003e custom-made green light LED source for 15 minutes (total of 5.4 J/cm2). During irradiation patients will continue to receive topical rose bengal at 5-minute intervals. Sham RB-PDT simulates this experience however a green light will be shined adjacent to the patient, careful to avoid exposure to the cornea and the cornea will be covered with a corneal light shield. In place of rose bengal we will use either saline drops. Rose bengal will not be used in the sham procedure due to concern that photochemical activation of the Rose Bengal may occur with exposure to ambient light and therefore produce some treatment effect. All study participants will have repeat corneal cultures 30 minutes after the RB-PDT or sham RB-PDT procedure.\u003c/p\u003e \u003cp\u003eDue to the nature of the surgical intervention, the surgeon and technician performing cross-linking will not be masked. The patient, physician performing repeat scraping and clinical follow up, microbiologist, and refractionist performing the BSCVA will be masked to treatment arm.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec10\" class=\"Section2\"\u003e \u003ch2\u003eData collection and management\u003c/h2\u003e \u003cp\u003eData collection is the responsibility of the clinical trial staff at the site under the supervision of the site investigator. The investigator is responsible for ensuring the accuracy, completeness, legibility, and timeliness of the data reported. Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e outlines the schedule of enrolment, interventions, and assessments.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eSchedule of enrolment, interventions and assessments for the Rose Bengal Electromagnetic Activation with Green Light for Infection Reduction Trial\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"9\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eVisit 1\u003c/p\u003e \u003cp\u003eDay 0\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eVisit 2\u003c/p\u003e \u003cp\u003eDay 1\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eVisit 3\u003c/p\u003e \u003cp\u003eDay 2\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eVisit 4\u003c/p\u003e \u003cp\u003eDay 3\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eVisit 5\u003c/p\u003e \u003cp\u003e3-Week\u003c/p\u003e \u003cp\u003eFollow-Up\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c7\"\u003e \u003cp\u003eVisit 6\u003c/p\u003e \u003cp\u003e3-Month Follow-Up\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c8\"\u003e \u003cp\u003eVisit 7\u003c/p\u003e \u003cp\u003e6-Month\u003c/p\u003e \u003cp\u003eFollow-Up\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c9\"\u003e \u003cp\u003eVisit 8\u003c/p\u003e \u003cp\u003e12-Month Follow-Up\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eEnrolment\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eConsent \u0026amp; Authorization\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eBaseline Form\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eClinical Drawing\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eVFQ\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFollow-up Form\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eFinal Form\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eInterventions\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCXL/Sham CXL\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eStudy medication\u003csup\u003e#\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAssessments\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eIOP\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePain Scale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eAS-OCT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eConfocal Microscopy\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePentacam Topography\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eClinical Photography*\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSlit Lamp Examination\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eBSCVA/ETDRS/MRx\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePinhole Visual Acuity\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eCulture/Smear\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eX\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTotal Visit Time\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2 hours\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2 hours\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e3 hours\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.5 hours\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1 hour\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c7\"\u003e \u003cp\u003e1 hour\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c8\"\u003e \u003cp\u003e1 hour\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c9\"\u003e \u003cp\u003e1 hour\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"9\"\u003e*Clinical photographs also taken upon adverse events\u003c/td\u003e\u003c/tr\u003e \u003ctr\u003e\u003ctd colspan=\"9\"\u003e\u003csup\u003e\u003cb\u003e#\u003c/b\u003e\u003c/sup\u003eDifluprednate versus placebo starting at 24 hours\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eClinical data (including adverse events (AEs), concomitant medications, and expected adverse reactions data) and clinical laboratory data will be entered into Research Electronic Data Capture (REDCap), a 21 CFR Part 11-compliant data capture system provided by the Data Coordinating Center at UCSF. These data will be kept confidential. The data system includes password protection and internal quality checks, such as automatic range checks, to identify data that appear inconsistent, incomplete, or inaccurate.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003ePrimary outcome measurement and statistical analyses\u003c/h2\u003e \u003cp\u003e \u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003eVisual Acuity\u003c/span\u003e. The primary outcome will be 6-month Best Spectacle Corrected Visual Acuity (BSCVA) BSCVA will be measured in a masked fashion using the EDTRS chart with the patient seated 4-meters away and the room lights will be set between 50 to 100 foot-candles. We will use multiple linear regression models to evaluate BSCVA measured with covariates for treatment arm, study site (randomization strata), and baseline pinhole visual acuity.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003eSecondary outcome measures and statistical analyses\u003c/h2\u003e \u003cp\u003e \u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003eVisual Acuity at additional time points\u003c/span\u003e. As secondary analyses we will also look at 3-week, 3-month, and 12-month BSCVA. We will use multiple linear regression models to evaluate BSCVA measured with covariates for treatment arm, study site (randomization strata), and baseline pinhole visual acuity. A number of subgroup analyses will be performed including organism subtype, infiltrate and/or scar location, and prior antimicrobial use.\u003c/p\u003e \u003cp\u003e \u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003eMicrobiological cure.\u003c/span\u003e Studies have suggested that in addition to providing an initial diagnosis, repeated culture can be used to assess response to treatment and is highly correlated with clinical outcomes such as visual acuity.\u003csup\u003e\u003cspan additionalcitationids=\"CR29 CR30\" citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e We will re-culture all study participants at Day 2 to assess the effect of RB-PDT on rate of microbiological cure. We hypothesize that those in the RB-PDT group will have a higher rate of microbiological cure on Day 2 cultures than those randomized to sham RB-PDT.\u003c/p\u003e \u003cp\u003eWe propose the primary analysis to be a Fisher\u0026rsquo;s exact test comparing the proportion of positivity at follow-up between initially culture positive individuals who were assigned to RB-PDT versus initially culture positive individuals assigned to sham RB-PDT. Additionally, we will report the results for initially culture negative individuals as a supplementary analysis in a logistic regression with assignment, indicators for site (randomization strata), and initial culture results as covariates.\u003c/p\u003e \u003cp\u003e \u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003eScar/infiltrate.\u003c/span\u003e Infiltrate and/or scar size will be measured at the slit lamp by a masked physician by taking the geometric mean of the longest diameter and longest perpendicular to that diameter in millimeters. Hypopyon height will also be recorded in millimeters at the slit lamp. The analysis for scar and/or infiltrate size will follow the templates for visual acuity given above. Multiple linear regression models will be used to evaluate 12-month scar size by treatment arm while correcting for baseline measurements. Corneal thinning and scarring will be evaluated similarly using Anterior Segment Optical Coherence Tomography (AS-OCT) correcting for baseline values.\u003c/p\u003e \u003cp\u003e \u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003eVisual Function Questionnaire (VFQ).\u003c/span\u003e VFQ will be compared between arms controlling for Day 1 VFQ. NEI-VFQ in the US, the Brazilian version of the NEI-VFQ will be used in Sao Paulo and the Indian-VFQ (IND-VFQ) in India. This will be conducted using linear regression with baseline and assignment variables.\u003c/p\u003e \u003cp\u003e \u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003ePentacam Scheimpflug Tomography\u003c/span\u003e. Pentacam is a rotating Scheimpflug camera, which provides 3 dimensional images of the cornea. In addition to topographic maps with keratometric readings of the anterior and posterior cornea, Pentacam reports on the total corneal power, corneal thickness maps, higher order aberrations, and densitometry. Statistical analysis will be similar to that describe above, linear mixed effects regression using treatment assignment and baseline values as covariates, using the same template as we did for BSCVA.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003eAdverse events and statistical analyses\u003c/h2\u003e \u003cp\u003e \u003cspan type=\"ItalicUnderline\" class=\"ItalicUnderline\" name=\"Emphasis\"\u003eTPK/Perforation\u003c/span\u003e. A Cox proportional hazards model will estimate the hazard of perforation, defined as perforation (flat anterior chamber with presence of iris plugging a defect in the cornea or seidel positivity) or the need for TPK while correcting for baseline infiltrate depth.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003eInterim analysis\u003c/h2\u003e \u003cp\u003eInterim reports for the DSMC are prepared by the Data Coordinating Center. These reports include (a) recruitment overall, and by study site, (b) compliance, and (c) retention. The reports will also list study outcomes, including 6-month BSCVA and microbiological outcomes, and all adverse outcomes, including mortality and perforations. All adverse events are tabulated and reported. Statistical comparisons will be conducted using Fisher\u0026rsquo;s exact test, but with the caution that failure to find a statistically significant difference cannot be used to infer a lack of risk difference, since the study is not powered to examine rare outcomes. Procedures for reporting both adverse events and serious adverse events, including notification of the Medical Monitor, were reviewed by the DSMC prior to opening enrollment. We will categorize adverse events, severe adverse events, and events of interest following recommended best practices for clinical trial monitoring and reporting.\u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003eSample size calculation\u003c/h2\u003e \u003cp\u003eThe trial\u0026rsquo;s sample size calculation was based on the primary outcome, 6-month BSCVA. We informed the calculation with measurements from the first Steroids for Corneal Ulcers Trial (SCUT), among patients enrolled with between 20/60 and 20/400 vision. The SCUT trial measured BSCVA at baseline, 3-months, and 12-months. We conservatively used the 12-month outcome measure for the calculations since there was no 6-month measurement. The standard deviation of BSCVA at 12 months was 0.293. Since the primary analysis will adjust for baseline BSCVA, we used an estimate of the residual standard deviation, which is\u003c/p\u003e \u003cp\u003e\u0026#119878;\u0026#119863;\u0026#119903;=\u0026#119878;\u0026#119863;1\u0026minus;\u0026#119903;2\u0026oline;\u0026oline;\u0026oline;\u0026oline;\u0026oline;\u0026oline;\u0026radic;SDr\u0026thinsp;=\u0026thinsp;SD1\u0026minus;r2\u003c/p\u003e \u003cp\u003ewhere \u003cem\u003er\u003c/em\u003e is the correlation between the baseline measure and primary endpoint. In SCUT, the correlation between baseline and 12-month BSCVA among patients with between 20/60 and 20/400 vision at enrollment was 0.216. We thus assumed a residual standard deviation of\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003e0.2931\u0026thinsp;\u0026minus;\u0026thinsp;0.2162\u0026oline;\u0026oline;\u0026oline;\u0026oline;\u0026oline;\u0026oline;\u0026oline;\u0026oline;\u0026oline;\u0026oline;\u0026radic;=0.2860.2931\u0026thinsp;\u0026minus;\u0026thinsp;0.2162\u0026thinsp;=\u0026thinsp;0.286\u003c/h2\u003e \u003cp\u003eAssuming a significance level of 0.05, allowing for approximately 15% loss to follow up, we estimate that we will have 90% power to detect a 1.1-line difference (logMAR 0.11) between groups with 165 study participants per arm (330 total). For the same sample size and under the same assumptions, the detectable difference at 80% power is 1.0-lines (logMAR 0.10). These calculations were based on the standard power formula for the T-test (using an estimated residual standard deviation).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003eDissemination plan\u003c/h2\u003e \u003cp\u003eThis study will comply with the NIH Data Sharing Policy and Policy on the Dissemination of NIH-Funded Clinical Trial Information and the Clinical Trials Registration and Results Information Submission rule. As such, this trial is registered at ClinicalTrials.gov (NCT05110001), and results from this trial will be submitted and published on ClinicalTrials.gov. In addition, every attempt will be made to publish results in peer-reviewed journals and to present these data at national and international meetings. Consistent with the collaborative nature of the proposed research, the PI anticipates sharing all data generated by the study with collaborators. Analytic datasets that will be developed through the project will comply with the NIH Data Sharing Policy. The analytical datasets from this project will include patient-level data generated from the study visits. Data from the trial will be made available upon reasonable request.\u003c/p\u003e \u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eAlthough bacterial corneal ulcers are more common in the US, fungal and acanthameoba keratitis (AK) presents a therapeutic challenge to clinicians because of poor outcomes and few treatment options.\u003csup\u003e\u003cspan additionalcitationids=\"CR33 CR34 CR35 CR36 CR37\" citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e\u003c/sup\u003e In the tropics, fungal infection can account for upwards of 50% of corneal ulcers.\u003csup\u003e\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e,\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e,\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e\u003c/sup\u003e In the United States fungal keratitis ranges from 35% of corneal ulcers in South Florida\u003csup\u003e40\u003c/sup\u003e to 4% in temperate climates such as Los Angeles.\u003csup\u003e\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e\u003c/sup\u003e These infections can occur after trauma, with contact lens wear, or after refractive surgery.\u003csup\u003e\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e,\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e\u003c/sup\u003e An outbreak of \u003cem\u003eFusarium\u003c/em\u003e keratitis among contact lens wearers was related to the ReNu Moistureloc\u0026trade; contact lens solution, which was subsequently removed from the market.\u003csup\u003e\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eThe best treatment strategies for fungal keratitis have not been well characterized. Topical natamycin, a polyene, is the only antifungal agent approved by the Food and Drug Administration (FDA) for treatment of fungal keratitis. The Mycotic Ulcer Treatment Trials (MUTT) I and II, were two NEI-funded randomized double-masked clinical trials that found topical natamycin to be superior to topical voriconazole and no additional benefit of adjuvant oral voriconazole. Two recent randomized clinical trials also failed to demonstrate a benefit of adjuvant intrastromal voriconazole or adjuvant UVX in the treatment fungal keratitis.\u003csup\u003e\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e,\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e\u003c/sup\u003e However, natamycin is fungistatic and has limited penetration into the corneal layers.\u003csup\u003e\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e\u003c/sup\u003e Furthermore, outcomes of fungal keratitis with topical natamycin are extremely poor as demonstrated in MUTT II where approximately 50% of patients had full thickness corneal perforation or required TPK despite topical natamycin, topical voriconazole, and adjuvant oral voriconazole.\u003c/p\u003e \u003cp\u003eAlthough much less common, acanthameoba keratitis (AK) may have the most prolonged and severe course of any corneal infection. AK is typically related to contact lens use and the incidence of these infections varies from as low as 1% to 4\u0026ndash;8% of culture positive microbial keratitis cases in countries where contact lens use is common.\u003csup\u003e\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e\u003c/sup\u003e Topical biguanides such as chlorhexidine 0.02% and Polyhexamethylene biguanide (PHMB) 0.02% are thought to be the most effective available medical therapy. However, large series suggest that only 60% of patients achieve complete cure with medical therapy alone by one year and that almost 50% end up with a poor outcome, defined as requiring TPK or having visual acuity less than 20/80.\u003csup\u003e49\u003c/sup\u003e Furthermore, these medications are highly toxic and cause permanent damage to delicate ocular structures such as limbal stem cells and trabecular meshwork.\u003csup\u003e\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eCorneal cross-linking (CXL) is a novel prospective therapy that may simultaneously reduce both ocular pathogens and inflammatory cells and strengthen the cornea.\u003csup\u003e\u003cspan additionalcitationids=\"CR2\" citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e CXL with riboflavin (UVX) and Rose Bengal with green light (RB-PDT) are both effective \u003cem\u003ein vitro\u003c/em\u003e against common bacterial ocular pathogens, such as \u003cem\u003ePseudomonas aeruginosa\u003c/em\u003e and \u003cem\u003eStreptococcus pneumoniae\u003c/em\u003e.\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e,\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e\u003c/sup\u003e However, UVX appears to have much less effect on fungal and acanthamoeba organisms \u003cem\u003ein vitro\u003c/em\u003e and one randomized clinical trial also did not show a benefit of adjuvant UVX in the treatment of filamentous fungal keratitis patients.\u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e\u003cdiv class=\"BlockQuote\"\u003e\u003cp\u003e \u003cem\u003eIn vitro\u003c/em\u003e RB-PDT, appears to be much more effective against fungal and acanthamoeba isolates.\u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e,\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e Rose Bengal (RB) is one of the most commonly used dyes in the diagnosis of ocular surface disease.\u003csup\u003e\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e\u003c/sup\u003e Rose Bengal is an effective photosensitizer, readily converting triplet oxygen (\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003eO\u003csub\u003e2\u003c/sub\u003e) to produce high singlet oxygen (\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003eO\u003csub\u003e2\u003c/sub\u003e) yields with exposure to green light.\u003csup\u003e\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e\u003c/sup\u003e Although RB dye penetration is to approximately 100\u0026micro;m into the stroma, subsequent free radical formation occurs up to 1/3 of the corneal stromal depth.\u003csup\u003e\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e,\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e\u003c/sup\u003e The ability of RB to continue free radical formation is self-limited after photo-irradiation has ceased.\u003csup\u003e\u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e\u003c/sup\u003e Multiple \u003cem\u003eIn vitro\u003c/em\u003e and \u003cem\u003eex vivo\u003c/em\u003e studies have suggested that RGX may be safer than UVX. Wound healing studies found more corneal haze and slower wound healing after UVX compared with RGX.\u003csup\u003e\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u003c/sup\u003e Rabbit studies have demonstrated the safety of RGX on limbal stem cells and endothelium and found anterior stromal keratocyte damage in RGX comparable to epithelial debridement alone.\u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e,\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e By contrast, UVX causes an immediate decrease in the sub-epithelial nerve plexus and loss of keratocytes in the anterior one-third of the corneal stroma, although this recovers after a few months.\u003csup\u003e\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e,\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eSmear and culture negative ulcers represent another therapeutic challenge for clinicians. Up to 60% of corneal cultures are smear and culture negative.\u003csup\u003e\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e\u003c/sup\u003e When these patients do not improve with topical antibiotics alone, clinicians must decide what alternative medical therapy to introduce. There is little guidance in the literature on how to manage these patients. These cases are challenging to study since they represent different underlying etiologies and one medical therapy is unlikely to address all of them. RB-PDT is unique in its potential to address bacterial, fungal and parasitic infections making it a particularly attractive novel therapy.\u003c/p\u003e \u003cp\u003eLimitations to our study include the fact that while our study design is practical in terms of resources, the organisms have different clinical courses will likely respond differently to RB-PDT. It is true that these cases are typically not studied together since one medical therapy is unlikely to address all of the underlying etiologies. RB-PDT is unique in its potential to address bacterial, fungal and parasitic infections, making it possible to enroll all of these ulcers in one study. This also makes RB-PDT a particularly appealing therapy for smear and culture negative cases, which are common and are a therapeutic challenge for clinicians.\u003csup\u003e\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e\u003c/sup\u003e Here, we propose pre-specified subgroup analyses for acanthameoba, fungal, and smear/culture negative, to analyze the effects of RB-PDT on each type of ulcer.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eHere we explore a novel adjuvant therapy for the treatment of fungal, acanthameoba and smear and culture negative keratitis. Reducing the global burden of vision loss from corneal opacification will likely require a multidisciplinary approach including corneal ulcer prevention, novel antimicrobial agents and adjuvant therapies such as RB-PDT.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTrial status\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eProtocol version 3.0 last edited March 15\u003csup\u003eth\u003c/sup\u003e 2023. Recruitment began in January 2022 and is expected to last until approximately January 2025.\u0026nbsp;\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eDeclarations\u003c/h2\u003e \u003cp\u003e \u003cstrong\u003eEthics Approval and Consent to Participate\u003c/strong\u003e \u003cp\u003eSingle IRB approval for US sites was obtained at the University of California, San Francisco (#1826045). Institutional IRB approval was obtained at the Aravind Eye Hospital, Madurai (2020009CLI). Council on Medical Research (ICMR) approval was also obtained for the study (011-26589492).\u003c/p\u003e \u003c/p\u003e\u003cp\u003e \u003ch2\u003eCompeting Interests\u003c/h2\u003e \u003cp\u003eThere are no conflicts of interest to report.\u003c/p\u003e \u003c/p\u003e\u003ch2\u003eFunding\u003c/h2\u003e \u003cp\u003eThis work was supported by grants UG1 EY028518 (Lietman/Rose-Nussbaumer) and K23 EY025025 (Rose-Nussbaumer) from the National Eye Institute an institutional P30 grant from the National Eye Institute (Stanford) and Research to Prevent Blindness (UCSF \u0026amp; Stanford University).\u003c/p\u003e\u003ch2\u003eAuthors\u0026rsquo; contributions\u003c/h2\u003e \u003cp\u003eTom Lietman, Jennifer Rose-Nussbaumer, NV Prajna and N Radhakrishnan have contributed to the design of the study and study implementation and these collaborators make up the steering committee. Tom Lietman is the head of the data coordinating center. Jennifer Rose-Nussbaumer is the head of the clinical coordinating center. P Lalitha and AL H\u0026ouml;fling-Lima are contributing to study implementation. Benjamin Arnold is the biostatistician and as such has directed the statistical analysis plan and will have access to the final dataset.\u003c/p\u003e\u003ch2\u003eAcknowledgements\u003c/h2\u003e \u003cp\u003eOur study team would like to thank Drs. Jean-Marie Parel and Guillermo Amescua from the Ophthalmic Biophysics Center at the Bascom Palmer Eye Institute for donation of RB-PDT equipment and their generous support with their technical expertise.\u003c/p\u003e\u003ch2\u003eAvailability of Data and Material\u003c/h2\u003e \u003cp\u003eEvery attempt will be made to publish results in peer-reviewed journals and to present these data at national and international meetings. Consistent with the collaborative nature of the proposed research, the PI anticipates sharing all data generated by the study with collaborators. Analytic datasets that will be developed through the project will comply with the NIH Data Sharing Policy. The analytical datasets from this project will include patient-level data generated from the study visits. Data from the trial will be made available upon reasonable request.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAlio JL, Abbouda A, Valle DD, et al. Corneal cross linking and infectious keratitis: a systematic review with a meta-analysis of reported cases. \u003cem\u003eJournal of ophthalmic inflammation and infection. \u003c/em\u003e2013;3(1):47.\u003c/li\u003e\n\u003cli\u003eSand D, She R, Shulman IA, et al. Microbial keratitis in los angeles: the doheny eye institute and the los angeles county hospital experience. \u003cem\u003eOphthalmology. \u003c/em\u003e2015;122(5):918-924.\u003c/li\u003e\n\u003cli\u003ePapaioannou L, Miligkos M, Papathanassiou M. Corneal Collagen Cross-Linking for Infectious Keratitis: A Systematic Review and Meta-Analysis. \u003cem\u003eCornea. \u003c/em\u003e2016;35(1):62-71.\u003c/li\u003e\n\u003cli\u003eKeating A, Pineda R, 2nd, Colby K. Corneal cross linking for keratoconus. \u003cem\u003eSeminars in ophthalmology. \u003c/em\u003e2010;25(5-6):249-255.\u003c/li\u003e\n\u003cli\u003eLamy R, Netto CF, Reis RG, et al. Effects of corneal cross-linking on contrast sensitivity, visual acuity, and corneal topography in patients with keratoconus. \u003cem\u003eCornea. \u003c/em\u003e2013;32(5):591-596.\u003c/li\u003e\n\u003cli\u003eRaiskup-Wolf F, Hoyer A, Spoerl E, et al. Collagen crosslinking with riboflavin and ultraviolet-A light in keratoconus: long-term results. \u003cem\u003eJournal of cataract and refractive surgery. \u003c/em\u003e2008;34(5):796-801.\u003c/li\u003e\n\u003cli\u003eVinciguerra P, Albe E, Trazza S, et al. Intraoperative and postoperative effects of corneal collagen cross-linking on progressive keratoconus. \u003cem\u003eArchives of ophthalmology. \u003c/em\u003e2009;127(10):1258-1265.\u003c/li\u003e\n\u003cli\u003eZhu H, Alt C, Webb RH, et al. Corneal Crosslinking With Rose Bengal and Green Light: Efficacy and Safety Evaluation. \u003cem\u003eCornea. \u003c/em\u003e2016;35(9):1234-1241.\u003c/li\u003e\n\u003cli\u003eLorenzo-Martin E, Gallego-Munoz P, Ibares-Frias L, et al. Rose Bengal and Green Light Versus Riboflavin-UVA Cross-Linking: Corneal Wound Repair Response. \u003cem\u003eInvest Ophthalmol Vis Sci. \u003c/em\u003e2018;59(12):4821-4830.\u003c/li\u003e\n\u003cli\u003eCherfan D, Verter EE, Melki S, et al. Collagen cross-linking using rose bengal and green light to increase corneal stiffness. \u003cem\u003eInvest Ophthalmol Vis Sci. \u003c/em\u003e2013;54(5):3426-3433.\u003c/li\u003e\n\u003cli\u003eBekesi N, Gallego-Munoz P, Ibares-Frias L, et al. Biomechanical Changes After In Vivo Collagen Cross-Linking With Rose Bengal-Green Light and Riboflavin-UVA. \u003cem\u003eInvest Ophthalmol Vis Sci. \u003c/em\u003e2017;58(3):1612-1620.\u003c/li\u003e\n\u003cli\u003eWang T, Zhu L, Zhu J, et al. Subacute effects of rose Bengal/Green light cross linking on rabbit thin corneal stability and safety. \u003cem\u003eLasers Surg Med. \u003c/em\u003e2018;50(4):324-332.\u003c/li\u003e\n\u003cli\u003eNaranjo A, Pelaez D, Arrieta E, et al. Cellular and molecular assessment of rose bengal photodynamic antimicrobial therapy on keratocytes, corneal endothelium and limbal stem cell niche. \u003cem\u003eExp Eye Res. \u003c/em\u003e2019;188:107808.\u003c/li\u003e\n\u003cli\u003eGallego-Munoz P, Ibares-Frias L, Lorenzo E, et al. Corneal Wound Repair After Rose Bengal and Green Light Crosslinking: Clinical and Histologic Study. \u003cem\u003eInvest Ophthalmol Vis Sci. \u003c/em\u003e2017;58(9):3471-3480.\u003c/li\u003e\n\u003cli\u003eVatansever F, de Melo WC, Avci P, et al. Antimicrobial strategies centered around reactive oxygen species--bactericidal antibiotics, photodynamic therapy, and beyond. \u003cem\u003eFEMS Microbiol Rev. \u003c/em\u003e2013;37(6):955-989.\u003c/li\u003e\n\u003cli\u003eHalili F, Arboleda A, Durkee H, et al. Rose Bengal- and Riboflavin-Mediated Photodynamic Therapy to Inhibit Methicillin-Resistant Staphylococcus aureus Keratitis Isolates. \u003cem\u003eAm J Ophthalmol. \u003c/em\u003e2016;166:194-202.\u003c/li\u003e\n\u003cli\u003eDurkee H, Arboleda A, Aguilar MC, et al. Rose bengal photodynamic antimicrobial therapy to inhibit Pseudomonas aeruginosa keratitis isolates. \u003cem\u003eLasers Med Sci. \u003c/em\u003e2019.\u003c/li\u003e\n\u003cli\u003ePrajna NV, Radhakrishnan N, Lalitha P, et al. Cross-Linking-Assisted Infection Reduction: A Randomized Clinical Trial Evaluating the Effect of Adjuvant Cross-Linking on Outcomes in Fungal Keratitis. \u003cem\u003eOphthalmology. \u003c/em\u003e2019.\u003c/li\u003e\n\u003cli\u003eOzbek-Uzman S, Yalniz-Akkaya Z, Burcu A. Corneal Collagen Cross-Linking With Photoactivated Chromophore for Infectious Keratitis After Penetrating Keratoplasty. \u003cem\u003eCornea. \u003c/em\u003e2020;39(3):283-289.\u003c/li\u003e\n\u003cli\u003eKasetsuwan N, Reinprayoon U, Satitpitakul V. Photoactivated Chromophore for Moderate to Severe Infectious Keratitis as an Adjunct Therapy: A Randomized Controlled Trial. \u003cem\u003eAm J Ophthalmol. \u003c/em\u003e2016;165:94-99.\u003c/li\u003e\n\u003cli\u003eSaid DG, Elalfy MS, Gatzioufas Z, et al. Collagen cross-linking with photoactivated riboflavin (PACK-CXL) for the treatment of advanced infectious keratitis with corneal melting. \u003cem\u003eOphthalmology. \u003c/em\u003e2014;121(7):1377-1382.\u003c/li\u003e\n\u003cli\u003eAtalay HT, Dogruman-Al F, Sarzhanov F, et al. Effect of Riboflavin/Rose Bengal-Mediated PACK-CXL on Acanthamoeba Trophozoites and Cysts in Vitro. \u003cem\u003eCurr Eye Res. \u003c/em\u003e2018;43(11):1322-1325.\u003c/li\u003e\n\u003cli\u003eArboleda A, Miller D, Cabot F, et al. Assessment of rose bengal versus riboflavin photodynamic therapy for inhibition of fungal keratitis isolates. \u003cem\u003eAm J Ophthalmol. \u003c/em\u003e2014;158(1):64-70.e62.\u003c/li\u003e\n\u003cli\u003ePrajna NV, Krishnan T, Rajaraman R, et al. Effect of Oral Voriconazole on Fungal Keratitis in the Mycotic Ulcer Treatment Trial II (MUTT II): A Randomized Clinical Trial. \u003cem\u003eJAMA Ophthalmol. \u003c/em\u003e2016;134(12):1365-1372.\u003c/li\u003e\n\u003cli\u003ePrajna NV, Krishnan T, Rajaraman R, et al. Predictors of Corneal Perforation or Need for Therapeutic Keratoplasty in Severe Fungal Keratitis: A Secondary Analysis of the Mycotic Ulcer Treatment Trial II. \u003cem\u003eJAMA Ophthalmol. \u003c/em\u003e2017;135(9):987-991.\u003c/li\u003e\n\u003cli\u003eArboleda A, Miller D, Cabot F, et al. Assessment of rose bengal versus riboflavin photodynamic therapy for inhibition of fungal keratitis isolates. \u003cem\u003eAm J Ophthalmol. \u003c/em\u003e2014;158(1):64-70 e62.\u003c/li\u003e\n\u003cli\u003eNaranjo A, Arboleda A, Martinez JD, et al. Rose Bengal Photodynamic Antimicrobial Therapy (RB-PDAT) for Patients with Progressive Infectious Keratitis: A Pilot Clinical Study. \u003cem\u003eAm J Ophthalmol. \u003c/em\u003e2019.\u003c/li\u003e\n\u003cli\u003eRay KJ, Lalitha P, Prajna NV, et al. The Utility of Repeat Culture in Fungal Corneal Ulcer Management: A Secondary Analysis of the MUTT I Randomized Clinical Trial. \u003cem\u003eAm J Ophthalmol. \u003c/em\u003e2017.\u003c/li\u003e\n\u003cli\u003eBhadange Y, Das S, Kasav MK, et al. Comparison of culture-negative and culture-positive microbial keratitis: cause of culture negativity, clinical features and final outcome. \u003cem\u003eThe British journal of ophthalmology. \u003c/em\u003e2015;99(11):1498-1502.\u003c/li\u003e\n\u003cli\u003eMcLeod SD, Kolahdouz-Isfahani A, Rostamian K, et al. The Role of Smears, Cultures, and Antibiotic Sensitivity Testing in the Management of Suspected Infectious Keratitis. \u003cem\u003eOphthalmology. \u003c/em\u003e1996;103(1):23-28.\u003c/li\u003e\n\u003cli\u003eVemuganti GK, Garg P, Gopinathan U, et al. Evaluation of agent and host factors in progression of mycotic keratitis: A histologic and microbiologic study of 167 corneal buttons. \u003cem\u003eOphthalmology. \u003c/em\u003e2002;109(8):1538-1546.\u003c/li\u003e\n\u003cli\u003eGopinathan U, Garg P, Fernandes M, et al. The epidemiological features and laboratory results of fungal keratitis: a 10-year review at a referral eye care center in South India. \u003cem\u003eCornea. \u003c/em\u003e2002;21(6):555-559.\u003c/li\u003e\n\u003cli\u003eFlorCruz NV, Evans JR. Medical interventions for fungal keratitis. \u003cem\u003eThe Cochrane database of systematic reviews. \u003c/em\u003e2015(4):CD004241.\u003c/li\u003e\n\u003cli\u003eFeilmeier MR, Sivaraman KR, Oliva M, et al. Etiologic diagnosis of corneal ulceration at a tertiary eye center in Kathmandu, Nepal. \u003cem\u003eCornea. \u003c/em\u003e2010;29(12):1380-1385.\u003c/li\u003e\n\u003cli\u003eDunlop AA, Wright ED, Howlader SA, et al. Suppurative corneal ulceration in Bangladesh. A study of 142 cases examining the microbiological diagnosis, clinical and epidemiological features of bacterial and fungal keratitis. \u003cem\u003eAustralian and New Zealand journal of ophthalmology. \u003c/em\u003e1994;22(2):105-110.\u003c/li\u003e\n\u003cli\u003eWong TY, Ng TP, Fong KS, et al. Risk factors and clinical outcomes between fungal and bacterial keratitis: a comparative study. \u003cem\u003eThe CLAO journal : official publication of the Contact Lens Association of Ophthalmologists, Inc. \u003c/em\u003e1997;23(4):275-281.\u003c/li\u003e\n\u003cli\u003eSrinivasan M, Gonzales CA, George C, et al. Epidemiology and aetiological diagnosis of corneal ulceration in Madurai, south India. \u003cem\u003eThe British journal of ophthalmology. \u003c/em\u003e1997;81(11):965-971.\u003c/li\u003e\n\u003cli\u003eWhitcher JP, Srinivasan M. Corneal ulceration in the developing world--a silent epidemic. \u003cem\u003eThe British journal of ophthalmology. \u003c/em\u003e1997;81(8):622-623.\u003c/li\u003e\n\u003cli\u003eDeorukhkar S, Katiyar R, Saini S. Epidemiological features and laboratory results of bacterial and fungal keratitis: a five-year study at a rural tertiary-care hospital in western Maharashtra, India. \u003cem\u003eSingapore medical journal. \u003c/em\u003e2012;53(4):264-267.\u003c/li\u003e\n\u003cli\u003eLiesegang TJ, Forster RK. Spectrum of microbial keratitis in South Florida. \u003cem\u003eAmerican journal of ophthalmology. \u003c/em\u003e1980;90(1):38-47.\u003c/li\u003e\n\u003cli\u003eMcLeod SD, Kolahdouz-Isfahani A, Rostamian K, et al. The role of smears, cultures, and antibiotic sensitivity testing in the management of suspected infectious keratitis. \u003cem\u003eOphthalmology. \u003c/em\u003e1996;103(1):23-28.\u003c/li\u003e\n\u003cli\u003eVerma S, Tuft SJ. Fusarium solani keratitis following LASIK for myopia. \u003cem\u003eThe British journal of ophthalmology. \u003c/em\u003e2002;86(10):1190-1191.\u003c/li\u003e\n\u003cli\u003eBernal MD, Acharya NR, Lietman TM, et al. Outbreak of Fusarium keratitis in soft contact lens wearers in San Francisco. \u003cem\u003eArchives of ophthalmology. \u003c/em\u003e2006;124(7):1051-1053.\u003c/li\u003e\n\u003cli\u003eYildiz EH, Abdalla YF, Elsahn AF, et al. Update on fungal keratitis from 1999 to 2008. \u003cem\u003eCornea. \u003c/em\u003e2010;29(12):1406-1411.\u003c/li\u003e\n\u003cli\u003ePrajna NV, Radhakrishnan N, Lalitha P, et al. Cross-Linking-Assisted Infection Reduction: A Randomized Clinical Trial Evaluating the Effect of Adjuvant Cross-Linking on Outcomes in Fungal Keratitis. \u003cem\u003eOphthalmology. \u003c/em\u003e2020;127(2):159-166.\u003c/li\u003e\n\u003cli\u003eNarayana S, Krishnan T, Ramakrishnan S, et al. Mycotic Antimicrobial Localized Injection: A Randomized Clinical Trial Evaluating Intrastromal Injection of Voriconazole. \u003cem\u003eOphthalmology. \u003c/em\u003e2019;126(8):1084-1089.\u003c/li\u003e\n\u003cli\u003eO\u0026apos;Day DM, Head WS, Robinson RD, et al. Corneal penetration of topical amphotericin B and natamycin. \u003cem\u003eCurrent eye research. \u003c/em\u003e1986;5(11):877-882.\u003c/li\u003e\n\u003cli\u003eDart JK, Saw VP, Kilvington S. Acanthamoeba keratitis: diagnosis and treatment update 2009. \u003cem\u003eAm J Ophthalmol. \u003c/em\u003e2009;148(4):487-499.e482.\u003c/li\u003e\n\u003cli\u003ePapa V, Rama P, Radford C, et al. Acanthamoeba keratitis therapy: time to cure and visual outcome analysis for different antiamoebic therapies in 227 cases. \u003cem\u003eBr J Ophthalmol. \u003c/em\u003e2019.\u003c/li\u003e\n\u003cli\u003eCarrijo-Carvalho LC, Sant\u0026apos;ana VP, Foronda AS, et al. Therapeutic agents and biocides for ocular infections by free-living amoebae of Acanthamoeba genus. \u003cem\u003eSurv Ophthalmol. \u003c/em\u003e2017;62(2):203-218.\u003c/li\u003e\n\u003cli\u003eMartins SA, Combs JC, Noguera G, et al. Antimicrobial efficacy of riboflavin/UVA combination (365 nm) in vitro for bacterial and fungal isolates: a potential new treatment for infectious keratitis. \u003cem\u003eInvestigative ophthalmology \u0026amp; visual science. \u003c/em\u003e2008;49(8):3402-3408.\u003c/li\u003e\n\u003cli\u003eKhan-Lim D, Berry M. Still confused about rose bengal? \u003cem\u003eCurr Eye Res. \u003c/em\u003e2004;29(4-5):311-317.\u003c/li\u003e\n\u003cli\u003eWachter E, Dees C, Harkins J, et al. Topical rose bengal: pre-clinical evaluation of pharmacokinetics and safety. \u003cem\u003eLasers Surg Med. \u003c/em\u003e2003;32(2):101-110.\u003c/li\u003e\n\u003cli\u003eAmescua G, Arboleda A, Nikpoor N, et al. Rose Bengal Photodynamic Antimicrobial Therapy: A Novel Treatment for Resistant Fusarium Keratitis. \u003cem\u003eCornea. \u003c/em\u003e2017;36(9):1141-1144.\u003c/li\u003e\n\u003cli\u003eTonogai Y, Ito Y, Iwaida M, Tati M, et al. Studies on the toxicity of coal-tar dyes. III. Reason of acute toxicity to fish caused by coal-tar dyes and their industrial effluents. \u003cem\u003eJ Toxicol Sci. \u003c/em\u003e1980;5(1):23-33.\u003c/li\u003e\n\u003cli\u003eMazzotta C, Hafezi F, Kymionis G, et al. In Vivo Confocal Microscopy after Corneal Collagen Crosslinking. \u003cem\u003eOcul Surf. \u003c/em\u003e2015;13(4):298-314.\u003c/li\u003e\n\u003cli\u003eSharma N, Suri K, Sehra SV, et al. Collagen cross-linking in keratoconus in Asian eyes: visual, refractive and confocal microscopy outcomes in a prospective randomized controlled trial. \u003cem\u003eInt Ophthalmol. \u003c/em\u003e2015;35(6):827-832.\u003c/li\u003e\n\u003cli\u003ePeng MY, Cevallos V, McLeod SD, et al. Bacterial Keratitis: Isolated Organisms and Antibiotic Resistance Patterns in San Francisco. \u003cem\u003eCornea. \u003c/em\u003e2018;37(1):84-87.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"trials","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"trls","sideBox":"Learn more about [Trials](http://trialsjournal.biomedcentral.com/)","snPcode":"13063","submissionUrl":"https://www.editorialmanager.com/trls","title":"Trials","twitterHandle":"MedicalEvidence","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Infectious keratitis, cornea, RB-PDT","lastPublishedDoi":"10.21203/rs.3.rs-4165312/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-4165312/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cem\u003e\u003cstrong\u003eBackground:\u0026nbsp;\u003c/strong\u003e\u003c/em\u003eInfectious keratitis secondary to fungus or acanthamoeba often has a poor outcome despite receiving the best available medical therapy.\u0026nbsp;\u003cem\u003eIn vitro\u003c/em\u003e\u0026nbsp;Rose Bengal Photodynamic therapy (RB-PDT) appears to be effective against fungal and acanthamoeba isolates.\u003csup\u003e22,23\u003c/sup\u003e\u0026nbsp;In one published series RB-PDT reduced the need for therapeutic penetrating keratoplasty in severe bacterial, fungal, and acanthameoba keratitis not responsive to medical therapy.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods: \u003c/strong\u003eThis international, randomized, sham and placebo controlled 2-arm clinical trial, randomizes patients with smear positive fungal and acanthameoba and smear negative corneal ulcers in a 1:1 fashion to one of two treatment arms: 1) Topical antimicrobial plus sham RB-PDT or 2) Topical antimicrobial plus RB-PDT\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e\u003cstrong\u003eDiscussion:\u003c/strong\u003e\u003c/em\u003e\u0026nbsp;We anticipate that RB-PDT will improve best spectacle corrected visual acuity and also reduce complications such as corneal perforation and the need for therapeutic penetrating keratoplasty. This study will comply with the NIH Data Sharing Policy and Policy on the Dissemination of NIH-Funded Clinical Trial Information and the Clinical Trials Registration and Results Information Submission rule. Our results will be disseminated via clinicaltrials.gov website, meetings, and journal publications. Our data will also be available upon reasonable request.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTrial Registration: \u003c/strong\u003eNCT, NCT05110001, Registered November 5, 2021.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003ehttps://www.clinicaltrials.gov/study/NCT05110001\u0026nbsp;\u003c/p\u003e","manuscriptTitle":"A double-masked, sham-controlled trial of rose bengal photodynamic therapy for the treatment of fungal and acanthameoba keratitis: Rose Bengal Electromagnetic Activation with Green Light for Infection Reduction (REAGIR) Study","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-07-02 20:42:16","doi":"10.21203/rs.3.rs-4165312/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Major revision","date":"2024-06-11T18:41:47+00:00","index":"","fulltext":""},{"type":"reviewerAgreed","content":"","date":"2024-06-11T09:48:33+00:00","index":0,"fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-06-10T12:18:33+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-04-05T10:24:25+00:00","index":"","fulltext":""},{"type":"submitted","content":"Trials","date":"2024-03-25T15:12:09+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"trials","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"trls","sideBox":"Learn more about [Trials](http://trialsjournal.biomedcentral.com/)","snPcode":"13063","submissionUrl":"https://www.editorialmanager.com/trls","title":"Trials","twitterHandle":"MedicalEvidence","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"e193450e-424c-4079-957e-d2b724006ddf","owner":[],"postedDate":"July 2nd, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2024-09-02T16:00:54+00:00","versionOfRecord":{"articleIdentity":"rs-4165312","link":"https://doi.org/10.1186/s13063-024-08376-3","journal":{"identity":"trials","isVorOnly":false,"title":"Trials"},"publishedOn":"2024-08-28 15:57:23","publishedOnDateReadable":"August 28th, 2024"},"versionCreatedAt":"2024-07-02 20:42:16","video":"","vorDoi":"10.1186/s13063-024-08376-3","vorDoiUrl":"https://doi.org/10.1186/s13063-024-08376-3","workflowStages":[]},"version":"v1","identity":"rs-4165312","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-4165312","identity":"rs-4165312","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}