Fusarium Keratitis: A Systematic Review (1969 to 2023).

BACKGROUND: Mycotic keratitis (MK) represents a corneal infection, with Fusarium species identified as the leading cause. Fusarium is a genus of filamentous fungi commonly found in soil and plants. While many Fusarium species are harmless, some can cause serious infections in humans and animals, particularly Fusarium keratitis, that can lead to severe ocular infections, prevalent cause of monocular blindness in tropical and subtropical regions of the world. Due to its incidence and importance in ophthalmology, we conducted a systematic analysis of clinical cases to increase our understanding of Fusarium keratitis by gathering clinical and demographic data. METHODS: To conduct an analysis of Fusarium keratitis, we looked through the literature from the databases PubMed, Embase, Lilacs, and Google Scholar and found 99 papers that, between March 1969 and September 2023, corresponded to 163 cases of Fusarium keratitis. RESULTS: Our analysis revealed the Fusarium solani species complex as the predominant isolate, with females disproportionately affected by Fusarium keratitis. Notably, contact lens usage emerged as a significant risk factor, implicated in nearly half of cases. Diagnosis primarily relied on culture, while treatment predominantly involved topical natamycin, amphotericin B, and/or voriconazole. Surprisingly, our findings demonstrated a prevalence of cases originating from the United States, suggesting potential underreporting and underestimation of this mycosis in tropical regions. This shows the imperative for heightened vigilance, particularly in underdeveloped regions with substantial agricultural activity, where Fusarium infections may be more prevalent than currently reported. CONCLUSION: Our study sheds light on the clinical complexities of Fusarium keratitis and emphasizes the need for further research and surveillance to effectively tackle this vision-threatening condition. Furthermore, a timely identification and early initiation of antifungal treatment appear to be as important as the choice of initial treatment itself.

Favorable outcome of Lasiodiplodia theobromae keratomycosis : a clinical case and systematic review.

BACKROUD: Keratitis caused by Lasiodiplodia theobromae is rare and typically associated with a poor prognosis. Current literature lacks sufficient evidence on effective management of patients with this condition. CASE PRESENTATION: A 74-year-old former agricultural worker presented with a red right eye, discomfort, and decreased visual acuity, progressing over three days without treatment. Examination revealed type 2 diabetes and a non-perforating, spiculated corneal abscess with a hypopyon in the right eye. Initial treatment included a triple antibiotic therapy and supportive care. Direct mycological examination identified numerous septate mycelial filaments. Antifungal treatment with natamycin and voriconazole, both topically and orally, was initiated. Cultures confirmed Lasiodiplodia theobromae. The patient showed significant improvement. Treatment continued for eight weeks, with a final visual acuity of 20/50 due to a stromal scar. CONCLUSION: An extensive literature review conducted in November 2023, using databases such as PubMed and Google Scholar with the keywords "lasiodiplodia" and "keratitis" yielded no previous cases of this specific condition being managed solely with the combined use of natamycin and voriconazole. This antifungal combination is commonly included in most management protocols for fungal keratitis. Factors such as the use of corticosteroids and delayed diagnosis were noted to adversely affect the prognosis. This case and this systematic review underscores the potential for non-surgical management options in severe fungal keratitis.

Platelet-derived biomaterial controls aspergillus fumigatus keratitis by decreasing fungal burden: an in vivo study.

Fungal keratitis is a severe corneal infection characterized by suppurative and ulcerative lesions. Aspergillus fumigatus is a common cause of fungal keratitis. Antifungal drugs, such as natamycin, are currently the first-line treatment for fungal keratitis, but their ineffectiveness leads to blindness and perforation. Additionally, the development of fungal resistance makes treating fungal keratitis significantly more challenging. The present study used platelet-derived biomaterial (PDB) to manage A. fumigatus keratitis in the animal model. Freezing and thawing processes were used to prepare PDB, and then A. fumigatus keratitis was induced in the mice. Topical administration of PDB, natamycin, and plasma was performed; quantitative real-time PCR (qPCR) and histopathologic examination (HE) were used to assess the inhibitory effect of the mentioned compounds against fungal keratitis. The qPCR results showed that PDB significantly decreased the count of A. fumigatus compared to the control group (P-value ≤ 5). Natamycin also remarkably reduced the count of fungi in comparison to the untreated animal, but its inhibitory effect was not better than PDB (P-value > 5). The findings of HE also demonstrated that treatment with PDB and natamycin decreased the fungal loads in the corneal tissue. However, plasma did not show a significant inhibitory effect against A. fumigatus. PDB is intrinsically safe and free of any infections or allergic responses; additionally, this compound has a potential role in decreasing the burden of A. fumigatus and treating fungal keratitis. Therefore, scientists should consider PDB an applicable approach to managing fungal keratitis and an alternative to conventional antifungal agents.

Treatment Strategies for Filamentous Fungi Keratitis.

INTRODUCTION: This research aims to describe clinical findings, epidemiology and treatment outcomes in patients with filamentous fungi keratitis of a tertiary centre in Germany over a 7-year period and to compare the efficacy of different antifungal treatments and the effect of additive topical steroids. METHODS: This retrospective study included 25 eyes of 23 patients from October 2013 to December 2020 with cultural isolates of filamentous fungi and corresponding keratitis. Best-corrected visual acuity (BCVA), clinical signs, symptoms, risk factors and outcome were extracted from medical records. RESULTS: Improvement of BVCA was noted in 68% of eyes. Mean BCVA of the study population increased from 0.75 logMAR [median 0.40, standard deviation (SD) 0.82, range 0-2.3] to 0.48 logMAR (median 0.10, SD 0.88, range - 0.1 to 3). The most commonly used antifungal topical treatment was a combination of natamycin 5% and voriconazole 2% (44% of eyes), followed by voriconazole 2% in 36% of cases. An antiinflammatory topical steroid was applied in 52%. In 16% of the eyes, penetrating keratoplasty (pKP) was performed. CONCLUSION: Diagnosis of filamentous fungi keratitis is often difficult or delayed. Outcomes can be poor even with intensive treatment because of high resistance to common antifungals. Access to natamycin 5% seems to lead to favourable outcomes in filamentous fungi keratitis.

Utilizing confocal microscopy and topical natamycin in metarhizium keratitis of Caribbean origin.

A 32-year-old contact lens-wearing man with recent travel history to the Caribbean was referred for a corneal infiltrate in the left eye that worsened following 1-week of steroid-antibiotic therapy. Corneal cultures were obtained and sent to our facility's clinical microbiology laboratory for analysis. Same-day in vivo confocal microscopy revealed fungal elements. Nucleic acid sequencing performed on the isolated determined it to be a member of the entomopathogenic genus Metarhizium. Over the course of 3 months, the patient's corneal infiltrate ultimately resolved following topical natamycin 5 % therapy. This is the first reported case to have originated in the Caribbean and to utilize in vivo confocal microscopy to aid diagnosis. Our case also supports previous reports of success with natamycin therapy in treatment of Metarhizium sp. keratitis.

Mesoporous zinc oxide-based drug delivery system offers an antifungal and immunoregulatory strategy for treating keratitis.

Fungal keratitis is a refractory eye disease that is prone to causing blindness. Fungal virulence and inflammatory responses are two major factors that accelerate the course of fungal keratitis. However, the current antifungal drugs used for treatment usually possess transient residence time on the ocular surface and low bioavailability deficiencies, which limit their therapeutic efficacy. In this work, natamycin (NATA)-loaded mesoporous zinc oxide (Meso-ZnO) was synthesized for treating Aspergillus fumigatus keratitis with excellent drug-loading and sustained drug release capacities. In addition to being a carrier for drug delivery, Meso-ZnO could restrict fungal growth in a concentration-dependent manner, and the transcriptome analysis of fungal hyphae indicated that it inhibited the mycotoxin biosynthesis, oxidoreductase activity and fungal cell wall formation. Meso-ZnO also promoted cell migration and exhibited anti-inflammatory role during fungal infection by promoting the activation of autophagy. In mouse models of fungal keratitis, Meso-ZnO/NATA greatly reduced corneal fungal survival, alleviated tissue inflammatory damage, and reduced neutrophils accumulation and cytokines expression. This study suggests that Meso-ZnO/NATA can be a novel and effective treatment strategy for fungal keratitis.

Improved Topical Ophthalmic Natamycin Suspension for the Treatment of Fungal Keratitis.

Purpose: Natamycin (NT) is used as a first-line antifungal prescription in the treatment of fungal keratitis (FK) and is commercially available as a 5% w/v ophthalmic suspension. NT shows poor water solubility and light sensitivity. Thus, the present investigation is aimed to enhance the fraction of NT in solution in the commercial formulation by adding cyclodextrins (CDs), thereby improving the delivery of the drug into deeper ocular tissues. Methods: The solubility of NT in different CDs, the impact of ultraviolet (UV) light exposure, stability at 4°C and 25°C, in vitro release, and ex vivo transcorneal permeation studies were performed. Results: NT exhibited the highest solubility (66-fold) in randomly methylated-ß-cyclodextrin (RM-ßCD) with hydroxypropyl-ßCD (HP-ßCD) showing the next highest solubility (54-fold) increase in comparison to market formulation Natacyn® as control. The stability of NT-CD solutions was monitored for 2 months (last-time point) at both storage conditions. The degradation profile of NT in NT-RM-ßCD and NT-HP-ßCD solutions under UV-light exposure followed first-order kinetics exhibiting half-lives of 1.2 h and 1.4 h, respectively, an almost 3-fold increase over the control solutions. In vitro release/diffusion studies revealed that suspensions containing RM-ßCD and HP-ßCD increased transmembrane flux significantly (3.1-fold) compared to the control group. The transcorneal permeability of NT from NT-RM-ßCD suspension exhibited an 8.5-fold (P < 0.05) improvement compared to Natacyn eyedrops. Furthermore, the addition of RM-ßCD to NT suspension increases the solubilized fraction of NT and enhances transcorneal permeability. Conclusion: Therefore, NT-RM-ßCD formulations could potentially lead to a decreased frequency of administration and significantly improved therapeutic outcomes in FK treatment.

The preparation and therapeutic effects of ß-glucan-specific nanobodies and nanobody-natamycin conjugates in fungal keratitis.

Fungal keratitis (FK) is a severe infectious corneal disease. Since traditional eye drops exhibit poor dissolution and high corneal toxicity, the efficacy of current treatments for FK remains limited. It is needed to develop new approaches to control the cornea damage from FK. In this study, a nanobody (Nb) specific to ß-glucan in the fungal cell wall was prepared. The conjugate of the Nb with natamycin (NAT), a traditional antifungal drug, was synthesized. Firstly, we found the Nb specific to ß-glucan inhibited fungal growth by disrupting cell wall and biofilm formation.. In addition, the content of ß-glucan in the fungal cell wall decreased after Nb treatment. The Nb also reduced the adhesion ability of fungal conidia to human corneal epithelial cells (HCECs). Further, we examined the difference between NAT and Nb-NAT in antifungal growth. Nb-NAT showed better antifungal effects than NAT which was caused by the interaction between Nb and ß-glucan. Moreover, Nb concentration below 0.5 mg/mL did not affect the viability of HCECs. Nb-NAT had less cytotoxicity and ocular surface irritation than NAT. Nb specific to ß-glucan attenuated Aspergillus fumigatus (A. fumigatus) virulence and relieved inflammatory responses in FK. Nb-NAT treatment of the cornea improved therapeutic effects compared with NAT. It decreased clinical scores and expression level of inflammatory factors. To our knowledge, this study is the first to report a Nb specific to ß-glucan and Nb-NAT for the treatment of FK. These unique functions of the Nb specific to ß-glucan and Nb-NAT would render it as an alternative molecule to control fungal infections including FK. STATEMENT OF SIGNIFICANCE: Fungal keratitis is a corneal disease with a high rate of blindness. Due to the poor dissolution and high corneal toxicity exhibited by traditional eye drops, the efficacy of current therapeutic treatments for fungal keratitis (FK) remains limited. To enhance the therapeutic effect of natamycin in treating fungal keratitis, this study developed an innovative approach by preparing a ß-glucan-specific nanobody and loading it with the antifungal drug natamycin. The ß-glucan-specific nanobody has the ability to control both fungal pathogen invasion and inflammation, which can cause damage to the cornea in FK. The conjugation with the ß-glucan-specific nanobody significantly increased the antifungal capacity of natamycin and reduced its toxicity. The further application of natamycin conjugated with the ß-glucan-specific nanobody could be expanded to other diseases caused by fungal pathogen infections.

Natamycin Ocular Delivery: Challenges and Advancements in Ocular Therapeutics.

Fungal keratitis, an ocular fungal infection, is one of the leading causes of monocular blindness. Natamycin has long been considered the mainstay drug used for treating fungal keratitis and is the only US Food and Drug Administration (USFDA)-approved drug, commercially available as a topical 5% w/v suspension. Furthermore, ocular fungal infection treatment takes a few weeks to months to recover, and the available marketed antifungal suspensions are associated with poor residence time, limited bioavailability (< 5%) and high dosing frequency as well as minor irritation and discomfort. Despite these challenges, natamycin is still the preferred drug choice for treating fungal keratitis, as it has fewer side effects and less ocular toxicity and is more effective against Fusarium species than other antifungal agents. Several novel therapeutic approaches for the topical delivery of natamycin have been reported to overcome the challenges posed by the conventional dosage forms and to improve ocular bioavailability for the efficient management of fungal keratitis. Current progress in the delivery systems uses approaches aimed at improving the corneal residence time, bioavailability and antifungal potency, thereby reducing the dose and dosing frequency of natamycin. In this review, we discuss the various strategies explored to overcome the challenges present in ocular drug delivery of natamycin and improve its bioavailability for ocular therapeutics.

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.

Pentoxifylline treats Aspergillus fumigatus keratitis by reducing fungal burden and suppressing corneal inflammation.

Fungal keratitis (FK) is a blinding ocular disease, which mainly results from fungal damage and excessive inflammation. Pentoxifylline, a kind of methylxanthine, has been discovered to have anti-inflammatory properties in various infectious diseases, hinting a potential therapeutic effect on treating corneal fungal infection. Whereas, the therapeutic impact of pentoxifylline on fungal keratitis is still uncertain. This study investigated the antifungal capability against Aspergillus fumigatus and the anti-inflammatory role of pentoxifylline by activating nuclear factor, erythroid 2 like 2 (Nrf2)/heme oxygenase1 (HO1) pathway in the process of FK. In our research, we demonstrated that pentoxifylline could effectively inhibit fungal growth and inflammatory reaction. Pentoxifylline reduced the production of pro-inflammatory factors by stimulating the Nrf2/HO1 pathway. Although there was no statistical difference between the curative efficacy of pentoxifylline and natamycin application to FK, pentoxifylline could promote corneal epithelial repair and was less toxicity to the ocular surface than natamycin. In conclusion, pentoxifylline performs antifungal and anti-inflammatory effects by lessening the fungus burden and activating the Nrf2/HO1 pathway, hinting that it has the potential to be a new therapeutic medication for Aspergillus fumigatus keratitis.

The therapeutic role and mechanism of 4-Methoxycinnamic acid in fungal keratitis.

Fungal keratitis is an infectious vision-threatening disease that has a poor prognosis, and the clinical therapeutic drugs have multiple limitations, such as epithelial toxicity and low bioavailability. Therefore, new antifungal treatment strategies must be developed. 4-Methoxycinnamic acid (MCA) is a widely occurring natural phenolic acid that has been proven to have multiple effects, such as antibacterial, antifungal, anti-inflammatory, neuroprotective, and inhibiting cancer. In this research, we explored the effects and underlying mechanisms of MCA on A. fumigatus keratitis and the antifungal effects of the combination of MCA and natamycin (NATA) on A. fumigatus. We found that MCA exerts antifungal effects by inhibiting the synthesis of the fungal cell wall, changing the permeability of fungal cell membranes. Moreover, the MCA-NATA combination exhibited synergy for A. fumigatus. In addition, MCA exerted an anti-inflammatory effect by downregulating the inflammatory factors (IL-1ß, TNF-α, IL-6, and iNOS) in C57BL/6 mice and RAW264.7 cells. The anti-inflammatory mechanism of MCA was associated with the Mincle signal pathway. In summary, MCA acts as a potential therapeutic drug for fungal keratitis and a potential antifungal sensitizer for natamycin. MCA inhibits fungal cell wall synthesis, destroys the permeability of fungal cell membranes, and mediates the anti-inflammatory, immune response of the host.

Therapeutic effect of Atractylenolide I on Aspergillus fumigatus keratitis by affecting MyD88/ NF-κB pathway and IL-1ß, IL-10 expression.

PURPOSE: Atractylenolide I (AT-I) is a natural sesquiterpene with anti-inflammatory effects. The purpose of this study was to research the anti-inflammatory effect of AT-I on Aspergillus fumigatus(A. fumigatus) keratitis in mice. METHODS: Cytotoxicity test and cell scratch test were used to determine the therapeutic concentrations of corneal infections. In vivo and in vitro studies, mouse cornea and human corneal epithelial cells (HCECs) infected with A. fumigatus were treated with AT-I or dimethyl sulfoxide (DMSO). Then, to analyze the effect of AT-I on inflammatory response, namely neutrophil or macrophage recruitment and the expression of cytokines involving MyD88, NF-κB, interleukin 1ß (IL-1ß) and interleukin 10 (IL-10). To study the effects of the drug, the techniques used include slit-lamp photography, immunofluorescence, myeloperoxidase (MPO) detection, quantitative real-time polymerase chain reaction (QRT-PCR), and western blot. At the same time, in order to explore the combined effect of the drug and natamycin, slit-lamp photographs and clinical scores were used to visually display the disease process. RESULTS: No cytotoxicity was observed under the action of AT-I at a concentration of 800 µM. In mouse models, AT-I significantly suppressed inflammatory responses, reduced neutrophil and macrophage recruitment, and decreased myeloperoxidase levels early in infection. Studies have shown that AT-I may reduce the levels of IL-1ß and IL-10 by inhibiting the MyD88/ NF-κB pathway. The drug combined with natamycin can increase corneal transparency in infected mice. CONCLUSION: AT-I may inhibit MyD88 / NF-κB pathway and the secretion of inflammatory factors IL-1 ß and IL-10 to achieve the therapeutic effect of fungal keratitis.

Acremonium keratitis: Risk factors, clinical characteristics, management, and outcome in 65 cases.

Purpose: To study the risk factors, clinical presentation, management options, and outcomes in cases of culture-proven Acremonium keratitis. Methods: Medical and microbiology records of culture-proven Acremonium keratitis from Jan 2007 to Dec 2019 at a tertiary eye care center were reviewed. Details of clinical findings on each visit and operating notes were reviewed from the medical records. All cases were subjected to corneal scraping at the first visit for microbiological investigation consisting of direct smear examination and culture. Topical natamycin 5% was the mainstay of medical treatment. Surgical treatment was considered for nonresponding patients. Results: During the 13-year study period, 65 cases of culture-proven Acremonium keratitis were identified out of 1605 cases of fungal keratitis. Trauma was the most common predisposing factor in 32 cases (49.2%). The average area of the corneal stromal infiltrate was 24.8 mm2 at the initial presentation. Hypopyon at the time of presentation was evident in 28 (43.1%) cases. Staphylococcus spp. was the most common (n = 22, 33.8%) organism coexistent with Acremonium. Direct microscopy of corneal scraping was positive for fungal filaments in 57/65 (87.6%) cases. Medical management alone was given in 44 patients (67.6%). Age (>50 years) and treatment delay (>15 days) were found to be independent risk factors for the poor final visual outcome (VA <20/60). Conclusion: When treated early, Acremonium keratitis responds well to medical therapy with currently available topical antifungals. However, advanced and nonresponding cases require surgical intervention for resolution of the infection.

Mechanism of antifungal activity and therapeutic action of ß-ionone on Aspergillus fumigatus keratitis via suppressing LOX1 and JNK/p38 MAPK activation.

PURPOSE: To investigate the anti-inflammatory and antifungal role of ß-ionone (BI) in fungal keratitis (FK). METHODS: In vitro antifungal activity of BI against Aspergillus fumigatus (A. fumigatus) was evaluated by using minimum inhibitory concentration (MIC), crystal violet staining, biofilm biomass measurement, propidium iodide uptake test, and adherence assay. And RT-PCR was carried out to measure the levels of RodA, RodB, Rho, FKs, CshA-D, RlmA, Cyp51A-B and Cdr1B. Network pharmacology analysis was applied to predict the relationship between BI and FK. Cell Count Kit-8 (CCK8) assay was utilized to detect the cytotoxicity of BI to RAW264.7 and immortalized human corneal epithelial cells (HCECs). The underlying mechanism of BI at regulating the level of inflammatory factors in FK was assessed by RT-PCR, ELISA and Western blot in vitro and in vivo. The therapeutic effect of BI has investigated in A. fumigatus keratitis by employing the clinical score, pathological examination, plate count, immunofluorescence and myeloperoxidase (MPO) assay. We also used the slit-lamp microscopy, clinical scores, and HE staining to assess the effect of natamycin compared with BI treatment in vivo. RESULTS: BI suppressed the growth of A. fumigatus and had a significant effect on A. fumigatus biofilms and membrane permeability. RT-PCR demonstrated that exposure of A. fumigatus to BI inhibited the expression of genes that function in hydrophobin (RodA, RodB), cell wall integrity (Rho, FKs, CshA-D, RlmA), azole susceptibility (Cyp51A-B, Cdr1B). Network pharmacology showed that the effects of BI in FK implicate with C-type lectin receptor signaling pathway. In vivo, after A. fumigatus infection, BI treatment markedly reduced the severity of FK by decreasing clinical score, neutrophil recruitment, and fungal load. And BI treatment also obviously reduced the expression of inflammatory cytokines, Lectin-like oxidized LDL receptor (LOX-1), phosphorylation of p38MAPK and p-JNK versus the DMSO-treated group. BI and natamycin both significantly increased corneal transparency and decreased inflammatory cell recruitment in the FK in the mice model. CONCLUSION: These results indicated that BI had fungicidal activities against A. fumigatus. It also ameliorated FK in mice by reducing inflammation, which was regulated by LOX-1, p-p38MAPK and p-JNK.

Clinical Profile and Treatment Outcomes of Patients With Acremonium Species Positive Keratitis Managed in a Tertiary Eye Care Center.

PURPOSE: To report the clinical profile and treatment outcomes of patients with culture-positive Acremonium keratitis. METHODS: This is a retrospective observational study. Medical records of all patients treated in a tertiary eye hospital for culture positive infective keratitis from March 2016 to February 2021 were screened, of which those positive for Acremonium species on fungal culture were reviewed. Demographic details, clinical presentation, clinical course, treatment given, total follow-up duration, time taken for ulcer to heal, scar size, and final visual acuity in the last follow-up were recorded. RESULTS: Fifty three cases of fungal keratitis caused by Acremonium species were identified, 22 females and 31 males, with average age of 46.39±18.64 years. The mean duration of symptoms being 54.47±50 days. Only five patients had a history of trauma with vegetative matter. Clinical presentation of patients showed a large number of variations, with 2 patients presenting as peripheral ulcerative keratitis and 1 with epithelial plaque. The mean visual acuity of patients at presentation was 2.43±0.46 logMAR units. Thirty-three of 53 patients presented with perforated corneal ulcer and underwent penetrating keratoplasty; 20 patients were medically managed on topical voriconazole 1%, natamycin 5%, and oral voriconazole. The mean duration of healing of epithelial defect was 95±60.62 days (range 60-165 days). CONCLUSION: Acremonium keratitis has a long and indolent course. A prolonged combination therapy of natamycin and voriconazole seems to be effective in the management. A delay in the diagnosis of Acremonium keratitis often leads to clinical worsening requiring keratoplasty.

In vitro antifungal susceptibility of Fusarium species and Aspergillus fumigatus cultured from eleven horses with fungal keratitis.

PURPOSE: To examine the relationship between Minimum Inhibitory Concentration (MICs) and response to therapy of 6 Fusarium spp. and 5 Aspergillus fumigatus isolated from equine ulcerative keratitis cases. PROCEDURE: Fungi were identified by morphology and Internal Transcribed Spacer (ITS) polymerase chain reaction (PCR) with sequencing and evaluated at the University of Texas Fungal Testing Laboratory for susceptibility to three azole antifungals (miconazole, voriconazole, posaconazole), natamycin, and two echinocandins (anidulafungin, caspofungin). A Mann-Whitney rank sum test was used for the comparison of time to heal between infections of different fungal genera and in vitro susceptibility to the drug administered. RESULTS: Fusarium spp. were resistant to azole antifungals in 6/6 cases (100%). Fusarium spp. were susceptible to echinocandins and natamycin in all cases. A. fumigatus was resistant to anidulafungin in 1/5 cases (20%) and posaconazole in 1/5 cases (20%) The remainder of A. fumigatus isolates were susceptible to all antifungal agents tested. Fusarium isolates were treated with antifungals to which they were not susceptible; however, all cases of A. fumigatus were treated with antifungals to which they were susceptible. All Fusarium cases and A. fumigatus cases experienced clinical resolution, regardless of surgical intervention. There was no statistical correlation between fungal genus and time to heal (p < .082). CONCLUSIONS: The in vitro susceptibility indicated that all cases of Fusarium spp. were resistant to azole antifungal drugs which were used as treatment. Clinical outcomes, however, showed that all cases healed despite resistance to antifungals.

Laccase-mediated functionalization of natamycin by gallic acids for the therapeutic effect on Aspergillus fumigatus keratitis.

To improve the therapeutic effect of natamycin on fungal keratitis (FK), the grafted derivatives of natamycin and gallic acid were obtained, and the effects of the grafted derivatives on Aspergillus fumigatus (A. fumigatus) keratitis were investigated. The structure of natamycin grafted with gallic acid was identified by FT-IR and UV-Vis, and the successful synthesis of Gallic-Natamycin (GA-NAT) was proved. CCK-8 and the Draize eye test showed that GA-NAT had less cytotoxicity. Then, through in vitro antibacterial experiments such as minimum inhibitory concentration (MIC), adhesion, biofilm formation, and calcium fluorescence staining and in vivo experiments such as clinical score and plate counting, the results showed that GA-NAT had similar antifungal activity to natamycin, but had a better therapeutic effect than natamycin. Myeloperoxidase assay and immunofluorescence staining also showed that GA-NAT significantly inhibited neutrophil recruitment and activity. Moreover, It was further found that GA-NAT could inhibit the mRNA and protein expressions of LOX-1, TNF-α, and IL-1ß. These results indicated that GA-NAT inhibited the fungal growth, reduced the neutrophil infiltration into cornea, and down-regulated the expression of inflammatory factors in lesions, which provides a new choice for FK treatment.

Thermosensitive Tri-Block Polymer Nanoparticle-Hydrogel Composites as Payloads of Natamycin for Antifungal Therapy Against Fusarium Solani.

Purpose: Fusarium Solani is the principal pathogen associated with fungal keratitis. As a sensitive drug to F. Solani, natamycin (NAT) was limited by the poor penetration and low bioavailability in clinical application. The aim of this study was to develop a new type of tri-block polymer nanoparticle-gel complex (Gel@PLGA-PEI-PEG@NAT) for delivering NAT and evaluate its physicochemical properties, antifungal activity, safety, penetrability, adhesion, and efficacy in treating fungal keratitis. Methods: PLGA-PEI-PEG@NAT was prepared and characterized with a nano-particle size analyzer, transmission electron microscopy (TEM), scanning electron microscopy (SEM), and Fourier transform infrared spectroscopy (FTIR). The minimum inhibitory concentration (MIC), cytotoxicity, penetrability of NAT (Natacyn® 5% ophthalmic suspension; Alcon) and PLGA-PEI-PEG@NAT with different concentrations were assessed. The eye surface retention time, ocular irritation, and curative effect of the NAT ophthalmic suspension and Gel@PLGA-PEI-PEG@NAT on a rabbit fungal keratitis model were evaluated. Results: PLGA-PEI-PEG@NAT had a particle size of 150 nm, a positive surface charge, and a sustained-release effect. The MIC for F. Solani was 2 µg/mL. A cytotoxicity test and ocular irritation test showed that PLGA-PEI-PEG@NAT and Gel@PLGA-PEI-PEG@NAT had good biocompatibility and no obvious irritation for rabbit corneas. Penetration experiments confirmed that PLGA-PEI-PEG@NAT can successfully enter corneal epithelial cells and through the cornea to enter the anterior chamber. Compared with NAT ophthalmic suspension, Gel@PLGA-PEI-PEG@NAT had stronger cornea permeation at the same concentration. The therapeutic effect and precorneal retention ability of the NAT ophthalmic suspension and Gel@PLGA-PEI-PEG@NAT on the fungal keratitis rabbit model were compared. Gel@PLGA-PEI-PEG@NAT achieved a better therapeutic effect at a lower drug concentration, and its eye surface retention time was significantly longer than that of the NAT ophthalmic suspension. Conclusion: Gel@PLGA-PEI-PEG@NAT was shown to be a safe and effective nanodrug delivery system for NAT. It has great potential to improve the cure rate of fungal keratitis, reduce the administration frequency during the treatment process, and improve patient compliance.

Patterns of Antifungal Resistance in Adult Patients With Fungal Keratitis in South India: A Post Hoc Analysis of 3 Randomized Clinical Trials.

IMPORTANCE: Antifungal resistance has been shown to impact treatment success, but research analyzing antifungal resistance is scarce. OBJECTIVE: To evaluate changes in antifungal resistance over time. DESIGN, SETTING, AND PARTICIPANTS: Ad hoc analysis of 3 randomized clinical trials including consecutive patients 18 years and older presenting with smear-positive fungal ulcers to Aravind Eye Hospitals in Madurai, Coimbatore, Pondicherry, and Tirunelveli in South India who participated in 1 of 3 clinical trials: the Mycotic Ulcer Treatment Trials (MUTT) I (2010 to 2011) or II (2010 to 2015) or the Cross-Linking Assisted Infection Reduction (CLAIR) trial (2016 to 2018). This post hoc analysis was designed in March 2021 and data were analyzed in May and November 2021. INTERVENTIONS: Minimum inhibitory concentration (MIC) of natamycin and voriconazole was determined from corneal cultures obtained using standardized methods outlined in the Clinical and Laboratory Standards Institute. MAIN OUTCOMES AND MEASURES: The primary outcome of this post hoc analysis was MIC of natamycin and voriconazole. RESULTS: A total of 890 fungal isolates were obtained from 651 patients (mean [SD] age, 49.6 [13.0]; 191 [43.3%] female) from 2010 to 2018. MICs were available for 522 samples in 446 patients. Fungal isolates overall demonstrated a 1.02-fold increase per year in voriconazole resistance as measured by MICs (95% CI, 1.00-1.04; P = .06). In subgroup analyses, Fusarium species demonstrated a 1.04-fold increase in voriconazole resistance per year (95% CI, 1.00-1.06; P = .01). Fungal isolates showed a 1.06-fold increase in natamycin resistance per year overall (95% CI, 1.03-1.09; P < .001). Fusarium species had a 1.06-fold increase in natamycin resistance (95% CI, 1.05-1.08; P < .001), Aspergillus had a 1.09-fold increase in resistance (95% CI, 1.05-1.15; P < .001), and other filamentous fungi had a 1.07-fold increase in resistance to natamycin per year (95% CI, 1.04-1.10; P < .001). CONCLUSIONS AND RELEVANCE: This post hoc analysis suggests that susceptibility to both natamycin and voriconazole may be decreasing over the last decade in South India. While a trend of increasing resistance could impact treatment of mycoses in general and infectious fungal keratitis in particular, further study is needed to confirm these findings and determine their generalizability to other regions of the world. TRIAL REGISTRATION: ClinicalTrials.gov Identifiers: NCT00996736 and NCT02570321.