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Purpose: Microbial keratitis (MK) is a major cause of corneal blindness worldwide. Variations in season and temperature can affect MK incidence due to specific causative organisms; however, few studies have examined these factors in the UK., Methods: Retrospective review of all corneal scrapes from patients with MK presenting to Manchester Royal Eye Hospital, UK, between January 2004 and December 2015. Manchester's monthly temperature data were obtained from Met Office UK. Analysis was performed using logistic regression., Results: From 4229 corneal scrapes, 1539 organisms grew (90.6% bacteria, 7.1% fungi, and 2.3% Acanthamoebae sp.). Gram-positive bacteria grew with increasing temperature [odds ratio (OR) 1.62, 95% CI: 1.11-2.39, P = 0.014], and fungi grew with decreasing temperature (OR 0.29, 95% CI: 0.16-0.51, P < 0.001). Moraxella sp. grew with decreasing temperature (OR 0.91, 95% CI: 0.86-0.96, P = 0.001). Compared with winter, overall culture positivity was significantly less likely in summer (OR 0.57, 95% CI: 0.38-0.87, P = 0.008) and spring (OR 0.65, 95% CI: 0.43-0.99, P = 0.045). Gram-negative bacteria were more likely in summer (OR 1.48, 95% CI: 1.06-2.09, P = 0.022) and autumn (OR 1.75, 95% CI: 1.24-2.47, P = 0.001). Candida sp. were less likely in summer (OR 0.25, 95% CI: 0.07-0.82, P = 0.027) and autumn (OR 0.18, 95% CI: 0.05-0.62, P = 0.009), and Acanthamoeba sp. were less likely in summer (OR 0.39, 95% CI: 0.15-0.92, P = 0.037) and spring (OR 0.26, 95% CI: 0.08-0.69, P = 0.011)., Conclusions: Herein we report variation in the incidence of MK-causing organisms by season and temperature; this finding may aid clinicians in predicting possible causative organisms for MK at differing times of the year.

Purpose: Fungal keratitis is a major cause of corneal ulcers, resulting in significant visual impairment and blindness. Fenretinide, a derivative of vitamin A, has been shown to suppress inflammation in a multitude of diseases. In this study, we aimed to characterize the effect of fenretinide in Aspergillus fumigatus keratitis of the eye in a mouse model., Methods: In vivo and in vitro experiments were performed in mouse models and THP-1 macrophage cell cultures infected with A. fumigatus, respectively. Experimental subjects were first pretreated with fenretinide, and then the effect of the compound was assessed with clinical evaluation, neutrophil staining, myeloperoxidase assay, quantitative polymerase chain reaction (qRT-PCR), and western blot., Results: We confirmed that fenretinide contributed to protection of corneal transparency during early mouse A. fumigatus keratitis by reducing neutrophil recruitment, decreasing myeloperoxidase (MPO) levels and increasing apoptosis. Compared with controls, fenretinide impaired proinflammatory cytokine interleukin 1 beta (IL-1β) production in response to A. fumigatus exposure with contributions by lectin-type oxidized LDL receptor 1 (LOX-1) and c-Jun N-terminal kinase (JNK)., Conclusions: Together, these findings demonstrate that fenretinide may suppress inflammation through reduced neutrophil recruitment and inflammatory cytokine production in A. fumigatus keratitis.

Objective: To investigate the role of NLRP12 in regulating Pseudomonas aeruginosa (P. aeruginosa) keratitis., Materials and Methods: Real-Time-PCR and Western blot were performed to measure the NLRP12 level in corneas and bone marrow-derived macrophages (BMDMs) of C57BL/6 (B6) mice. B6 mice received a subconjunctival injection of lentivirus expressing active NLRP12 (NLRP12-lentivirus) or Ctl-lentivirus (as control), followed by infection of P. aeruginosa. The clinical score, slit lamp and bacterial plate count of mice were evaluated. In addition, myeloperoxidase (MPO) was detected to assess the infiltration of polymorphonuclear neutrophil (PMN). Cytokine levels were measured by Real Time-PCR and ELISA. Meanwhile, the bacterial burden was also evaluated. The activation of NF-κB signaling was determined by pIκBα/IκBα levels based on Western blot and NF-κB-dependent Luciferase activity on the basis of Luciferase assays using 293T cells., Results: NLRP12 mRNA and protein levels were decreased in B6 corneas and BMDMs after P. aeruginosa infection. The over-expression of NLRP12 in B6 corneas significantly ameliorated the severity of corneal disease, bacterial burden, PMN infiltration and pro-inflammatory cytokine expression. In vitro analysis demonstrated that the up-regulation of NLRP12 suppressed pro-inflammatory cytokine production and enhanced bacterial clearance in RAW264.7 cells. The protein levels of pIκBα and IκBα were significantly decreased after NLRP12-lentivirus treatment compared with that of Ctl-lentivirus. NF-κB-dependent Luciferase activity was potently inhibited by NLRP12 infected with P. aeruginosa or cotransfected with the downstream signaling molecules including IKKα and IKKβ in 293T cells., Conclusions: NLRP12 decreases the severity of P. aeruginosa keratitis, reduces corneal inflammation and bacterial burden through the down-regulation of the NF-κB signaling pathway.

Purpose: To report an unusual case of isolated Acremonium eumycetoma presenting as a protuberant mass over the cornea., Methods: Case report and literature review., Results: A 55-year-old male patient referred to our center with a case of perforated corneal ulcer with uveal tissue prolapse was examined in the casualty department and found to have central melt, approximately 8 mm, along with suspected uveal tissue prolapse. A provisional diagnosis of sloughed corneal ulcer with uveal prolapse was made along with differential diagnoses of fungal ball and infected foreign body granuloma. Tectonic penetrating keratoplasty under general anesthesia was planned. Intraoperatively, the suspected uveal (brown colored) tissue was found to be an epicorneal mass growing over an intact and infiltrated cornea. Histopathological and microbiological analysis of the epicorneal mass and host cornea revealed it to be a fungal ball (mass full of septate hyphae) with growth of Acremonium species on culture. The patient was administered topical and oral antifungal agents postoperatively, in addition to topical antibiotics and cycloplegics., Conclusions: Isolated corneal Acremonium eumycetoma masquerading as a perforated corneal ulcer with prolapsed uveal tissue is a rare entity. Surgical intervention and appropriate antimicrobial therapy are key to successful outcome.

Purpose: The incidence of postkeratoplasty fungal infection is increasing in the United States, and our most commonly used corneal storage medium, Optisol-GS, contains antibiotics but no antifungal agents. We previously demonstrated the efficacy of amphotericin B additives in eliminating Candida albicans contaminants in Optisol-GS. The purpose of this study was to determine whether amphotericin B would also be efficacious against Fusarium solani and Aspergillus fumigatus., Methods: Vials of Optisol-GS were supplemented with 0.255 μg/mL of amphotericin B. Half of the vials were inoculated with F. solani and half with A. fumigatus. Positive control vials were inoculated with the fungi but no amphotericin B. The vials were refrigerated, sampled, and plated at different time points. The plates were then incubated at 36°C for 48 hr after which fungal colony counts were performed., Results: There was an average reduction in the growth of F. solani in the amphotericin B-supplemented vials of 44% on day 2, 79% on day 7, and 80% on day 14 when compared with the positive control vials. There was an average reduction in the growth of A. fumigatus in the amphotericin B-supplemented vials of 40% on day 2 and 14% on day 7 when compared with the positive control vials. Both amphotericin B-supplemented and control vials grew less than 2 colonies of A. fumigatus on day 14., Conclusions: This study suggests that amphotericin B additives in Optisol-GS reduce the growth of F. solani and A. fumigatus.

Purpose: To investigate voriconazole (VRZ) penetration and fungal load in the cornea after applying VRZ therapy with various treatment combinations in a fungal keratitis model., Methods: Fifty-four eyes of 27 young albino rabbits were provided for this experimental study. Twelve corneas were inoculated with Candida albicans, 12 corneas were inoculated with Fusarium solani, and 6 eyes were selected as controls. Infected corneas received various treatment combinations including VRZ 1% drop therapy alone, VRZ 1% plus amphotericin B 1% drop combination therapy, iontophoretic VRZ therapy, and VRZ 1% drop therapy after corneal cross-linking. Fungal load was measured by log reduction, and VRZ levels were quantified by liquid chromatography-tandem mass spectrometry., Results: Iontophoresis-assisted VRZ application showed the highest antifungal activity against F. solani keratitis (4-log reduction) and C. albicans keratitis (5-log reduction) compared with other treatment applications. VRZ levels were also found to be the highest in corneas that received iontophoretic VRZ treatment (3.6313 ± 0.0990 ppb for F.solani keratitis and 1.7001 ± 0.0065 ppb for C. albicans keratitis) compared with other treatment applications., Conclusions: Iontophoresis seems to provide the highest VRZ concentration and highest antifungal activity in the cornea compared with other treatment applications for C. albicans and F. solani keratitis.

Aim: To investigate the relationships between corneal suture bacteriology, biofilm and the clinical setting using culture studies and scanning electron microscopy (SEM)., Methods: This is a prospective, observational study of patients with a history of penetrating keratoplasty presenting to a tertiary cornea clinic for routine or symptoms-related corneal suture removal. We documented for each patient the suture clinical setting (quiescent, exposed and keratitis-related), retention time, antimicrobial therapy, bacterial growth on culture studies, and bacterial presence and biofilm coverage on SEM., Results: There were significantly different culture positivity rates between the quiescent (8%), exposed (12%) and keratitis-related (60%) suture groups (p=0.039). As expected, keratitis-related sutures had the longest retention time compared with quiescent and exposed ones (p=0.02). The biofilm coverage score was higher for sutures from the keratitis-related and exposed groups, although this trend was not statistically significant (p=0.90). Higher biofilm scores were seen in samples that also yielded a positive culture result (p=0.36) and in samples with bacterial presence on SEM images (p=0.16 and p=0.73). Both of these were important trends but not statistically significant., Conclusions: Evidence for active bacterial and biofilm presence on corneal sutures was found. Corneal sutures should be considered for removal sooner, before becoming exposed and/or keratitis-related. Traditional culture studies and SEM imaging are helpful in investigating biofilm and its clinical importance. More studies of the spectrum of bacterial growth on embedded biomedical devices such as corneal sutures are needed., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2018. No commercial re-use. See rights and permissions. Published by BMJ.)

Objective: To evaluate microbiological, histological, and ultrastructural characteristics of short-term cryopreserved (STC) canine corneoscleral tissue (<1 year) and to compare it with long-term cryopreserved (LTC) tissue (>6 years)., Animals Studied: Thirty-six healthy canine globes., Procedure: After a decontamination protocol, globes were enucleated and stored at -20 °C. Corneoscleral tissue was evaluated at different periods: <1 year (20 eyes) and >6 years (12 eyes). Four eyes were used as controls. Microbiologic study included direct (blood, McConkey and Sabouraud agars) and enrichment (brain-heart infusion broth) cultures. Cryopreservation artifacts were evaluated by hematoxylin-eosin. Corneoscleral collagen organization and number of normal and dead keratocytes were established by transmission electron microscopy (TEM). Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) was also used for keratocyte characterization., Results: Corneal microbial growth was observed in 25% of the direct STC cultures, and in 47.4% and 16.7% of the enriched STC and LTC cultures, respectively. Scleral STC direct cultures were 30% positive, while enrichment cultures were positive in 66.7% and 16.7% of the STC and LTC, respectively (P = 0.011). Cryopreservation artifacts were higher in LTC tissues (P < 0.001). Apoptotic keratocytes were predominant by TEM and TUNEL, in both STC and LTC. Minimal structural differences were detected in collagen organization between STC and LTC., Conclusions: Cryopreservation of canine corneoscleral tissue seems to reduce bacterial contamination over time. Apoptosis is the main way of death of cryopreserved canine keratocytes. Based on the lack of significant structural differences between STC and LTC samples, these cryopreserved tissues could potentially be used for tectonic support for at least 8 years without structural or microbiological impediment., (© 2017 American College of Veterinary Ophthalmologists.)

Development of efficient ocular drug delivery system for antifungal drugs becomes a must nowadays to face and eradicate the widely spread ophthalmic fungal infections. Itraconazole, a triazole antifungal, is struggling to penetrate the cornea and subsequently, its efficacy is limited. The aim of this study was to enhance itraconazole corneal penetration through utilizing the minimum surfactant amount in presence of β-cyclodextrin which acted as a dissolution and permeation enhancer. β-Cyclodextrin consolidated micellar dispersions (CCMD) were prepared after an initial screening to select the composition of surfactant(s). The preparation was done according to a modified melt dispersion technique. The prepared CCMD were characterized through the analysis of their particle size, zeta potential and solubilization efficiency. The optimum formula was chosen based on a factorial response surface analysis and it was composed of 17:1 w/w surfactant/drug, 30:1 w/w cyclodextrin/drug ratios and 0.02% polyethylene oxide. This formula was subjected to in vitro characterization including release, imaging by transmission electron microscope, mucoadhesion, stability, in addition to the determination of the minimum inhibitory concentration. Moreover, the ex vivo/in vivo permeation, safety and efficacy profiles were determined. The optimized CCMD formula was found to be significantly safe, stable, mucoadhesive and efficient to permeate the drug through rabbits' corneas. Consequently, the optimized CCMD formulation can be a promising, safe and efficient platform for the transcorneal delivery of lipophilic drugs including most antifungals., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Purpose: The aim of the study was to develop a novel formulation of levofloxacin and besifloxacin to achieve improved mucoadhesion and permeability of besifloxacin and levofloxacin through cornea for the effective treatment of ocular infections., Methods: A multicomponent hydrogel formulation containing chitosan-polyvinyl alcohol (PVA)-poly(N-vinylpyrrolidone) (PVP) was designed. Lysophosphatidylcholine was used to enhance corneal penetration of the drugs. The hydrogel preparations were characterized for various parameters, including clarity, pH, viscosity, in vitro release kinetics, mucoadhesion, ex vivo human corneal permeation, and antimicrobial efficacy. The formulations were compared with standard drug solution and marketed eye drops (Besix ® and Levotop ® )., Results: Compared to commercial ophthalmic preparations and free drug solutions, hydrogel formulation of both besifloxacin and levofloxacin was found to have 3.5- and 8-fold higher (P < 0.001) mucoadhesion and superior cumulative corneal permeation. The formulations showed superior in vitro anti-infective properties. Incubation of besifloxacin and levofloxacin formulations with Staphylococcus aureus-infected cornea model for 0.5 h showed greater potency of the hydrogel formulations compared to the marketed eye drops and standard solutions., Conclusions: The results of the study show the multicomponent hydrogel formulations of besifloxacin and levofloxacin to have superior corneal permeation with the potential for being used as topical ophthalmic preparations.

Purpose: The objective of this study was to describe the microbiological and clinical features of nine cases of Exserohilum keratitis., Patients and Methods: Fungal isolates from corneal scrapings were identified based on macroscopic and microscopic characteristics of the colonies and DNA sequencing of ITS1-5.8S-ITS2 region in the rRNA gene. All patients were treated with topical and if required systemic antifungals. Therapeutic penetrating keratoplasty (TPK) was done in case of failed medical therapy., Results: Morphologically, all fungal isolates were Exserohilum rostratum except one Exserohilum mcginnisii. Based on the BLAST analysis, 6 isolates showed 100% similarity to Setosphaeria rostrata (CBS 112815) and E. mcginnisii (CBS 20308). The other three isolates were Setosphaeria holmii (CBS 128053), not reported earlier in fungal keratitis. Three patients were lost to follow-up and response to medical therapy was good (Healed scar - 4 patients). Two out of nine patients were advised TPK., Conclusions: Diagnosis and clinical features of Exserohilum keratitis are akin to other dematiaceous keratitis. The two morphological species of E. mcginnisii and E. rostratum are indistinguishable from Setosphaeria rostratum at DNA sequence level, which justifies the retention of the latter nomenclature., Competing Interests: None

Purpose: To detect co-infections in the culture-proven acanthamoebic keratitis (AK) cases, and to test the capability of biofilm formation in the isolated microbiota. The clinical findings, habit of wearing contact lens and in-vitro antibiotic resistance were analyzed further according to the biofilm formation capability., Methods: After clinical examination, corneal scraps and swabs were taken from 240 clinically suspected AK cases, for Acanthamoeba and microbiological cultures. In cases of keratoplasty, trimmed corneal tissue was collected and sent for histopathological examination. Scanning electron microscopy was done for some samples. Biofilm formation capability was investigated using a tissue culture plate method. Antibiotic resistance pattern was determined using a modified-Kirby-Bauer disc diffusion method., Results: In 102 AK culture proven cases, 11 had no co-infection, 74 had a single co-infection and 17 had double co-infections. Enterobactericae and Aspergillus were the commonest bacterial and fungal isolates, respectively. Regarding the biofilm formation, 64.7% of Enterobactericae, 50% of Pseudomonas aeuroginosa, 43.75% of Staph aureus, 76.92% of Streptococcus pneumoniae, 28.57% of Corynebacterium, 60% of α-haemolytic streptococci, 40% of Acinetobacter, 100% of Candida and 77.8% Aspergillus isolates were biofilm producers. Severe manifestations were more frequently reported in cases co-infected with biofilm producers than with non-biofilm producers. Generally, high percentages of the biofilm forming bacterial isolates were sensitive to antibiotics in-vitro., Conclusions: Routine investigations for co-infection and biofilm formation in addition to Acanthamoeba culture are strongly recommended in suspected AK cases. Co-infection with biofilm producers may precipitate extrinsic in-vivo drug resistance despite of the in-vitro sensitivity. Designing a biofilm-dissolving topical drug is highly recommended to enhance the response to the standard therapeutic regimen especially in the resistant AK cases., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Purpose: Extemporaneously prepared miconazole eye drops (EPMD) are used by some eye care practitioners to manage keratomycosis in Ghana. This study therefore aimed to determine the efficacy and safety of EPMD using in vitro and in vivo experimental models., Methods: The minimum inhibitory concentration (MIC) of EPMD was determined by the agar-well diffusion method. In vivo, the activity of EPMD on corneal ulcer, neovascularization, clouding, edema, carring and on keratomycotic conjunctivitis and corneal scarring (clinical features) associated with Candida albicans-induced keratomycosis in rabbits was determined by treating them with 0.034-1.08% (weight-in-volume) EPMD for a period of 30 days. The safety of EPMD on the healthy eye was determined by instilling various concentrations into the intact eye of the rabbits., Results: The MIC of EPMD on Candida albicans was 1.08% (zone of inhibition of 13 mm ± 0.578), which resulted in significantly better improvements (p ≤ 0.001) in clinical findings than eyes treated with sterile water (p > 0.05), and showed no significant difference (p > 0.05) compared to eyes treated with 0.3% fluconazole. There were no visible signs of ocular toxicity on instilling it into healthy eyes of rabbits., Conclusion: The extemporaneously prepared miconazole eye drops are effective and safe to use in keratomycosis.

The effect of storage time and temperature on the recovery of pathogen DNA from polytetrafluorethylene (PTFE) was investigated. PTFE impression membranes were inoculated with Pseudomonas aeruginosa, Herpes Simplex Virus-1 (HSV-1) or Acanthamoeba and stored at -70 °C, -20 °C, +4 °C or +35 °C. PCR was performed on days 0, 1, 2, 3, 7 and months 1, 3 and 10 post-inoculation. We found no reduction in the DNA recovery of any of the studied microorganisms for the first 3 days of storage up to +35 °C. For HSV-1 and P. aeruginosa, storage for 3 months at +35 °C was associated with a significant reduction in DNA recovery (P<0.001), but not at +4 °C, -20 °C or -70 °C for 1 month for P. aeruginosa and for 10 months for HSV-1. Acanthamoeba DNA recovery was not affected by any storage parameters (P=0.203). These results will inform the investigation of microbial keratitis where access to microbiological testing is not readily available.

We describe a case of keratitis caused by Paecilomyces lilacinus in a contact lens wearer with a history of diabetes.

Purpose: To assess donor rim culture results and outcomes of ocular infections in the Cornea Preservation Time Study (CPTS)., Methods: Donor corneal rim cultures were optional. Donor characteristics were assessed for association with positive cultures using the Fisher exact test and Poisson regression analyses. Incidence rates of ocular infections were estimated, and 95% confidence intervals were calculated., Results: Cultures were performed in 784 (58.9%) of the 1330 cases. For the 0 to 7-day versus 8 to 14-day preservation time groups, respectively, positive fungal growth occurred in 10 of 397 (2.5%) versus 5 of 387 (1.3%) corneas (P = 0.30), whereas positive bacterial cultures occurred in 6 of 397 (1.5%) versus 4 of 387 (1.0%) corneas (P = 0.75). Surgeon-prepared tissue remained a significant risk for positive fungal cultures [relative risk (RR) of surgeon- versus eye-bank-prepared, 2.85; 95% CI (1.02-7.98)], whereas younger donors [RR per year of age, 0.96; 95% CI (0.93-1.00)] and accidental death donors [RR of accident versus disease, 3.71; 95% CI (1.36-10.13)] were at a greater risk for positive bacterial cultures. Fungal infection (Candida glabrata) developed in 1 (6.7%) of 15 recipients with a positive fungal culture, and no recipient infections occurred with positive bacterial culture. With one additional fungal keratitis (Candida albicans) and one bacterial endophthalmitis (E. coli) with no rim culture performed, a total of 2 of 1330 eyes (0.15%) developed fungal and 1/1330 eyes (0.08%) developed bacterial postkeratoplasty infections., Conclusions: A longer preservation time was not associated with a higher rate of positive donor rim cultures. The overall rate of infection across the entire cohort was low.

This study assessed the influence of donor, environmental, and logistical factors on the contamination rates of the conjunctival swabs and organ culture media of human donor eyes.In total, 1008 conjunctival swabs and 418 organ culture media samples from 504 consecutive human donor eyes were analyzed. Cross-tabulation, chi-squared tests, and Fisher's exact tests were used to assess the influence of the different factors on the contamination rates of the conjunctival swabs and organ culture media.The overall contamination rates were 28.4% for the conjunctival swabs and 1.0% for the organ culture media. A prolonged time between death and the conjunctival swab collection was associated with an increased conjunctival swab contamination rate [odds ratio (OR) = 1.9, 95% confidence interval (CI) = 1.2-3.0, P = .007]. The highest conjunctival swab contamination rate was found in the corneas procured in external institutions (outside the university hospital) (44.1%, OR = 3.6, 95%CI = 1.5-8.4, P = .003). Hospitalization times of 2 to 7 days prior to death were associated with an increased conjunctival swab contamination risk (OR = 2.6, 95%CI = 1.1-5.8, P = .021). However, the sex, age, cause of donor death, differentiation between septic and aseptic donors, differentiation between heart-beating brain-dead multiorgan donors and cadaveric donors, a warmer mean monthly temperature, and death to corneoscleral disc excision time did not significantly increase the conjunctival swab contamination risk. In addition, none of these factors affected the organ culture media contamination risk. Moreover, a positive conjunctival swab did not significantly increase the media contamination risk (P = .08). Surprisingly, the microorganisms causing media contamination were present at 50% of the amount detected on the conjunctival surface of the respective donor eye.A prolonged time between death and the conjunctival swab collection, a hospitalization time of 2 to 7 days prior to death, and corneal collection outside the university hospital seemed to be the main factors responsible for an increased conjunctival swab contamination risk. In addition, our investigation illustrated that a positive conjunctival swab is not a strong indicator for organ culture media contamination. Critical discussion is necessary regarding the validity of conjunctival swabs as prognostic parameters for organ culture media contamination.

Competing Interests: There are no conflicts of interest

Serratia liquefaciens is a rarely encountered gram-negative organism in ophthalmology practice. The only reported ocular infections are from contamination of contact lenses. The authors report the first case of a patient who developed orbital cellulitis secondary to severe S. liquefaciens microbial keratitis.

Objective: To present a case of diagnostic confusion in Nocardia keratitis in a contact lens wearer and to illustrate the characteristic clinical findings of this rare entity., Methods: Case report of Nocardia keratitis that was mistaken for acanthamoeba, herpetic, and fungal keratitis in three tertiary corneal referral centers before the correct diagnosis of Nocardia infection was made., Results: A 29-year-old contact lens wearer was referred to our hospital for a contact lens-associated bacterial keratitis, not improving under standard bacterial treatment. Biomicroscopy revealed a circular corneal ulcer and pinhead lesions arranged in a wreath pattern. Initial scraping revealed no positive cultures and confocal microscopy findings were suspicious for fungal keratitis. Only after a poor response to protozoal and fungal treatment, and a negative herpes serology, Nocardia was suspected. This suspicion was confirmed with a positive culture and the topical amikacin and ciprofloxacin were started. The infiltrate responded promptly and resolved, leaving a small corneal scar and a good visual recovery., Conclusion: Because of its infrequent occurrence and its variable clinical picture, Nocardia keratitis is easily misdiagnosed. Although a rare entity, this infection should be added to the differential diagnosis in contact lens-related keratitis not responding to first-line antibiotics and presenting with patchy anterior stromal infiltrates.

Purpose: To report novel surgical and medical management of intraocular and corneal infection with Candida glabrata that manifested 3 months after Descemet membrane endothelial keratoplasty (DMEK) and to review demographic reasons for increasing fungal contamination of donor tissue., Methods: Demographics, donor rim cultures, diagnostic tests, management, clinical course, outcomes, and donor mate outcomes are reported for a fungal infection after DMEK., Results: The fungal infection was treated for 3 weeks with a combination of intracameral and intracorneal voriconazole and intracameral and topical amphotericin B (off-label use). After initial improvement, the infection appeared to spread to the posterior chamber and the cornea decompensated. Pars plana vitrectomy was performed, and the fungal plaque and underlying area of the graft (approximately 3 mm area) were excised with the vitrector. After surgical intervention, the voriconazole and amphotericin B injections were discontinued, and oral posaconazole was prescribed for 2 months (off-label use). This combination of surgical and medical management successfully cleared the infection. Surprisingly, corneal edema completely resolved, and central DMEK endothelial cell density was 2506 cells/mm 1 month after discontinuing the antifungal injections, which apparently caused reversible endothelial toxicity., Conclusions: The incidence of nosocomial fungal infections, particularly non-albicans Candida, is increasing, as are the rates of positive fungal cultures from corneal donor tissue and postkeratoplasty fungal infections. Prospective studies are needed to assess the value of routine donor cultures, compare the safety and efficacy of various prophylactic treatments, and evaluate addition of antifungals to cold storage media.

Purpose: We compare results from regression discontinuity (RD) analysis to primary results of a randomized controlled trial (RCT) utilizing data from two contemporaneous RCTs for treatment of fungal corneal ulcers., Methods: Patients were enrolled in the Mycotic Ulcer Treatment Trials I and II (MUTT I & MUTT II) based on baseline visual acuity: patients with acuity ≤ 20/400 (logMAR 1.3) enrolled in MUTT I, and >20/400 in MUTT II. MUTT I investigated the effect of topical natamycin versus voriconazole on best spectacle-corrected visual acuity. MUTT II investigated the effect of topical voriconazole plus placebo versus topical voriconazole plus oral voriconazole. We compared the RD estimate (natamycin arm of MUTT I [N = 162] versus placebo arm of MUTT II [N = 54]) to the RCT estimate from MUTT I (topical natamycin [N = 162] versus topical voriconazole [N = 161])., Results: In the RD, patients receiving natamycin had mean improvement of 4-lines of visual acuity at 3 months (logMAR -0.39, 95% CI: -0.61, -0.17) compared to topical voriconazole plus placebo, and 2-lines in the RCT (logMAR -0.18, 95% CI: -0.30, -0.05) compared to topical voriconazole., Conclusions: The RD and RCT estimates were similar, although the RD design overestimated effects compared to the RCT.

Purpose: Here, we report risk factors associated with outcome in severe bacterial keratitis (BK), fungal keratitis (FK), and Acanthamoeba keratitis (AK) in India., Methods: Prospective observational cohort study conducted in Aravind Eye Hospital, India. Adults presenting with severe microbial keratitis (MK) were enrolled (size ≥3 mm) and followed to 21 days post-enrolment. Ulcer clinical features were recorded at presentation. Outcomes by final visit were classified as good (completely healed or reduced infiltrate size) or poor (enlarged infiltrate size, perforated, or surgery performed)., Results: Of 252 participants with severe MK, 191 had FK, 18 had AK, 19 had BK, 4 had mixed BK/FK, and 20 were microbiologically negative. Median age was 50 years (interquartile range [IQR]: 37-60 years), 64% were male, 63% were agriculturalists, and 45% had no formal education. Corneal trauma occurred in 72%, and median symptom duration before presentation was 7 days (IQR: 5-15 days). Clinical features associated with FK were feathery margins (p < 0.001), raised profile (p = 0.039), or dry surface (p = 0.007). Hypopyon was more likely in BK (p = 0.001) and ring infiltrate in AK (p < 0.001). Ulcers with poor outcome (n = 106/214) were more likely to be larger (odds ratio [OR]: 1.63, 95% confidence interval [CI]: 1.30-2.05, p < 0.001), involve the posterior cornea at presentation (OR: 2.31, 95% CI: 1.16-4.59, p = 0.017), involve Aspergillus sp. (OR: 3.23, 95% CI: 1.26-8.25, p = 0.014), or occur in females (OR: 2.04, 95% CI: 1.03-4.04, p = 0.04). Even after treatment, 34% (n = 76/221) had severe visual impairment by the final visit., Conclusions: Severe MK occurred predominantly in agriculturalists post-corneal trauma and often had poor outcomes. Provision of community-based eyecare may allow earlier treatment and improve outcomes.

Objective: To report the epidemiology of culture-positive bacterial corneal ulcers in Vancouver, B.C., Canada., Design: Retrospective, observational case series., Methods: Predetermined search terms were entered into the hospitals' electronic microbiology databases to create a cohort of patients who had undergone corneal scrapings for ulcers from April 2006 to March 2011. All specimens were plated on culture media. Cultured species were identified, and antimicrobial sensitivities were obtained. Clinical charts were then reviewed to identify associated risk factors., Results: In total 281 corneal scrapings were included, with a positive culture recovery rate of 75%. Bacterial keratitis accounted for 84.8% of culture-positive ulcers, followed by fungi (10%) and finally Acanthamoeba (5.2%); 73% of ulcers were monomicrobial in origin and 28% polymicrobial. We found an increase in Gram-negative micro-organisms over time. General sensitivity to antibiotics did not change over time. A major risk factor for Gram-positive involvement was ocular surface disease, whereas contact lens wear was a major risk factor for Gram-negative involvement., Conclusions: Bacterial keratitis was found to be the major cause of infectious keratitis in Vancouver, B.C. The majority of bacterial ulcers were caused by Gram-positive bacteria. However, we found an increase in Gram-negative involvement over time. Contact lens wear was identified as the major risk factor for development of Gram-negative ulcers. Pre-existing ocular disease was associated with Gram-positive infection. Susceptibility of Gram-negative bacteria to common broad-spectrum antibiotics was high, but susceptibility of Gram-positive bacteria to these antibiotics was lower and more variable., (Copyright © 2018 Canadian Ophthalmological Society. Published by Elsevier Inc. All rights reserved.)

Purpose: To report a difficult-to-identify case of keratitis due to Fusarium solani, diagnosed with the help of exhaustive gene analysis., Case: A 47-year-old woman attended our hospital with a refractory corneal ulcer associated with contact lens wear in her left eye that had appeared two weeks earlier. On her initial visit, slit lamp examination revealed a small double-ringed irregular hyphate ulcer in the center of the cornea, which had no epithelial defect, suggesting fungal infection. Microscopic visualization and culture of corneal scrapings were performed repeatedly, but the results were negative. Despite intensive antifungal treatment, infiltration of the cornea worsened and therapeutic keratoplasty was performed. Specimens from a corneal button were microbiologically and histologically negative for microorganisms. During irrigation of the anterior chamber with amphotericin B, aspirated corneal infiltrations were submitted for culture and metagenomic analysis. Genes belonging to F. solani were identified by metagenomic analysis, and an isolate presumed to be a Fusarium species was cultured. Genotypic identification of the isolates confirmed F. solani., Conclusion: The detection of microorganisms, especially fungi, can be extremely difficult. In difficult-to-culture cases, metagenomic analysis seems to be a promising technique for the identification of microbial pathogens., (Copyright © 2017 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Objective: To compare oral voriconazole vs placebo in addition to topical antifungals in the treatment of filamentous fungal keratitis., Design: Non-prespecified, secondary case-control analysis from a multicenter, double-masked, randomized placebo-controlled clinical trial., Methods: Study Participants: Patients with smear-positive filamentous fungal ulcers and visual acuity of 20/400 or worse who eventuated to therapeutic penetrating keratoplasty (TPK)., Intervention: Study participants were randomized to oral voriconazole vs oral placebo; all received topical antifungal drops., Main Outcome Measures: TPK button culture positivity., Results: A total of 95 of 194 (49.5%) study participants enrolled at Madurai, Coimbatore, or Pondicherry, India eventuated to TPK in an average of 20.9 days (standard deviation 15.2 days, range 2-71 days). TPK button cultures were available for 67 of 95 (71%) of the TPKs performed and were positive for filamentous fungus in 45 of 67 (67%) cases. For each 1-day increase in the time to TPK there was 0.94-fold decreased odds of fungal culture positivity (95% confidence interval [CI] 0.90-0.98, P = .005). Those randomized to oral voriconazole had 1.26-fold increased odds of TPK button culture positivity after controlling for time to TPK and baseline organism, but this was not statistically significant (95% CI 0.32-4.87; P = .74). Those who underwent TPK for lack of response to medical therapy were 10.64-fold more likely to be culture positive than if the indication for surgery was perforation and this was statistically significant (95% CI 2.16-51.70; P = .003)., Conclusions: There appears to be no benefit to adding oral voriconazole to topical antifungal agents in the treatment of severe filamentous fungal ulcers. Infection rather than inflammation appears to be the reason for the worsening clinical picture in many of these patients., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Purpose: To document the findings of corneal biopsies for progressive microbial keratitis in a large tertiary referral institution., Methods: A retrospective medical records review of all patients who underwent at least one corneal biopsy for the diagnosis of microbial keratitis at Sydney Eye Hospital, Australia between January 1, 2010 and December 31, 2016 was performed., Results: Thirty-eight patients (18 men and 20 women) underwent a corneal biopsy for progressive microbial keratitis unresponsive to broad-spectrum topical antimicrobials. Risk factors for microbial keratitis included contact lens wear in 8 (21%), recent intraocular surgery in 5 cases (13%), recent agricultural trauma in 3 cases (8%), exposure keratopathy due to Graves' orbitopathy in 1 case (3%), and profound systemic immunosuppression due to chemotherapy for leukaemia in 1 case (3%). The remaining 20 patients had no identifiable risk factors. Fifteen patients (39%) had a positive biopsy result, which identified bacteria in 6 cases and Mycobacteria in 1 case, both by culture of the biopsy specimen. Three cases of fungus were identified on culture of biopsy specimen, two of which were also confirmed on histopathology and an additional case was identified from histopathology alone. A single case of Acanthamoeba was diagnosed by culture and histopathology, and an additional 3 cases were diagnosed on histopathology alone. A corneal biopsy yielded new organisms in 73% (11/15) cases where the culture results of biopsy specimens were positive., Conclusion: Corneal biopsy is an important tool in the diagnosis of progressive keratitis, often identifying causal organisms not found on corneal scraping alone.

Purpose: To identify mycotoxin genes among clinical ocular isolates of Fusarium species and to correlate these with clinical outcomes of Fusarium keratitis., Methods: Fifty-four clinical isolates of Fusarium were retrieved from the Bascom Palmer Eye Institute Ocular Microbiology Laboratory data bank. Multiplex polymerase chain reactions were run to confirm the identification of Fusarium species [internal transcribed spacer sequence, translation elongation factor 1-α (TEF) and β-tubulin] and to detect the presence of genes encoding production of fumonisin B mycotoxins (FUM1 and FUM8) and trichothecene mycotoxins (deoxynivalenol and nivalenol). The presence or absence of mycotoxins was compared with patient outcomes., Results: Forty-three (79%) of the 54 isolates were confirmed as Fusarium species, by an internal transcribed spacer sequence in 3 (5.6%) and by TEF in 43 (79.6%) of the 54 isolates. Fumonisin biosynthetic gene 1 (FUM1) was detected in 57.4% (n = 31/54) of the Fusarium isolates. No FUM8, deoxynivalenol genes, and nivalenol genes were detected among these in the clinical isolates group. Initial best-corrected visual acuity ranged from 20/25 to 20/80 in the FUM1 gene-negative group and from 20/20 to light perception in the FUM1 gene-positive group. There was no difference in the time to cure between both groups. The presence of FUM1 genes in 5 fungal isolates seemed to be associated with progression to penetrating keratoplasty in the 5 patients from whom the fungi were isolated. Fusarium solani was recovered from all patients requiring penetrating keratoplasty., Conclusions: Fumonisin B biosynthetic gene 1 may be common among clinical Fusarium isolates and contribute to worse initial visual acuity and high-risk progression to penetrating keratoplasty.

Microsporidia are protists close to the kingdom of fungi that may cause eye infections. Most cases are reported in Asia and affect both immunocompromised and immunocompetent patients. Here, we report a rare case of microsporidial keratoconjunctivitis in an immunocompetent French patient 3 weeks after returning from India. In our patient, Weber trichrome staining of conjunctival scrapings revealed rounded elements approximately 1-3 μm in size. Conventional polymerase chain reaction analysis by ribosomal RNA subunit sequencing showed 100% identity with Vittaforma corneae . Treatment by corneal debridement combined with fluoroquinolone eye drops allowed complete resolution of the lesions. Although rare, ocular microsporidiosis should be investigated in a patient who is native to Asia or has returned from an endemic area and presents with keratoconjunctivitis of undetermined etiology.

Purpose: The purpose of this study is to determine predisposing factors, common bacterial causes, and antibiotic sensitivity of corneal ulcers in Murtala Muhammad Specialist Hospital., Materials and Methods: A prospective, cross-sectional, observational study was carried out on patients with corneal ulcers. Information on relevant clinical, sociodemographic profile was obtained. Examination includes visual acuity test and slit-lamp biomicroscopy, especially fluorescein staining. Corneal scrapings from the ulcer were inoculated onto the selected solid culture media plates. Gram staining for microscopic examination was done. Antibiotic sensitivity test on different antibiograms was done using modified Kirby-Bauer technique, determining bacterial isolate sensitivity or resistance to relevant antibiotics (chloramphenicol, ciprofloxacin, ofloxacin, gentamicin, tetracycline, ceftazidime, ceftriaxone, cefotaxime, and penicillin)., Results: A total of 77 patients with corneal ulcer were examined. Ocular trauma, application of harmful traditional eye medication, and use of unspecified topical medication before presentation were among the predisposing factors. Bacterial growth was seen in 46.8% of the samples, of which 28.6% of the growth were Gram-positive. The common bacteria isolated were Staphylococcus epidermidis, Staphylococcus aureus, Pseudomonas aeruginosa, Streptococcus pneumoniae, Proteus species, and Klebsiella pneumoniae. Isolates were sensitive to ciprofloxacin and ofloxacin while an intermediate sensitivity was seen with chloramphenicol and gentamicin. Isolates were resistant to tetracycline, ceftazidime, ceftriaxone, cefotaxime, and penicillin., Conclusion: Ocular trauma was the major predisposing factor to corneal ulcer, and Staphylococcus species was the major bacterial organism isolated. Isolates were found to be sensitive to ciprofloxacin and ofloxacin, while an intermediate sensitivity was seen with chloramphenicol and gentamicin., Competing Interests: There are no conflicts of interest

Purpose: To report a case of keratitis caused by Providencia alcalifaciens., Methods: A 70-year-old man presented with pain, redness, and decreased vision in both of his eyes. On initial presentation, the visual acuity in both of his eyes was hand motion, and slit lamp examination showed diffuse infiltrates and total epithelial defects of the cornea, conjunctival hyperemia, and severe anterior chamber reactions with 2-mm-height hypopyon. The results of laboratory studies revealed elevated liver function tests, which indicated the presence of alcoholic liver disease., Results: The culture result was positive for P. alcalifaciens and showed sensitivity to ceftazidime and amikacin but resistant to cefazolin, gentamicin, fluoroquinolone, and ampicillin. He was treated with topical application and intravenous injection of ceftazidime. His keratitis resolved gradually over 3 months as the treatment dose was tapered and the treatment was eventually discontinued. His final visual acuity was 20/200 in the right eye and 20/60 in the left eye because of the remaining central corneal opacity., Conclusions: We report a first case of keratitis due to P. alcalifaciens in a chronic heavy alcohol drinker. Ophthalmologists should consider this bacterium as a potential pathogen of corneal ulceration, when clinically suspect gram-negative bacterial keratitis especially in immunocompromised state.

Objective: (i) To assess the rate of positive microbiological cultures of corneas prepared by the Eye Bank of Canada (Ontario Division) between January 1, 2012, and December 31, 2013; (ii) to review the microbiology protocols at the 5 major transplant centres in Ontario; and (iii) to assess the incidence of endophthalmitis during the study period., Design: Retrospective chart review., Participants: A total of 4186 consecutive cultured corneal tissues prepared by the Eye Bank from January 1, 2012, to December 31, 2013., Methods: Rates of culture-positive cornea rims and incidence of postkeratoplasty endophthalmitis at 5 surgical centres in Ontario were determined, and the protocols used to culture rims at each site were concurrently reviewed. Culture results were analyzed via logistic regression for positive cultures., Results: The rate of positive cultures at each sites were as follows: centre A, 3.74%; centre B, 3.26%; centre C, 0.51%; centre D, 0.48%; and centre E, 0.04%. Centres A, B, and D were noted to have significantly higher positive rates than centre E. In comparing microbiology protocols, longer incubation period (11 days) was 12 times more likely to be associated with higher positive culture rates than shorter period (4-5 days). Six-month follow-up of all keratoplasties revealed zero reported cases of endophthalmitis., Conclusions: A literature review regarding the predictive value of routine culturing reveals conflicting data. Our findings suggest that differences in the microbiology protocols directly influence the rates of positive rim cultures. Without a standardized protocol, it is not possible to evaluate the predictive value of routine corneal rim culturing in predicting postkeratoplasty endophthalmitis., (Copyright © 2018. Published by Elsevier Inc.)

Penetration of antibiotics into and through the cornea is a major limiting factor in the treatment of ocular infections. Several strategies are in vogue to overcome this limitation such as use of fortified drops, gels, and subconjunctival injections. Here, we present the fabrication of rapidly dissolving polymeric microneedle array to effectively deliver besifloxacin through the cornea. Microneedles were prepared using polyvinyl alcohol and polyvinyl pyrrolidone by the micromolding technique. The model fluoroquinolone antibiotic, besifloxacin, was loaded in 36 microneedles arranged in a 6 × 6 array format within a 1 cm 2 area. The average height and base width of microneedles was 961 ± 27 and 366 ± 16 μm, respectively. Each microneedle array contained 103.4 ± 8.5 μg of besifloxacin. Cryosectioning and confocal microscopy of excised human cornea revealed that microneedles penetrated to a depth of up to 200 μm. Microneedles were found to completely dissolve in the cornea within 5 min. Application of microneedles for 5 min significantly (p < 0.05) improved the besifloxacin deposition and permeation through the cornea compared with free besifloxacin solution. Similarly, besifloxacin-loaded microneedles showed greater antibacterial activity in Staphylococcus aureus-infected cornea in comparison to free besifloxacin solution. Taken together, rapidly dissolving microneedles can be developed to effectively deliver besifloxacin to treat bacterial infections in the cornea and eye.

Purpose: To report a series of cases of Acanthamoeba keratitis (AK) in scleral lens wearers with keratoconus to determine whether this type of contact lens presents a greater risk for development of infection., Methods: This study reports three patients who wore scleral contact lenses to correct keratoconus and developed AK. The diagnoses of AK were established based on cultures of the cornea, scleral contact lenses, and contact lens paraphernalia. This study investigated the risk factors for infections., Results: The possible risks for AK in scleral contact lens wearers are hypoxic changes in the corneal epithelium because of the large diameter and minimal tear exchange, use of large amounts of saline solution necessary for scleral lens fitting, storing the scleral lens overnight in saline solution rather than contact lens multipurpose solutions, not rubbing the contact lens during cleaning, and the space between the cornea and the back surface of the scleral lens that might serve as a fluid reservoir and environment for Acanthamoeba multiplication. Two patients responded well to medical treatment of AK; one is still being treated., Conclusions: The recommendations for use and care of scleral contact lenses should be emphasized, especially regarding use of sterile saline (preferably single use), attention to rubbing the lens during cleaning, cleaning of the plunger, and overnight storage in fresh contact lens multipurpose solutions without topping off the lens solution in the case., (Copyright © 2017 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.)

The role of mast cells (MCs) in fungal infection is largely unknown. This study was to explore a protective role and mechanism of MCs in fungal keratitis. Experimental fungal keratitis (FK) mouse model was developed. Mice untreated (UT) or receiving corneal wound without fungal infection (Mock) were used as controls. Large number of connective tissue MCs was found in normal mice. MC activation with degranulation was largely observed, and the percentage of degranulated/total cells was high in FK. Dilated limbal vasculature with increased permeability, as well as largely infiltrated neutrophils with stimulated ICAM-1 protein levels were observed in corneas of FK mice, when compared with Mock and UT mice. Interestingly, pretreatment with cromolyn sodium (Block) significantly blocked MC degranulation, dramatically suppressed vascular dilation and permeability, and markedly reduced neutrophil infiltration with lower ICAM-1 levels in FK mice at 6-24 hours. Furthermore, the Block mice manifested prolonged disease course, increased pathological damage, and vigorous fungus growth, with much higher corneal perforation rate than FK mice at 72 h. These findings reveal a novel phenomenon that MCs play a vital role in protecting cornea against fungal infection through degranulation that promotes neutrophil infiltration via stimulating ICAM-1 production and limbal vascular dilation and permeability.

We compared the clinical characteristics, treatments, and prognoses of fungal keratitis in patients with and without diabetes. Patients diagnosed with fungal keratitis at Shandong Eye Institute between January 2010 and December 2016 were retrospectively reviewed and classified as diabetic and nondiabetic groups. One-hundred-and-eleven patients (111 eyes) with diabetes and 740 patients (740 eyes) without diabetes were included. The diabetic patients showed significantly older (p< 0.05) and lower male:female ratio (p<0.05). Plants trauma was the primary risk factor in both groups, and there was no significant difference of pathogen type (the most common was Fusarium genus, followed by Alternaria and Aspergillus genera). Multivariate logistic regression analyses revealed that diabetes and topical glucocorticoid use were the independent risk factors for the severity of fungal keratitis. The recurrent infection rate between the diabetic and nondiabetic patients during the follow-up (6 to 24 months) after penetrating keratoplasty (PKP) was not significantly different. Although the recurrent epithelial defect, rejection, and best-corrected visual acuity were similar between the patients with matched bed/graft size (7.75/8.0 mm) in the two groups 1 year after PKP, the incidence of delayed re-epithelialization (>7 days) was significantly higher in diabetic patients (3/10 versus 2/43 in nondiabetic patients, p<0.05). More specially, the diabetic patients with the duration ≥10 years showed more significantly delayed re-epithelialization than those with the diabetic duration less than 10 years (3/5 versus 1/26, p<0.05). In conclusion, the diabetes mellitus is an independent risk factor that affect the severity of fungal keratitis. Corneal re-epithelialization was significantly delayed after PKP in the diabetic patients, especially with the duration ≥10 years.

Econazole is a feasible alternative treatment in the management of fungal keratitis. Nevertheless, its low water solubility is considered the main limitation to the incorporation into ophthalmic formulations. In this work, econazole nitrate is solubilized by using cyclodextrins to achieve an optimum therapeutic concentration. Phase solubility diagrams suggest α-cyclodextrin as the most effective cyclodextrin and later the inclusion complex formed with this one was characterized in solution by 1D, 2D-NMR, and molecular modeling. Econazole-α-cyclodextrin inclusion complex was included in 2 types of ocular hydrogels: a natural polysaccharides ion-sensitive hydrogel and a hyaluronic acid hydrogel. Both of them show no ocular irritation in the hen's egg test on chorioallantoic membrane assay and a controlled econazole release over time. Permeability studies suggest that hydrogels do not modify the econazole nitrate permeability through bovine cornea in comparison with an econazole-α-cyclodextrin inclusion complex solution. Finally, ocular biopermanence studies performed using positron emission tomography show these hydrogels present a high retention time on the eye. Results suggest the developed formulations have a high potential as vehicles for the econazole topical ocular administration as fungal keratitis treatment., (Copyright © 2018 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.)

Objective: To evaluate the incidence of corneal abrasions/ulceration and microbial contamination in horses undergoing general anaesthesia., Study Design: Prospective, observational, clinical study., Animals: A total of 40 client-owned healthy horses scheduled for elective non-ophthalmic procedures., Methods: Conjunctival sac swabs were taken, fluorescein dye applied and digital images recorded from both eyes of the horses after preanaesthetic medication and 24 hours after recovery from general anaesthesia. A paraffin-based bland ophthalmic ointment was applied on the ocular surface intraoperatively following collection of a sample into a sterile container. All samples underwent aerobic, anaerobic and fungal culture. Subject demographics, chronology of ophthalmic ointment use, anaesthesia duration, recumbency after induction, during surgery and recovery, fluorescein uptake and culture results were recorded. Descriptive statistics were performed., Results: Complete data were collected from 34 horses; six (17.6%) developed mild unilateral generalized fluorescein uptake consistent with corneal abrasions. Recumbency on the operating table was the only risk factor significantly associated with corneal abrasions. A total of 11 bacterial species were identified; Staphylococcus spp. (15 eyes) and Micrococcus spp. (eight eyes) were the most frequently isolated bacteria. Two fungal species were isolated postoperatively (Aspergillus spp., Saccharomyces spp.) in two eyes. Ointment contamination was recorded in two cases (5%) but cross-contamination was not recognized., Conclusionsand Clinical Relevance: Incidence of corneal abrasion/ulceration in horses undergoing general anaesthesia and contamination rate of ophthalmic solutions are similar to those previously reported in dogs., (Copyright © 2018 Association of Veterinary Anaesthetists and American College of Veterinary Anesthesia and Analgesia. Published by Elsevier Ltd. All rights reserved.)

Acanthamoeba keratitis (AK) is a severe corneal infection that may occur as a serious outcome of improper use of contact lenses (CL)., Objectives: The study aimed to diagnose AK in soft CL users presenting with infectious keratitis, and to identify the prevalent genotypes isolated from different cases. Another aim was to determine the CL hygiene-related risk behaviors, and to explore the risk of water exposure for developing AK., Methods: A cross sectional study was performed. A questionnaire was carried out including 260 clinically diagnosed cases as infectious keratitis (170 females and 90 males); all of them were soft CL users for the suspected risk factors. Corneal scrapes from the affected eyes were cultured to diagnose bacterial and AK. PCR was performed and the amplified products were sequenced and compared with GenBank data., Results: The parasite was positively amplified from 32 samples (12.3%). Acanthamoeba T4 genotype was identified in 27/32 (84.4%) of isolates. Other detected genotypes belonged to T5 and T3 genotypes at rates of 9.4%, and 6.25%, respectively. The most important risk factors associated with development of AK were female sex, sleeping while wearing CL, and exposure to water resources through different practices. These practices included rinsing the CL case in tap water, swimming and/or showering while wearing CL, using multipurpose solution for cleaning the lenses, using water from over-building tanks. Rubbing the eyes due to discomfort when applying CL was an additional important risk factor associated with AK. The protective factor was regular hand washing before using CL., Conclusion: CL users were more exposed to AK and should gain enough health education regarding proper lens hygiene and dangers of tap water exposure., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Purpose: The aim of this paper is to present the case report of a patient developing endophthalmitis after penetrating keratoplasty caused by a multidrug-resistant Pseudomonas aeruginosa, detected only in the contralateral donor tissue., Case Report: A 77-year-old man underwent an uneventful penetrating keratoplasty with a preoperative culture-negative donor cornea; however, the fellow cornea grew multidrug-resistant Pseudomonas aeruginosa. The patient developed and was treated for endophthalmitis after penetrating keratoplasty, and aqueous and vitreous taps grew P. aeruginosa with antibiotic resistance identical to the isolate from the mate cornea. Sequence analysis of the 16S ribosomal gene from the two isolates and confirmation analyzing the sequence of P. aeruginosa heat shock protein gene (groES) were performed showing the same strain for both organisms., Conclusion: This case report documents the presence of the same multidrug-resistant P. aeruginosa causing endophthalmitis after penetrating keratoplasty and in the contralateral donor tissue, suggesting that we must be cautious in deciding to transplant tissues with positive culture in the contralateral donor cornea., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Competing Interests: Die Autoren geben an, dass kein Interessenkonflikt besteht.

Purpose: Acanthamoeba keratitis (AK) is an uncommon but serious corneal infection, in which delayed diagnosis carries a poor prognosis. Conventional culture requires a long incubation period and has low sensitivity. Polymerase chain reaction (PCR) and in vivo confocal microscopy (IVCM) are available alternative diagnostic modalities that have increasing clinical utility. This study compares confocal microscopy, PCR, and corneal scrape culture in the early diagnosis of AK., Methods: We reviewed the case notes of patients with a differential diagnosis of AK between June 2016 and February 2017 at the Bristol Eye Hospital, United Kingdom. Clinical features at presentation, and results of IVCM, PCR, and corneal scrape cultures were analyzed., Results: A total of 25 case records were reviewed. AK was diagnosed in 14 patients (15 eyes). Based on the definition of "definite AK," the diagnostic sensitivities of IVCM, PCR, and corneal scrape cultures were 100% [95% confidence interval (CI), 63.1%-100%], 71.4% (95% CI, 41.9%-91.6%) and 33.3% (95% CI, 9.9%-65.1%), respectively. The 3 methods showed a specificity of 100% and a positive predictive value of 100%. Using a reference standard of only positive corneal cultures, IVCM, and PCR had a sensitivity of 100% (95% CI, 29.2%-100%) and 75% (95% CI, 19.4%-99.4%), respectively., Conclusions: All 3 diagnostic tests are highly specific, and a positive test result is highly predictive of disease presence. IVCM is both highly sensitive and specific when performed by an experienced operator. PCR is a useful adjunct in the diagnosis of AK because of its wider availability compared with IVCM, and it may be used in combination with IVCM for microbiologic confirmation.

Purpose: To investigate the expression and roles of type I and II interferons (IFNs) in fungal keratitis, as well as the therapeutic effects of tacrolimus (FK506) and voriconazole on this condition., Methods: The mRNA and protein expression levels of type I (IFN-α/β) and II (IFN-γ) IFNs, as well as of related downstream inflammatory cytokines (interleukin (IL)-1α, IL-6, IL-12, and IL-17), were detected in macrophages, neutrophils, lymphocytes, and corneal epithelial cells (A6(1) cells) stimulated with zymosan (10 mg/ml) for 8 or 24 h. A fungal keratitis mouse model was generated through intrastromal injection of Aspergillus fumigatus , and the mice were then divided into four groups: group I, the PBS group; group II, the voriconazole group; group III, the FK506 group; and group IV, the voriconazole plus 0.05% FK506 group. Corneal damage was evaluated with clinical scoring and histological examination. In addition, the mRNA and protein expression levels of type I (IFN-α/β) and type II (IFN-γ) IFNs, as well as related inflammatory cytokines, were determined at different time points using quantitative real-time PCR (qRT-PCR) and western blotting., Results: After zymosan stimulation of mouse neutrophils, lymphocytes, macrophages, and A6(1) cells, the IFN mRNA and protein expression levels were markedly increased until 24 h, peaking at 8 h (p<0.001). The mRNA and protein expression levels of inflammatory cytokines (IL-1α, IL-6, IL-12, and IL-17) were also upregulated after zymosan stimulation. Moreover, type I (IFN-α/β) and type II (IFN-γ) IFN expression levels were increased and positively correlated with the progression of fungal keratitis in vivo. FK506 administered with voriconazole reduced the pathological infiltration of inflammatory cells into the cornea and downregulated the expression levels of IFNs and related inflammatory cytokines., Conclusions: In conclusion, this study demonstrated that type I and II IFN levels were markedly increased in fungal keratitis and that FK506 combined with voriconazole decreased the severity of fungal keratitis by suppressing type I and II IFNs and their related inflammatory responses.

Combined antifungal and antioxidant therapy may help to reduce oxidative stress in fungal keratitis. Experimental Fusarium solani keratitis was induced by application of F. solani conidia to scarified cornea (right eye) of 16 rabbits (another four rabbits were negative controls [Group I]). Five days later, F. solani-infected animals began receiving hourly topical saline alone (Group II), voriconazole (10 mg/mL) alone (Group III), epigallocatechin gallate (EGCG, 10 mg/mL) alone (Group IV) or voriconazole and EGCG (Group V). Twenty days post-inoculation, corneal lesions were graded. After animal sacrifice, excised corneas underwent histopathological and microbiological investigations. Corneal tissue levels/activities of interleukin 1 beta (IL-1β) and tumour necrosis factor alpha (TNF-α) gene mRNA transcripts, matrix metalloproteinase (MMP) 2 and 9 proteins, malondialdehyde (MDA) and reduced glutathione (GSH), and superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx), were also measured. Clinical and histopathological scores (severity of corneal lesions; [P < .05]) and mean levels (P < .05) of IL-1β and TNF-α mRNA transcripts, MMP 2, MMP 9 and MDA were Group II > Groups IV and III > Groups V and I. Mean SOD, CAT, GPx and GSH levels (P < .05) were Group II < Groups IV and III < Groups V and I. Topical voriconazole with EGCG apparently reduces inflammation in experimental F. solani keratitis, as manifested by improved clinical, histological, microbiological and molecular parameters., (© 2017 Blackwell Verlag GmbH.)