Biophilic Positive Carbon Dot Exerts Antifungal Activity and Augments Corneal Permeation for Fungal Keratitis.

Fungal keratitis (FK) is an infectious eye disease that poses a significant risk of blindness. However, the effectiveness of conventional antifungal drugs is limited due to the intrinsic ocular barrier that impedes drug absorption. There is an urgent need to develop new therapeutic strategies to effectively combat FK. Herein, we synthesized an ultrasmall positively charged carbon dot using a simple stage-melting method. The carbon dot can penetrate the corneal barrier by opening the tight junctions, allowing them to reach the lesion site and effectively kill the fungi. The results both in vitro and in vivo demonstrated that it exhibited good biocompatibility and antifungal activity, significantly improving the therapeutic effect in a mouse model of FK. Therefore, this biophilic ultrasmall size and positive carbon dot, characterized by its ability to penetrate the corneal barrier and its antifungal properties, may offer valuable insights into the design of effective ocular nanomedicines.

Mesenchymal stem cell-based adjunctive therapy for Pseudomonas aeruginosa-induced keratitis: A proof-of-concept in-vitro study.

PURPOSE: Pseudomonas aeruginosa-induced keratitis is one of the most severe and challenging forms of corneal infection, owing to its associated intense inflammatory reactions leading to corneal necrosis and dense corneal scar with loss of vision. Since mesenchymal stem cells (MSCs) are reported to possess antimicrobial and immunomodulatory properties, they can be tested as an adjuvant treatment along with the antibiotics which are the current standard of care. This study aims to investigate the anti-bacterial and immunomodulatory roles of human bone marrow MSC-derived conditioned medium (MSC-CM) in P. aeruginosa-infected human corneal epithelial cells (HCECs) in vitro. METHODS: The effect of MSC-CM on the growth of clinical isolates of P. aeruginosa was evaluated by colony-forming unit assay. The expression of inflammatory cytokines (IL-6 and TNF-α) and an antimicrobial peptide (Lipocalin 2) in lipopolysaccharide-treated MSCs and HCECs was analyzed through ELISA. Corneal epithelial repair following infection with P. aeruginosa was studied through scratch assay. RESULTS: Compared to control (P. aeruginosa (5*105) incubated in DMEM (1 ml) at 37 °C for 16 h), MSC-CM significantly: i) inhibits the growth of P. aeruginosa (159*109 vs. 104*109 CFU/ml), ii) accelerates corneal epithelial repair following infection with P. aeruginosa (9% vs. 24% closure of the wounded area after 12 h of infection), and iii) downregulates the lipopolysaccharide-induced expression of IL-6, TNF-α and Lipocalin 2 in HCECs. A combination of MSC-CM with an antibiotic, Ciprofloxacin moderately regulated the expression of IL-6, TNF-α, and Lipocalin 2. CONCLUSION: MSC-CM holds promise as an adjunctive therapeutic approach for P. aeruginosa-induced corneal epithelial damage.

Infectious keratitis following corneal transplantation: A long-term cohort study.

BACKGROUND: To assess the long-term incidence and risk factors for post-keratoplasty infectious keratitis (IK), associated ocular pathogens, and antibiotic resistance profiles. METHODS: Cohort study including 2553 consecutive penetrating, endothelial, and anterior lamellar keratoplasties performed between 1992 and 2020. Medical and microbiological records of patients clinically diagnosed with IK were retrospectively reviewed. MAIN OUTCOME MEASURES: cumulative incidence of IK, infectious agent species, and antibiotics resistance profiles. RESULTS: The average follow-up time after transplantation was 112 ± 96 months. Eighty-nine IK episodes were recorded; microbiological tests were positive in 55/89 (62%). The cumulated incidence of postoperative IK was 5.50%/10.25% at 10/20 years. The occurrence of at least one episode of IK after transplantation was associated with lower graft survival in the long term (p < 0.0001). Rejection risk (adjusted Hazard Ratio, 2.29) and postoperative epithelial complications (HR, 3.44) were significantly and independently associated with a higher incidence of postoperative IK. Infectious agents included 41 bacteria, 10 HSV, 6 fungi, and 1 Acanthamoeba. The rate of antibiotic resistance was 0% for vancomycin, 13% for fluoroquinolones, 20% for rifamycin, 59% for aminoglycosides, and 73% for ticarcillin. In 41% of cases, patients were under prophylactic topical antibiotics before the infectious episode. Topical antibiotics were significantly associated with increased resistance to penicillin, carbapenems, and aminoglycosides. CONCLUSION: IK (mainly bacterial) is a frequent complication of corneal transplantation in the long term. Vancomycin and fluoroquinolones can be considered as first-line treatments. Prolonged postoperative antibiotic preventive treatment is not advisable as it may increase antibiotic resistance.

In Vivo Confocal Microscopy Characterization of Candida parapsilosis Keratitis.

ABSTRACT: The present clinical case concerns two patients with mycotic keratitis because of Candida parapsilosis in which corneal confocal microscopy presented a characteristic feature of this pathogen. Both described patients used a therapeutic contact lens and administered a therapy with steroid eye drops which are well known predisposing factors for the onset of corneal mycoses. This report can be useful for correctly identifying the pathologic condition and quickly directing the therapy.

Corneal stromal ulcerations in a referral population of dogs and cats in the Netherlands (2012-2019): Bacterial isolates and antibiotic resistance.

OBJECTIVE: To evaluate bacterial isolates from corneal stromal ulcerations in dogs and cats in the Netherlands, review their antibiotic susceptibility, determine whether recent topical treatment affected bacterial culture results, and investigate whether (multi-drug) resistance patterns changed over time. ANIMALS STUDIED: Client-owned dogs and cats were diagnosed with corneal stromal ulceration at the Utrecht University Clinic for Companion Animals between 2012 and 2019. PROCEDURES: Retrospective analysis. RESULTS: In total, 163 samples were collected from 122 dogs (130 samples) and 33 cats. Positive cultures were obtained from 76 canine and 13 feline samples (59% and 39%, respectively) and included Staphylococcus (42 in dogs, 8 in cats), Streptococcus (22 in dogs, 2 in cats), and Pseudomonas (9 in dogs, 1 in cats) species. Significantly fewer positive cultures were found in dogs and cats previously treated with topical antibiotics (χ2  = 6.52, p = .011 and χ2  = 4.27, p = .039, respectively). Bacterial resistance to chloramphenicol was more common in dogs previously treated with chloramphenicol (χ2  = 5.24, p = .022). The incidence of acquired antibiotic resistance did not increase significantly over time. In dogs, the incidence of multi-drug-resistant isolates increased significantly between 2012-2015 and 2016-2019 (9.4% vs. 38.6%, p = .0032). CONCLUSIONS: Staphylococcus, Streptococcus, and Pseudomonas species were the most common bacteria associated with canine and feline corneal stromal ulcerations. Previous treatment with antibiotics affected bacterial culture results and antibiotic sensitivity. Although the overall incidence of acquired antibiotic resistance did not change over time, the incidence of multi-drug-resistant isolates in dogs increased over an 8-year period.

Microbial keratitis and its management at a rural centre: achieving success with limited resources.

PURPOSE: Microbial keratitis is a sight-threatening condition with a higher incidence in agrarian populations. In countries with a high indigent population, due to financial and other constraints, patients prefer to seek therapy locally rather than travel to advanced centres. The aim of this study is to describe the epidemiology, clinical characteristics, and outcomes of 60 consecutive patients with microbial keratitis managed at a rural centre. METHODS: Descriptive case series. All patients clinically diagnosed with infectious keratitis were included. Corneal scrapings were obtained and microbiological identification was done by Gram stain. Anti-microbial therapy was commenced based on smear findings and the patients were followed up till disease resolution. RESULTS: Sixty eyes of 60 patients were diagnosed with microbial keratitis in the study period. The mean age was 47.43 ± 18.69 years. Male:female ratio was 47:53. Risk factors included ocular trauma in the majority of patients (46/60; 76.7%). Microorganisms were identified on 75.6% of smears, with fungal filaments (65.4%) being the most common. Ulcers were central in over half (32/60; 53.3%), and > 3 mm in diameter in over three-fourths (81.6%) of patients. Forty-four patients (73.3%) achieved treatment success whereas 16/60 (26.6%) required referral to our tertiary-eye care facility for management. The median time to resolution was 14 days (IQR 10-26 days). CONCLUSION: Our series demonstrates the feasibility of microbiology-guided therapy in microbial keratitis by ophthalmologists at the secondary rural eye-care level. Two-thirds of the patients could be successfully managed at the rural centre and only severe cases needed a referral to tertiary centres.

Investigating and Treating a Corneal Ulcer Due to Extensively Drug-Resistant Pseudomonas aeruginosa.

Resistant Gram-negative bacteria are a growing concern in the United States, leading to significant morbidity and mortality. We identified a 72-year-old female patient who presented with unilateral vision loss. She was found to have a large corneal ulcer with hypopyon. Culture of corneal scrapings grew extensively drug-resistant Pseudomonas aeruginosa. Treatment involved a combination of systemic and topical antibiotics. Whole genome sequencing revealed the presence of blaVIM-80, blaGES-9, and other resistance determinants. This distinctive organism was linked to an over-the-counter artificial tears product.

Innovative cold atmospheric plasma (iCAP) decreases corneal ulcer formation and bacterial loads and improves anterior chamber health in methicillin resistant Staphylococcus aureus keratitis.

Bacterial keratitis is a vision-threatening infection of the cornea that is typically treated with antibiotics. However, antibiotics sometimes fail to eradicate the infection and do not prevent or repair the damage caused directly by the bacteria or the host immune response to the infection. Our group previously demonstrated that treatment of Pseudomonas aeruginosa keratitis in rabbits with innovative cold atmospheric plasma (iCAP) resulted in reduced edema, ulcer formation, and bacterial load. In this study, we investigated the efficacy of iCAP treatment in methicillin-resistant Staphylococcus aureus (MRSA). New Zealand white rabbits were infected intrastromally with MRSA then treated with iCAP, moxifloxacin, vancomycin, or combination of iCAP with each antibiotic to assess the safety and efficacy of iCAP treatment compared to untreated controls and antibiotics. iCAP treatment significantly reduced bacterial loads and inflammation, improved anterior chamber clarity, and prevented corneal ulceration compared to untreated controls and antibiotic treatment. Safety assessments of grimace test scores and tear production showed that iCAP was not significantly different from either antibiotic treatment in terms of distress or tear production. Combination iCAP/antibiotic treatment did not appear to provide significant added benefit over iCAP alone. Our findings suggest that the addition of iCAP may be a viable tool in reducing damage to the cornea and anterior chamber of the eye following S. aureus keratitis.

Solid Dispersion Incorporated into Dissolving Microneedles for Improved Antifungal Activity of Amphotericin B: In Vivo Study in a Fungal Keratitis Model.

Fungal keratitis (FK) is a fungal infection of the cornea, which is part of the eye and causes corneal ulcers and an increased risk of permanent blindness, which is often found in Candida albicans species. Amphotericin B (AMB), which is a group of polyenes as the first-line treatment of FK, is effective in annihilating C. albicans. However, AMB preparations such as eye drops and ointments have major drawbacks, for instance, requiring more frequent administrations, loss of the drug by the drainage process, and rapid elimination in the precornea, which result in low bioavailability of the drug. An ocular dissolving microneedle containing the solid dispersion amphotericin B (DMN-SD-AMB) had been developed using a mixture of poly(vinyl alcohol) (PVA) and poly(vinylpyrrolidone) (PVP) polymers, while the solid dispersion AMB (SD-AMB) was contained in the needle as a drug. This study aims to determine the most optimal and safest DMN-SD-AMB formula for the treatment of FK in the eye as well as a solution to overcome the low bioavailability of AMB eye drops and ointment preparations. SD-AMB had been successfully developed, which was characterized by increased antifungal activity and drug release in vitro compared to other treatments. Furthermore, DMN-SD-AMB studies had also been successfully performed with the best formulation, which exhibited the best ex vivo corneal permeation profile and antifungal activity as well as being safe from eye irritation. In addition, an in vivo antifungal activity using a rabbit infection model shows that the number of fungal colonies was 0.98 ± 0.11 log10 CFU/mL (F3), 5.76 ± 0.32 log10 CFU/mL (AMB eye drops), 4.01 ± 0.28 log10 CFU/mL (AMB ointments), and 9.09 ± 0.65 log10 CFU/mL (control), which differed significantly (p < 0.05). All of these results evidence that DMN-SD-AMB is a new approach to developing intraocular preparations for the treatment of FK.

Continuous voriconazole lavage in managing moderate and severe fungal keratitis: a randomized controlled trial.

PURPOSE: To assess the effectiveness and safety of continuous lavage with 1% voriconazole (CL) for moderate and severe fungal keratitis. METHODS: Thirty-one patients were randomized to receive topical eye drops either alone (T) or combined with continuous 1% voriconazole lavage (CL-T). The primary outcome was the cure rate at 3 months. The secondary outcomes were the 6-day efficacy, 3-day infiltration size and depth, hypopyon height, central corneal thickness (CCT), epithelial defect size, and subject feelings and clinical signs assessment scores. RESULTS: At 3 months, the cure rate was comparable between the groups in patients with moderate fungal keratitis (66.7% vs. 62.5%, P = 0.60). However, among severe cases, 4 cases (44.4%) in the CL-T group healed successfully, while none in the T group; this difference was not significant (P = 0.08), although it was very close to 0.05. This may be related to the small sample size. After 6 days, the percentage of patients with "worsened" ulcers in the CL-T group was lower than that in the T group (0% vs. 31%, P = 0.043). The infiltration size, infiltration depth, and hypopyon height in the CL-T group were smaller than those in the T group after 3 days (all P < 0.05). There was no difference in CCT, epithelial defect size, subject feelings scores, or clinical signs scores between groups. CONCLUSION: These outcomes suggest that CL is an effective and safe adjuvant method for controlling the progression of moderate and severe fungal keratitis. TRIAL REGISTRATION NUMBER: ChiCTR2100050565.

Multi-drug resistant Klebsiella-induced corneal ulcer following pterygium surgery: a case report.

BACKGROUND: Management of pterygium is dependent on the grading of pterygium and its clinical presentation (inflamed or quiescent), and surgical excision is the final choice of treatment for the pterygium extending beyond the limbus. Infectious keratitis is one of the most commonly reported complications in recent years. To the best of our knowledge, Klebsiella keratitis after pterygium surgery has not been described in the current literature. Here, we report a patient with corneal ulcer formation following pterygium surgical excision. CASE PRESENTATION: A 62-year-old woman presented with complaints of pain, blurred vision, photophobia and redness in her left eye for a month. She had a history of pterygium surgical excision two months ago. Slit-lamp examination showed conjunctival congestion, a central whitish corneal ulcer with a central epithelial defect, and hypopyon. Corneal scraped sample revealed multidrug resistant (MDR) Klebsiella pneumonia and the strain was found to be sensitive to cefoxitin and ciprofloxacin. Intracameral cefuroxime (1 mg/0.1 mL) injection, fortified cefuroxime ophthalmic suspension (50 mg/mL) and moxifloxacin ophthalmic suspension (0.5%) were successfully administered to control the infection. Since residual central stromal opacification remained persistent, final visual acuity did not improve beyond finger counting at two meters. CONCLUSIONS: Klebsiella keratitis is a rare and sight-threatening complication following pterygium excision. This report emphasizes the importance of close follow-up examination following pterygium surgeries.

Case Series: Mixed Infectious Keratitis by Pythium insidiosum and Fungal Species.

SIGNIFICANCE: This case series is the first to illustrate mixed infection from Pythium sp. and fungal species in corneal ulcer. PURPOSE: This case series aimed to alert all toward the possibility of both Pythium sp. and fungal species infection in case of nonresponding corneal ulcer treated with either antifungals or antipythium drugs alone. Increased suspicion of mixed infection in case of nonresponding fungal/ Pythium keratitis may facilitate early and prompt management. CASE REPORTS: Six patients presented with signs of either fungal or Pythium keratitis. They underwent ophthalmological examinations, smear examinations, cultures, and polymerase chain reaction (PCR). Therapeutic penetrating keratoplasty was performed in cases where symptoms worsened after treatment with either antifungal or antipythium drugs. The half corneal button (HCB) was shared for histopathological and microbiological examinations. In the first case, smear examination from corneal scraping (CS) revealed Pythium -like filaments, which were confirmed with PCR; however, Aspergillus nidulans grew in culture. In the second case, iodine-potassium iodide (IKI) staining was positive for Pythium ; however, PCR was positive for both Pythium and fungus, which was further confirmed by DNA sequencing. In the third case, IKI staining and HCB were positive for Pythium ; however, PCR was positive for fungus, which was identified as Candida saitoana with DNA sequencing. In the fourth case, Pythium grew in the CS culture; however, Candida sp. grew in the HCB culture. In the fifth case, Cladosporium sp. grew in culture from CS; however, Pythium insidiosum grew from the anterior chamber exudate after therapeutic penetrating keratoplasty. In the sixth case, smear examination revealed septate fungal filaments, and Cladosporium sp. grew in culture; however, HCB on histopathological examination showed features of Pythium keratitis. CONCLUSIONS: In unresponsive cases of Pythium or fungal keratitis, diagnostic modalities such as IKI and PCR should be implemented as a routine practice, in addition to smears and cultures.

In vitro susceptibility of canine corneal bacterial pathogens to three cross-linking protocols.

OBJECTIVE: To assess in vitro antibacterial efficacy of three cross-linking (XL) protocols on bacteria associated with canine ulcerative keratitis. METHODS: Three XL protocols: UVA 3 mW/cm2 for 60 min, UVA 3 mW/cm2 for 30 min, and UVA 30 mW/cm2 for 3 min with and without application of riboflavin and a riboflavin-only protocol were performed in vitro on the four most common bacterial genera isolated from cases of canine ulcerative keratitis treated at Dick White Referrals, UK. Zones of bacterial growth inhibition (GIZ) associated with treatment were measured and compared. RESULTS: The four most common isolates were Pseudomonas aeruginosa (PA) (48/140, 34.3%), Streptococcus spp. (32/140, 22.9%), Staphylococcus spp. (24/140, 17.1%) and Escherichia coli (EC) (11/140, 7.9%). PA, EC, Streptococcus canis (SC), and Staphylococcus pseudintermedius (SP), isolated from canine corneas, were selected for testing. EC and SC demonstrated growth inhibition following all UVA/riboflavin protocols. PA and SP only displayed growth inhibition following the 60 min UVA/riboflavin protocol. GIZ areas for 60 min UVA/riboflavin protocols were significantly greater than 30 and 3 min UVA/riboflavin protocols (p < .01) and there was no significant difference between 30 and 3 min UVA/riboflavin protocols. In respect to GIZ areas, EC was significantly more susceptible to XL than SP (p = <.01). CONCLUSIONS: All UVA/riboflavin XL protocols caused growth inhibition of EC and SC in vitro. PA and SP did not show clear growth inhibition in vitro following exposure to XL protocol settings of UVA 3 mW/cm2 for 30 min and UVA 30 mW/cm2 for 3 min.

Effects of commercial amniotic membrane extract on the re-epithelialization time and the early expression of matrix metalloproteinase-9 in cats with experimentally induced corneal ulcers.

OBJECTIVES: To investigate whether a commercially available amniotic membrane extract (AME) can accelerate corneal wound healing and suppress the early expression of MMP-9 in the tears of cats with experimentally induced superficial ulcerative keratitis. PROCEDURES: A total number of 16 cats were included. At the end of keratectomy, cats in the treatment group (TG, n = 8) received 40 µl of AME (EyeQ® Amniotic Eye Drops, Vetrix®) four times daily, while cats in the control group (CG, n = 8) received 40 µl of saline at the same time points. Tears were collected 24 and 48 h after keratectomy, and the total MMP-9 was quantified by ELISA. RESULTS: The corneal re-epithelialization rate did not differ between groups (p = .26), being 0.48 ± 0.05 mm2 /h in the CG and 0.41 ± 0.03 mm2 /h in the TG. Similarly, the average time to achieve corneal wound healing did not differ between groups (p = .25) and was 61.50 ± 3.54 h in the CG and 70.50 ± 6.71 h in the TG. The dimensions of the ulcerated areas also did not differ at any time point between the groups (p > .05). In both groups, corneas healed without scarring, pigmentation, or vascularization. The expression of MMP-9 in the tears was similar in both groups at 24 h post-keratectomy, with a slight decrease at 48 h (p > .05). CONCLUSIONS: The instillation of a commercial AME (EyeQ®) is safe, but it did not decrease the corneal re-epithelialization time or the early expression of MMP-9 in the tears of cats with experimentally induced superficial ulcerative keratitis in this study.

Presumed erlotinib-induced bilateral corneal ulcers in a dog with lung tumor.

OBJECTIVE: To report the corneal toxicity of erlotinib in dogs. ANIMAL STUDIED: A 13-year-old castrated male Maltese dog. RESULTS: A dog with lung cancer presented with a month-long history of mucoid discharge and blepharospasm in both eyes. Corneal ulcerations with stromal thinning were diagnosed in both eyes, which were refractory after 2 weeks of treatment with topical antibiotics and artificial tears. The dog was orally administered erlotinib (Tarceva®) by his owner for 2 months to treat his lung cancer. Urgent withholding of erlotinib was recommended, and after 2 weeks of discontinuation, the corneal defects resolved; however, corneal thinning remained until the six-month follow-up. CONCLUSIONS: To the best of author's knowledge, this is the first report in the veterinary literature that describes bilateral corneal ulcers associated with erlotinib administration in a dog.

Fungal Keratitis Caused by Talaromyces coalescens: A Case Report.

Fungal keratitis is a severe corneal infection, and the causative fungi include various rare fungal species. Fungal keratitis caused by Talaromyces species has yet to be reported, and there is no information about this fungus as a cause of keratitis. A 77-year-old man developed fungal keratitis while waiting for a donor cornea due to bullous keratopathy in his left eye. Fungal culture of a corneal scraping grew filamentous fungi, which were morphologically identified as Paecilomyces species. The corneal infection did not improve after topical administration of 1% voriconazole, and ribosomal DNA sequencing definitively verified the fungus to be Talaromyces coalescens. The lesion gradually improved after switching to topical 5% natamycin. Antifungal susceptibility tests determined the high minimum inhibitory concentrations of voriconazole to be > 8 µg/mL. This is the first report of Talaromyces fungal keratitis. Clinicians, especially those in ophthalmology, need to be aware of this rare fungus.

Recurrent Filamentous Fungal Keratitis Caused When the Primarily Selected Graft Diameter was Too Small.

PURPOSE: To establish the importance of using a sufficiently large corneal graft in primary penetrating keratoplasty in order to prevent recurrence of fungal keratitis. OBERSERVATIONS: A 58-year-old female patient underwent emergency penetrating keratoplasty (diameter 7.0 mm, double running suture) for therapy-resistant fungal keratitis (Fusarium solani) at an external eye clinic. Despite intensive antifungal therapy, new fungal infiltrates appeared in the host cornea after a few days. The patient was referred to our department for further treatment. On first presentation, circular infiltrates were seen around the corneal graft with anterior chamber involvement and therapy-resistant hypopyon. We performed an emergency penetrating repeat keratoplasty (diameter of 13.0 mm, 32 interrupted sutures) combined with anterior chamber lavage and intracameral and intrastromal drug injection. CONCLUSION AND IMPORTANCE: Fungal keratitis sometimes has a frustrating clinical course. Therefore, early diagnosis with effective therapy initiation is of the utmost importance. In cases of penetrating keratoplasty, optimal planning and timing (before anterior chamber involvement) should be provided. Sufficient safety distance must be ensured in the choice of graft diameter, fixation with multiple interrupted sutures, and anterior chamber lavage, as well as intracameral and intrastromal drug administration. Incomplete excision carries a risk of recurrence and endophthalmitis in the course. Close postoperative control is necessary to detect early recurrences.

Corneal cross-linking guards against infectious keratitis: an experimental model.

BACKGROUND: PACK-CXL (photo-activated chromophore for keratitis-corneal cross-linking) is an alternative option in treatment of corneal infections. It inhibits corneal melting by increasing the stromal resistance, besides the microbicidal effect of photo-activated riboflavin. METHODS: Corneal infection with Pseudomonas aeruginosa and Staph aureus bacteria was induced in 20 eyes of 10 rabbits after 6 weeks of corneal cross-linking in half of the eyes, while the other acted as control group. Clinical and corneal histopathological examination was done to evaluate the extent of inflammation, ulceration, organism penetration, and depth of corneal stromal affection. RESULTS: The control eyes developed severe inflammation compared to the cross-linked eyes. Corneal melting occurred in 6 eyes in the control versus none in cross-linked group. Histopathological examination showed that the inflammation was confined to the superficial part of the stroma with localization of the inflammation in the cross-linked eyes in contrast to the control eyes that showed deep infiltration. CONCLUSION: PACK-CXL provides infection localization through increasing the corneal rigidity and resistance to enzymatic digestion, even in the absence of the riboflavin microbicidal role. So, early PACK-CXL is worth to be considered in the IK treatment algorithm.

Topical Chlorhexidine 0.2% versus Topical Natamycin 5% for the Treatment of Fungal Keratitis in Nepal: A Randomized Controlled Noninferiority Trial.

PURPOSE: To investigate if topical chlorhexidine 0.2%, which is low cost and easy to formulate, is noninferior to topical natamycin 5% for the treatment of filamentous fungal keratitis. DESIGN: Randomized controlled, single-masked, noninferiority clinical trial. PARTICIPANTS: Adults attending a tertiary-level ophthalmic hospital in Nepal with filamentous fungal infection confirmed on smear or confocal microscopy. METHODS: Participants were randomly allocated to receive topical chlorhexidine 0.2% or topical natamycin 5%. Primary analysis (intention-to-treat) was by linear regression, using baseline logarithm of the minimum angle of resolution (logMAR) best spectacle-corrected visual acuity (BSCVA) and treatment arm as prespecified covariates. Mixed fungal-bacterial infections were excluded from the primary analysis but included in secondary analyses and secondary safety-related outcomes. The noninferiority margin was 0.15 logMAR. This trial was registered with ISRCTN, number ISRCTN14332621. MAIN OUTCOME MEASURES: The primary outcome measure was BSCVA at 3 months. Secondary outcome measures included perforation or therapeutic penetrating keratoplasty by 90 days. RESULTS: Between June 3, 2019, and November 9, 2020, 354 eligible participants were enrolled and randomly assigned: 178 to chlorhexidine and 176 to natamycin. Primary outcome data were available for 153 and 151 of the chlorhexidine and natamycin groups, respectively. Of these, mixed bacterial-fungal infections were found in 20 cases (12/153 chlorhexidine, 8/151 natamycin) and excluded from the primary analysis. Therefore, 284 patients were assessed for the primary outcome (141 chlorhexidine, 143 natamycin). We did not find evidence to suggest chlorhexidine was noninferior to natamycin and in fact found strong evidence to suggest that natamycin-treated participants had significantly better 3-month BSCVA than chlorhexidine-treated participants, after adjusting for baseline BSCVA (regression coefficient, -0.30; 95% confidence interval [CI], -0.42 to -0.18; P < 0.001). There were more perforations and emergency corneal grafts in the chlorhexidine arm (24/175, 13.7%) than in the natamycin arm (10/173, 5.8%; P = 0.018, mixed infections included), whereas natamycin-treated cases were less likely to perforate or require an emergency corneal graft, after adjusting for baseline ulcer depth (odds ratio, 0.34; 95% CI, 0.15-0.79; P = 0.013). CONCLUSIONS: Treatment with natamycin is associated with significantly better visual acuity, with fewer adverse events, compared with treatment with chlorhexidine. Natamycin remains the preferred first-line monotherapy treatment for filamentous fungal keratitis.

The role of Glabridin in antifungal and anti-inflammation effects in Aspergillus fumigatus keratitis.

PURPOSE: To investigate the effect of Glabridin (GLD) in Aspergillus fumigatus keratitis and its associated mechanisms. METHODS: Aspergillus fumigatus (A. fumigatus) conidia was inoculated in 96-well plate, and minimal inhibitory concentration (MIC) and biofilm formation ability were evaluated after GLD treatment. Spore adhesion ability was evaluated in conidia infected human corneal epithelial cells (HCECs). Keratitis mouse model was created by corneal intrastromal injection with A. fumigatus conidia, and GLD treatment started at the day after infection. The number of fungal colonies was calculated by plate count, and degree of corneal inflammation was assessed by clinical score. Flow cytometry, myeloperoxidase (MPO), and immunofluorescence staining (IFS) experiments were used to assess neutrophil infiltrations. PCR, ELISA and Western blot were conducted to determine levels of TLR4, Dectin-1 as well as downstream inflammatory factors. RESULTS: GLD treatment suppressed the proliferation, biofilm formation abilities and adhesive capability of A. fumigatus. In mice upon A. fumigatus infection, treatment of GLD showed significantly decreased severity of corneal inflammation, reduced number of A. fumigatus in cornea, and suppressed neutrophil infiltration in cornea. GLD treatment obviously inhibited mRNA and protein levels of Dectin-1, TLR4 and proinflammatory mediators such as IL-1ß, HMGB1, and TNF-α in mice corneas compared to the control group. CONCLUSION: GLD has antifungal and anti-inflammatory effects in fungal keratitis through suppressing A. fumigatus proliferation and alleviating neutrophil infiltration, and repressing the expression of TLR4, Dectin-1 and proinflammatory mediators.