Variations in irradiation energy and rose bengal concentration for photodynamic antimicrobial therapy of fungal keratitis isolates.

The purpose is to assess the efficacy of rose bengal photodynamic antimicrobial therapy (PDAT) using different irradiation energy levels and photosensitizer concentrations for the inhibition of fungal keratitis isolates. Seven different fungi (Aspergillus fumigatus, Candida albicans, Curvularia lunata, Fusarium keratoplasticum, Fusarium solani, Paecilomyces variotii, and Pseudallescheria boydii) were isolated from patients with confirmed infectious keratitis. Experiments were performed in triplicate with suspensions of each fungus exposed to different PDAT parameters including a control, green light exposure of 5.4 J/cm2, 2.7 J/cm2 (continuous and pulsed), and 1.8 J/cm2 and rose bengal concentrations of 0.1%, 0.05%, and 0.01%. Plates were photographed 72 h after experimentation, and analysis was performed to assess fungal growth inhibition. PDAT using 5.4 J/cm2 of irradiation and 0.1% rose bengal completely inhibited growth of five of the seven fungal species. Candida albicans and Fusarium keratoplasticum were the most susceptible organisms, with growth inhibited with the lowest fluence and minimum rose bengal concentration. Fusarium solani, Pseudallescheria boydii, and Paecilomyces variotii were inhibited by lower light exposures and photosensitizer concentrations. Aspergillus fumigatus and Curvularia lunata were not inhibited by any PDAT parameters tested. Continuous and pulsed irradiation using 2.7 J/cm2 produced similar results. Rose bengal PDAT successfully inhibits the in vitro growth of five fungi known to cause infectious keratitis. Differences in growth inhibition of the various fungi to multiple PDAT parameters suggest that susceptibilities to PDAT are unique among fungal species. These findings support modifying PDAT parameters based on the infectious etiology.

Subpalpebral Antibiotic Lavage for the Treatment of Refractory Infectious Scleritis.

Infectious scleritis is a potentially devastating condition that can result in severe vision loss. When traditional management fails, a subpalpebral antibiotic lavage system (SAL) can be considered to bathe the infected area with a high volume and concentration of antibiotics. Several reports show that this method can be curative for infectious scleritis, otherwise refractory to care. However, surgical approaches for this technique are either not well described, advocate for transecting the levator aponeurosis, or do not expose the entire width of the superior fornix, which can lead to postoperative ptosis. The authors describe a case where a novel approach was utilized, using a fenestrated angiocather in the superior lateral fornix, to maximize outcomes and minimize postoperative complications. In the setting of infectious scleritis refractory to traditional management, a SAL can be safely and effectively placed with the technique detailed in this report.

Role of corneal biopsy in the management of infectious keratitis.

PURPOSE OF REVIEW: The aim of this study was to review the existing literature and investigate the role of microbiologic culture and histopathologic examination of corneal biopsies in the management of infectious keratitis. RECENT FINDINGS: Corneal biopsy continues to be a significantly useful tool in the diagnosis and tailored management of infectious keratitis. Several techniques can be employed for tissue collection, handling and processing to optimize diagnostic yield and maximize safety, including emerging femtosecond laser-assisted biopsy. SUMMARY: Corneal opacities represent a significant cause of global blindness, and infectious keratitis is the most common cause. Organism identification in progressive infectious keratitis is essential for proper management. However, microbiological culture alone has a high rate of false-negative results. Records from the Bascom Palmer Eye Institute were retrospectively searched for patients between 1 January 2015, and 31 December 2019, who underwent corneal biopsy, therapeutic keratoplasty or endothelial graft removal for infectious keratitis and had specimens bisected and submitted for evaluation with both microbiologic culture and histopathologic examination. Detection of bacteria, fungus and mycobacteria was not statistically different between culture and histopathology. Microbiology and histopathology are complementary methods for the identification of causative microorganisms in corneal specimens with presumed infectious keratitis.

Levator Palpebrae Superioris With Inferior Oblique Muscle Flaps for Total Ocular Surface Reconstruction.

A healthy middle-aged man sustained extensive burns after a motor vehicle accident, including fourth-degree burns to the ocular adnexa. He had bilateral eyelid necrosis with complete tissue loss to the orbital rim. Progressive corneal thinning occurred bilaterally despite frequent lubrication, weekly amniotic membrane placement, and a Gunderson flap, ultimately requiring evisceration of his OS. In accordance with family wishes to perform all interventions for the OD, he underwent ocular surface reconstruction with levator palpebrae superioris (LPS) and inferior oblique (IO) muscle flaps, supplemented with an orbital fat transfer and bucket-handle muscle flap. These flaps were covered with porcine urinary bladder matrix and remained viable with complete globe coverage postoperatively. Although local eyelid flaps traditionally involve tissue supplied by branches of the external carotid artery, this case suggests that orbital-internal carotid artery based flaps utilizing the LPS and IO muscles may achieve ocular surface coverage after complete periocular and facial soft tissue loss.

Tumor necrosis factor-alpha and interferon-gamma induce inflammasome-mediated corneal endothelial cell death.

PURPOSE: Chronic corneal endothelial cell (CEC) loss results in corneal edema and vision loss in conditions such as pseudophakic bullous keratopathy (PBK), Fuchs' dystrophy, and corneal graft failure. Low CEC density has been associated with an elevation of intraocular pro-inflammatory cytokines such as tumor necrosis factor (TNF)-α and interferon (INF)-γ. These cytokines are capable of triggering pyroptosis, a programmed cell death mechanism mediated by the inflammasome, prompting the activation of the pro-inflammatory cytokine interleukin (IL)-1ß, the perpetuation of inflammation, and subsequent damage of corneal endothelial tissue. Therefore, the purpose of this study was to determine the deleterious contribution of the inflammasome and pyroptosis to CEC loss. METHODS: CECs from human donor corneas were treated ex vivo with TNF-α and IFN-γ for 48 h. Levels of caspase-1 and IL-1ß were then assayed by ELISA, and the expression of caspase-1 and gasdermin-D (GSDM-D) were confirmed by immunofluorescence. Endothelial cell damage was analyzed by a lactate dehydrogenase (LDH) release assay, and oxidative stress was determined by measuring the levels of reactive oxygen species (ROS) in the culture media. RESULTS: Inflammasome activation and oxidative stress were elevated in CECs following exposure to TNF-α and IFN-γ, which resulted in cell death by pyroptosis as determined by LDH release which was inhibited by the caspase-1 inhibitor Ac-YVAD-cmk. CONCLUSION: CEC death is induced by the pro-inflammatory cytokines TNF-α and IFN-γ, which contribute to inflammasome activation. Moreover, the inflammasome is a promising therapeutic target for the treatment of chronic CEC loss.

Rose bengal photodynamic antimicrobial therapy to inhibit Pseudomonas aeruginosa keratitis isolates.

To evaluate the in vitro efficacy of rose bengal and riboflavin photodynamic antimicrobial therapy for inhibition the growth of four Pseudomonas aeruginosa (P. aeruginosa) isolates. Four different clinical P. aeruginosa isolates were collected from patients with confirmed keratitis. Each strain was mixed with either sterile water, 0.1% riboflavin solution, or 0.1% rose bengal solution to yield a final bacteria concentration of 1.5 × 107 CFU/mL. Aliquots from each suspension were plated onto nutrient agar in triplicate. Plates were separated into two groups: (1) no irradiation and (2) 5.4 J/cm2 of radiant exposure with custom-made LED irradiation sources. Separate irradiation sources were used for each photosensitizer. The riboflavin groups used a UV-A light source (375 nm) and rose bengal groups used a green light source (525 nm). Plates were photographed at 72 h and custom software measured bacterial growth inhibition. Growth inhibition to riboflavin and rose bengal PDAT showed strain-dependent variability. All four strains of P. aeruginosa showed greatest growth inhibition (89-99%) in the green irradiated-rose bengal group. The UV-A-irradiated riboflavin showed inhibition of 24-44%. UV-A irradiation only showed minimal inhibition (7-14%). There was little inhibitory effect in the non-irradiated photosensitizer groups. Rose bengal PDAT had the greatest inhibitory effect on all four P. aeruginosa isolates. In the UV-A-irradiated riboflavin group, there was moderate inhibition within the irradiation zone; however, there was no inhibition in the non-irradiated groups. These results suggest that rose bengal PDAT may be an effective alternative treatment for Pseudomonas aeruginosa infections.

Cellular and molecular assessment of rose bengal photodynamic antimicrobial therapy on keratocytes, corneal endothelium and limbal stem cell niche.

Rose Bengal Photodynamic Antimicrobial Therapy (RB-PDAT) is a novel potential treatment for progressive infectious keratitis. The principle behind this therapy is using Rose Bengal as a photosensitizer that can be activated by green light and results in the production of oxygen free radicals which in turn eradicate the microorganism. Given RB-PDAT's mechanism of action and the potential cytotoxic effects, concerns regarding the safety of this technique have arisen. The purpose of this study was to evaluate the effect of RB-PDAT on keratocytes, while focusing on the safety profile that the photo-chemical reaction has on the limbal stem cell (LSC) niche and endothelial cell layer of the treated cornea. To perform RB-PDAT, Rose Bengal solution (0.1% RB in BSS) was applied to the right cornea of rabbits for 30 min and then irradiated by a custom-made green LED light source (525 nm, 6 mW/cm2) for 15 min (5.4 J/cm2). Three rabbits were sacrificed and enucleated after 24 h for evaluation. TUNEL assay and immunohistochemistry for endothelium and limbal stem cell viability were performed on whole mounts and frozen sections in treated and control eyes. LSC of both eyes were isolated and cultured to perform MTT viability and proliferation, and scratch wound healing assays under time-lapse microscopy. Interestingly, while Rose Bengal dye penetration was superficial, yet associated cellular apoptosis was evidenced in up to 1/3 of the stromal thickness on frozen sections. TUNEL assay on whole mounts showed no endothelial cell death following treatment. Immunohistochemistry on frozen sections of LSC displayed no structural difference between treated and non-treated eyes. There was no difference in LSC proliferation rates and scratch wound healing assay demonstrated adequate cell migration from treated and non-treated eyes. The current study suggests that even though penetration of the RB dye has been shown to be limited, oxidative stress produced by RB-PDAT can reach deeper into the corneal stroma. Nevertheless, our results show that performing RB-PDAT is safe on the corneal endothelium and has no effect on LSC viability or function.

Transcultural validation of the 5-Item Dry Eye Questionnaire for the Mexican population.

PURPOSE: To translate and validate a Spanish-language adaptation of the 5-Item Dry Eye Questionnaire for the Mexican population. METHODS: Based on the MAPI Institute guidelines, the linguistic validation procedures consisted of four steps. Every step was reviewed by the committee. The translated validated questionnaire was applied to 25 Mexican subjects. The questionnaires were completed by the same subjects at three time points, 8 h apart on the same day and then 3 days later. Sensitivity and specificity of the DEQ-5 to predict DE signs was subsequently estimated in 200 patients see in the Asociación para Evitar la Ceguera ophthalmology clinic. RESULTS: During the forward translation step analysis, the committee decided to change the severity scale, as the words "constantly" and "frequently" are synonymous in Spanish, so it was modified by changing "constantly" to "always" for better understanding. Overall, the intra-test intra-class correlation coefficient from tests administered on the same day was 0.9 (95% CI 0.77-0.95, p = 0.0005). The intra-test intra-class correlation coefficient from tests administered 3 days apart was 0.9 (95% CI 0.88-0.97, p = 0.0005). When applying the questionnaire to 200 patients seen in an eye clinic, we found a sensitivity of 76% and a specificity of 31% for a DEQ-5 score of ≥ 6, against 2 or more positive signs of dry eye. CONCLUSION: MAPI methodology proved to be a reliable strategy for the transcultural Dry Eye Questionnaire for translation from English to Spanish for the Mexican population.

Impact of Total Pars Plana Vitrectomy on Postoperative Complications in Aphakic, Snap-On, Type 1 Boston Keratoprosthesis.

PURPOSE: To determine the impact of total pars plana vitrectomies (PPVs) with peripheral shaving of the vitreous base on the rates of postoperative complications in patients with aphakic, snap-on type I Boston keratoprostheses (KPros). DESIGN: Retrospective, consecutive case series. PARTICIPANTS: A total of 48 eyes in 46 patients with implantation of aphakic, snap-on type 1 Boston KPros performed at a tertiary care facility between January 1, 2007, and December 31, 2013, were included. METHODS: The cumulative incidences of postoperative complications were compared between patients who underwent total PPVs with shaving of the vitreous base (n = 22) and those who had partial PPVs or anterior vitrectomies (AVs) at the time of KPro implantation (n = 26). MAIN OUTCOME MEASURES: Rates of complications between patients who underwent total PPVs and partial PPVs or AVs. RESULTS: The rate of total postoperative complications was lower in the total PPV group (P = 0.018, log-rank test). In particular, eyes that underwent total PPVs had lower rates of retroprosthetic membranes (RPMs) requiring intervention (P = 0.049) and less vision loss due to glaucoma progression (P = 0.046). There was also a trend for fewer corneal melts (P = 0.060) and less sight-threatening complications (P = 0.051) in the total vitrectomy group. There was no difference in the rates of KPro extrusion (P = 0.41), endophthalmitis or vitritis (P = 0.15), retinal detachments (P = 0.76), cystoid macular edema (P = 0.83), or timing of complications between the 2 groups. The mean preoperative and postoperative visual acuities were similar between the 2 groups (P = 0.97). The mean follow-up was 49±22 months. CONCLUSIONS: Eyes that underwent total PPVs during implantation of aphakic, snap-on, type I Boston KPros had less postoperative complications than eyes with partial PPVs or AVs during the average 4 years of follow-up.