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.

Films of biopolymers, pectin and gellan, enriched with natamycin and clove essential oils for the packaging of Corn tortilla: Protection against Staphylococcus aureus and Candida parapsilosis.

It was investigated the microbial protection of corn tortilla, traditional Mexican food with high acceptance, for food safety. We elaborated a functional film (FF) prepared with 0.4% (w/v) gellan gum, 2% (w/v) citrus pectin, 0.5% (w/v) glycerol, 0.0003% (w/v) natamycin, 0.03% (v/v) essential clove oils, and 0.1% (v/v) tween 80. The FF impeded the growth of indicator microorganisms in corn tortilla medium: Staphylococcus aureus (i.e., 35 °C, 50% RH, 7 days) and Candida parapsilosis (i.e., 27 °C, 42% RH, 7 days; and 9 °C, 95% RH, 30 days). In packaged artisanal corn tortilla storage at 22 °C and 50% RH for 30 days, the FF-treatments showed 5.5 log CFU/g total aerobes and 4.8 log CFU/g yeasts and moulds, being two and three logs lower than the concentrations recorded in the controls with no film, respectively. Some physical-mechanical properties of FF were Young's modulus, 500 MPa; elongation at break, 10%; stress at break, 18.5 MPa; oxygen permeability, 4 × 10-13 g m Pa-1 s-1 m-2; and water vapour permeability, 4.8 × 10-11 g m Pa-1 s-1 m-2. Also, the sensory evaluation of wrapped tortilla suggested no negative effects. The obtained results envisage potential food-packaging applications with the elaborated films.

Identification and Fungicide Sensitivity of Colletotrichum spp. from Apple Flowers and Fruitlets in Brazil.

Glomerella leaf spot (GLS) and bitter rot (BR), caused by Colletotrichum spp., are major diseases on apple in southern Brazil. Among integrated pest management tools for disease management in commercial orchards, fungicides remain an important component. This study aimed to identify Colletotrichum spp. from cultivar Eva in Paraná state orchards; evaluate their in vitro sensitivity to cyprodinil, tebuconazole, iprodione, and fluazinam; and determine the baseline in vitro sensitivity of these isolates to benzovindiflupyr and natamycin. Most isolates belonged to Colletotrichum melonis and C. nymphaeae of the C. acutatum species complex. The two species varied in sensitivity to fluazinam and tebuconazole, but no variability was found for any other fungicide. The lowest 50% effective concentration (EC50) values of Colletotrichum spp. were observed for cyprodinil (mean EC50 < 0.02) and benzovindiflupyr (mean EC50 < 0.05); EC50 values were intermediate for fluazinam (mean EC50 < 0.33) and tebuconazole (mean EC50 < 0.14), and they were highest for natamycin (mean EC50 < 5.56) and iprodione (mean EC50 > 12). Cyprodinil and fluazinam are registered for use in Brazil for apple disease management but not specifically for GLS and BR. Tebuconazole is one of the few products registered for Colletotrichum spp. control in apples. In conclusion, flowers and fruitlets can serve as sources of inoculum for GLS and BR disease; C. acutatum was the predominant species complex in these tissues; cyprodinil and fluazinam applications may suppress GLS and BR; and benzovindiflupyr and natamycin warrant further investigation for GLS and BR disease control of apple due to comparably high in vitro sensitivity.

Sodium alginate/gum arabic/glycerol multicomponent edible films loaded with natamycin: Study on physicochemical, antibacterial, and sweet potatoes preservation properties.

Sweet potato (Ipomonea batatas Lam) is easily damaged due to its thin skin, which is limited in shelf life and causes enormous economic losses in the food industry. A new type of safe, non-toxic, and edible antibacterial functional film was developed with sodium alginate (2.5 %), gum arabic (1 %), glycerol (2 %), and natamycin as an antimicrobial agent in this study. The physical and antibacterial properties of films, such as thickness, chromaticity, water vapor permeability, tensile strength, and elongation at break, were studied. Furthermore, the antibacterial film was applied in the preservation of sweet potatoes. The results showed that natamycin emulsion had good compatibility with sodium alginate. Besides reducing the transparency of the composite membrane, the mechanical properties, barrier properties, and thermal stability of the composite film were significantly enhanced by the addition of natamycin prepared by a pH-cycle method. When the concentration of natamycin in the membrane solution reached 40 µg/mL or more, the antibacterial film had a noticeable inhibitory effect on the growth of molds, and yeasts, significantly enhancing the bacteriostatic effect of the base film. During the sweet potatoes storage, the water content, total starch content, Vc content, and flavonoid glycoside content of sweet potato showed a downward trend. However, the treatment of antibacterial film containing natamycin could slow down the physiological and quality changes of sweet potatoes during conventional storage, and the sweet potatoes still had good processing quality after 120 days of storage.

Postharvest sour rot control in lemon fruit by natamycin and an Allium extract.

Citrus sour rot caused by Geotrichum citri-aurantii is one of the most important postharvest diseases in citrus fruit, causing huge economic losses. Traditionally, it has been controlled by the postharvest application of guazatine and propiconazole fungicides, but restrictions in their use make it urgent to find an alternative for sour rot management. Natamycin, a common food preservative, and the organosulfuric compounds extracted from Allium species are safe food additives that control different foodborne pathogens. In the present study, the curative activities of commercial formulations of natamycin (Fruitgard Nat 20) and an Allium extract (PTSO: propyl thiosulfinate oxide; Proallium FRD®), were evaluated for the control of G. citri-aurantii in artificially inoculated lemon fruit. Trials in laboratory and in commercial conditions were carried out to explore the feasibility of including both compounds as part of a safe postharvest sour rot disease control strategy. Under controlled laboratory conditions, sour rot was significatively reduced by 500 mg L-1 of natamycin, 580 mL L-1 of PTSO and 290 mL L-1 of PTSO + 4% of a food coat, applied by immersion. Nevertheless, the maximum dose of PTSO (580 mL L-1) caused phytotoxicity on the fruit rind. In commercial drenching conditions, 290 mL L-1 of PTSO + 4% of a food coat reduced sour rot incidence similar to conventional treatment. In a packing line treatment, spray application of 500 mg L-1 of natamycin with a previous dip in sodium bicarbonate, resulted in nearly 70% reduction of disease incidence compared to conventional salt application. A second commercial experiment revealed that fruit drenching with 290 mL L-1 of PTSO + 4% food coat followed by an in-line cascade application of 500 mg L-1 of natamycin is completely effective for sour rot control after 20 days at 5 °C. Further exposure at room temperature for 7 d showed a 61% reduction in sour rot incidence compared to the control. Results revealed that natamycin and PTSO are promising tools for sour rot control used alone or combined as part of an integrated postharvest strategy.

The combined utilization of Chlorhexidine and Voriconazole or Natamycin to combat Fusarium infections.

BACKGROUND: Fusarium species are the fungal pathogens most commonly responsible for the mycotic keratitis, which are resistant to the majority of currently available antifungal agents. The present study was designed to assess the efficacy of a combination of low doses chlorhexidine with two other commonly used drugs (voriconazole and natamycin) to treat Fusarium infections. RESULTS: We utilized combinations of chlorhexidine and natamycin or voriconazole against 20 clinical Fusarium strains in vitro using a checkerboard-based microdilution strategy. In order to more fully understand the synergistic interactions between voriconazole and chlorhexidine, we utilized a Galleria mellonella model to confirm the combined antifungal efficacy of chlorhexidine and voriconazole in vivo. We found that for voriconazole, natamycin, and chlorhexidine as single agents, the minimum inhibitory concentration (MIC) ranges were 2-8, 4-16, and > 16 µg/ml, respectively. In contrast, the MIC values for voriconazole and chlorhexidine were reduced to 0.25-1 and 1-2 µg/ml, respectively, when these agents were administered in combination, with synergy being observed for 90% of tested Fusarium strains. Combined chlorhexidine and natamycin treatment, in contrast, exhibited synergistic activity for only 10% of tested Fusarium strains. We observed no evidence of antagonism. Our in vivo model results further confirmed the synergistic antifungal activity of chlorhexidine and voriconazole. CONCLUSIONS: Our results offer novel evidence that voriconazole and chlorhexidine exhibit synergistic activity when used to suppress the growth of Fusarium spp., and these agents may thus offer value as a combination topical antifungal treatment strategy.

The crucial role of yeasts in the wet fermentation of coffee beans and quality.

The objective of this study was to investigate the role of yeasts in the wet fermentation of coffee beans and their contribution to coffee quality using a novel approach. Natamycin (300 ppm) was added to the fermentation mass to suppress yeast growth and their metabolic activities, and the resultant microbial ecology, bean chemistry and sensory quality were analyzed and compared to non-treated spontaneous fermentation we reported previously. The yeast community was dominated by Hanseniaspora uvarum and Pichia kudriavzevii and grew to a maximum population of about 5.5 log CFU/g in the absence of Natamycin, while when Natamycin was added yeasts were suppressed. The major bacterial species in both the spontaneous and yeast-suppressed fermentations included the lactic acid bacteria Leuconostoc mesenteroides and Lactococcus lactis, the acetic acid bacteria Gluconobacter cerinus and Acetobacter persici and the Enterobacteriaceae Enterobacter, Citrobacter and Erwinia. For both fermentations, the mucilage layers were completely degraded by the end of the process and the absence of yeast activities had no significant impact on mucilage degradation. During fermentation, reducing sugars were consumed while lactic acid was accumulated inside the beans, and its concentration was significantly higher in the spontaneous fermentation (3 times) than that where yeasts were suppressed by Natamycin. Glycerol was detected with a concentration of 0.08% in the absence of Natamycin and was not identified when Natamycin was added. Green beans fermented with yeast growth contained a higher amount of isoamyl alcohol (21 times), ethanol (3.7 times), acetaldehyde (8 times), and ethyl acetate (25 times) compared to beans fermented in the absence of yeast activities, which remained higher in the former after roasting. Beans fermented without yeast activities had a mild fruity aroma, and lower sensory scores of fragrances (7.0), flavor (6.5), acidity (6.3), body (7.0) and overall score (6.5) compared to the former. These findings demonstrated the crucial roles of yeasts in wet fermentation of coffee beans and for producing high quality coffee.

Cross-Linking-Assisted Infection Reduction: A Randomized Clinical Trial Evaluating the Effect of Adjuvant Cross-Linking on Outcomes in Fungal Keratitis.

PURPOSE: To determine if there is a benefit to adjuvant corneal crosslinking (CXL) and to compare natamycin versus amphotericin B for filamentous fungal keratitis. DESIGN: Outcome-masked, 2×2 factorial design, randomized controlled clinical trial. PARTICIPANTS: Consecutive patients presenting with moderate vision loss from a smear-positive fungal ulcer at Aravind Eye Hospital, Madurai, India. METHODS: Study eyes were randomized to 1 of 4 treatment combinations using an adaptive randomization protocol. The treatment arms included (1) topical natamycin 5% alone, (2) topical natamycin 5% plus CXL, (3) topical amphotericin B 0.15% alone, and (4) topical amphotericin 0.15% plus CXL. MAIN OUTCOME MEASURES: The primary outcome of the trial was microbiological cure at 24 hours on repeat culture. Secondary outcomes included best spectacle-corrected visual acuity (BSCVA) at 3 weeks and 3 months, percentage of study participants with epithelial healing at 3 days, 3 weeks, and 3 months, infiltrate or scar size at 3 weeks and 3 months, 3-day smear and culture, and adverse events. RESULTS: Those randomized to CXL regardless of medication (topical natamycin or amphotericin) had 1.32-fold increased odds of 24-hour culture positivity, although this was not statistically significant (95% confidence interval [CI], 0.57-3.06; P = 0.51). We were also unable to find a difference in 24-hour culture positivity between those randomized to amphotericin and those randomized to natamycin when evaluating as a group regardless of whether or not they received CXL (coefficient 1.10; 95% CI, 0.47-2.54; P = 0.84). The BSCVA was approximately 0.22 logarithm of the minimum angle of resolution (logMAR) (2.2 Snellen lines) worse on average at 3 weeks among those receiving CXL regardless of medication (95% CI, -0.04 to 0.40; P = 0.04) and 0.32 logMAR (3.2 Snellen lines) worse visual acuity at 3 months after controlling for baseline visual acuity (95% CI, 0.03-0.54; P = 0.02). There was no difference in infiltrate or scar size, percentage of epithelialized or adverse events when comparing CXL with no CXL or the 2 topical medications. CONCLUSIONS: There appears to be no benefit of adjuvant CXL in the primary treatment of moderate filamentous fungal ulcers, and it may result in decreased visual acuity.

Natamycin treatment reduces the quality changes of postharvest mulberry fruit during storage.

This study investigated the effect of natamycin (NA) treatment on the quality of mulberry fruit throughout the storage at 8°C and 90% relative humidity. The results of this study showed that NA treatment reduced the decay rate of mulberry fruit, inhibited respiration, decreased malondialdehyde (MDA) content, decreased phenylalanine ammonia-lyase (PAL) and polyphenoloxidase (PPO) activity, and maintained sucrose levels, total soluble solids (TSS), and total acids (TA) contents in the fruits throughout storage. NA treatment maintained the surface color and firmness of fruits throughout storage and suppressed the production of fructose, glucose, total phenolics, and the contents of anthocyanin. NA treatment maintained the quality of mulberry fruit throughout storage by enhancing the enzymatic activity of the reactive oxygen species (ROS) scavengers: catalase (CAT), superoxide dismutase (SOD), and peroxidase (POD) and inhibiting the production of hydrogen peroxide (H2 O2 ). Treatment of mulberries with increasing concentrations of NA could significantly inhibit the growth of fungal pathogens in mulberry fruits and improve their quality. PRACTICAL APPLICATIONS: The results from our previous study showed that NA was a safe preservative which could effectively inhibit the growth of molds and yeasts in mulberry fruits and improve their quality throughout the postharvest storage. Compared with other treatments for mulberry fruit, NA treatment has the advantages of low cost, simple operation, good application, and extension. The results in this study further prove that application of NA treatment has potential in the food industry to maintain the postharvest quality and extend the shelf life of mulberry fruits throughout storage.

Natamycin solid lipid nanoparticles - sustained ocular delivery system of higher corneal penetration against deep fungal keratitis: preparation and optimization.

BACKGROUND: Fungal keratitis (FK) is a serious pathogenic condition usually associated with significant ocular morbidity. Natamycin (NAT) is the first-line and only medication approved by the Food and Drug Administration for the treatment of FK. However, NAT suffers from poor corneal penetration, which limits its efficacy for treating deep keratitis. PURPOSE: The objective of this work was to prepare NAT solid lipid nanoparticles (NAT-SLNs) to achieve sustained drug release and increased corneal penetration. METHODS: NAT-SLNs were prepared using the emulsification-ultrasonication technique. Box- Behnken experimental design was applied to optimize the effects of independent processing variables (lipid concentration [X1], surfactant concentration [X2], and sonication frequency [X3]) on particle size (R1), zeta potential (ZP; R2), and drug entrapment efficiency (EE%) (R3) as responses. Drug release profile, ex vivo corneal permeation, antifungal susceptibility, and cytotoxicity of the optimized formula were evaluated. RESULTS: The optimized formula had a mean particle size of 42 r.nm (radius in nanometers), ZP of 26 mV, and EE% reached ~85%. NAT-SLNs showed an extended drug release profile of 10 hours, with enhanced corneal permeation in which the apparent permeability coefficient (Papp) and steady-state flux (Jss) reached 11.59×10-2 cm h-1 and 3.94 mol h-1, respectively, in comparison with 7.28×10-2 cm h-1 and 2.48 mol h-1 for the unformulated drug, respectively. Antifungal activity was significantly improved, as indicated by increases in the inhibition zone of 8 and 6 mm against Aspergillus fumigatus ATCC 1022 and a Candida albicans clinical isolate, respectively, and minimum inhibitory concentration values that were decreased 2.5-times against both of these pathogenic strains. NAT-SLNs were found to be non-irritating to corneal tissue. NAT-SLNs had a prolonged drug release rate, that improved corneal penetration, and increased antifungal activity without cytotoxic effects on corneal tissues. CONCLUSION: Thus, NAT-SLNs represent a promising ocular delivery system for treatment of deep corneal keratitis.

Colonization of therapeutic contact lens by dematiaceous fungi.

PURPOSE: To report 2 patients with colonization of therapeutic contact lens with dematiaceous fungi. METHODS: Case report. RESULTS: The first patient had a retained soft contact lens on an opaque cornea for 4 years with brownish black multiple colonies on the soft contact lens and culture grew Bipolaris spp. The second patient was on therapeutic contact lens for pseudophakic bullous keratopathy for 4 months and developed a brownish colonization of contact lens with unidentified dematiaceous fungi. Both the patients had conjunctivitis but did not develop fungal keratitis. CONCLUSION: Judicious use of therapeutic contact lens is required in agrarian countries with adequate emphasis on strict adherence to the standard protocols and frequent replacement of the lens.

Fungal Keratitis: Epidemiology, Rapid Detection, and Antifungal Susceptibilities of Fusarium and Aspergillus Isolates from Corneal Scrapings.

Fungal aetiology of keratitis/corneal ulcer is considered to be one of the leading causes of ocular morbidity, particularly in developing countries including India. More importantly, Fusarium and Aspergillus are reported commonly implicating corneal ulcer and against this background the present work was undertaken so as to understand the current epidemiological trend of the two fungal keratitis. During the project period, a total of 500 corneal scrapings were collected from suspected mycotic keratitis patients, of which 411 (82.2%) were culture positive for bacteria, fungi, and parasites. Among fungal aetiologies, Fusarium (216, 52.5% of 411) and Aspergillus (68, 16.5% of 411) were predominantly determined. While the study revealed a male preponderance with both the fungal keratitis , it further brought out that polyene compounds (natamycin and amphotericin B) and azoles were active, respectively, against Fusarium spp. and Aspergillus spp. Additionally, 94.1% of culture proven Fusarium keratitis and, respectively, 100% and 63.6% of A. flavus and A. fumigatus were confirmed by multiplex PCR. The sensitivity of the PCR employed in the present study was noted to be 10 fg/µl, 1 pg/µl, and 300 pg/µl of DNA, respectively, for Fusarium, A. flavus, and A. fumigatus. Alarming fact was that Fusarium and Aspergillus regionally remained to be the common cause of mycotic keratitis and the Fusarium isolates had a higher antifungal resistance than Aspergillus strains against most of the test drugs.

Natamycin niosomes as a promising ocular nanosized delivery system with ketorolac tromethamine for dual effects for treatment of candida rabbit keratitis; in vitro/in vivo and histopathological studies.

OBJECTIVES: This study was aimed to develop dual-purpose natamycin (NAT)-loaded niosomes in ketorolac tromethamine (KT) gels topical ocular drug delivery system to improve the clinical efficacy of natamycin through enhancing its penetration through corneal tissue and reducing inflammation associated with Fungal keratitis (FK). SIGNIFICANCE: Nanosized carrier systems, as niosomes would provide great potential for improving NAT ocular bioavailability.NAT niosomal dispersion formulae were prepared and then incorporated in 0.5%KT gels using different mucoadhesive viscosifying polymers. METHODS: Niosomes were prepared using the reverse-phase evaporation technique. In vitro experimental, and in vivo clinical evaluations for these formulations were done for assessment of their safety and efficacy for treatment of Candida Keratitis in Rabbits. In vitro release study was carried out by the dialysis method. In vivo and histopathological studies were performed on albino rabbits. RESULTS: NAT niosomes exhibited high entrapment efficiency percentage (E.E%) up to96.43% and particle size diameter ranging from 181.75 ± 0.64 to 498.95 ± 0.64 nm, with negatively charged zeta potential (ZP). NAT niosomal dispersion exhibited prolonged in vitro drug release (40.96-77.49% over 24h). NAT-loaded niosomes/0.5%KT gel formulae revealed retardation in vitro release, compared to marketed-product (NATACYN®) and NAT-loaded niosomes up to57.32% (F8). In vivo experimental studies showed the superiority for F8 in treatment of candida keratitis and better results on corneal infiltration and hypopyon level. These results were consistent with histopathological examination in comparison with F5 and combined marketed products (NATACYN® and Ketoroline®). CONCLUSIONS: This study showed that F8 has the best results from all pharmaceutical in vitro evaluations and a better cure percent in experimental application and enhancing the prolonged delivery of NAT and penetrating the cornea tissues.

Identification and Characterization of Fusarium proliferatum, a New Species of Fungi that Cause Fungal Keratitis.

Fusarium proliferatum (F. proliferatum) is known as a pathogen of corn and other crops, but its role in fungal keratitis has not been well investigated. Among 877 Fusarium isolates, we identified 155 (17.7%) stains as F. proliferatum according to their morphological features and partial DNA sequencing of translation elongation factor-[Formula: see text] (EF-[Formula: see text]) in this study. In vitro antifungal susceptibility tests showed that the F. proliferatum strains were sensitive to natamycin and vorionazole but resistant to amphotericin B, fluconazol, ketoconazole and itaconazole. Most of the F. proliferatum-positive keratitis patients (44/155,28.4%) were aged 51-60 years old. The main cause of infection was injury by a plant (51/155, 32.9%). A combination of 1% amphotericin B and 3% ketoconazole cured 45.2% (14/31) and a combination of 0.5% natamycin and 0.5% voriconazole cured 59.1% (13/22) of F. proliferatum-positive patients. The date suggests that F. proliferatum identified through EF-1ɑ DNA sequencing is an important new species that causes fungal keratitis. Based on antifungal susceptibility, treatment with a combination of 0.5% natamycin and 0.5% voriconazole improves the therapeutic efficacy in F. prolifertum-positive patients.

Necessity to identify the causative agent for appropriate treatment in fungal corneal ulcer: an in vitro study.

OBJECTIVE: Fungal corneal ulcers are a major cause of preventable blindness. Different antifungal agents as natamycin, nystatin, fluconazole, itraconazole, voriconazole are used to treat these ulcers. Among these, natamycin is most widely used as a treatment modality. In natamycin non-responding cases, other drugs especially voriconazole is used. This study was done to assess the use of antifungal drugs in the treatment of fungal corneal ulcer by determining the minimum inhibitory concentration against common fungal pathogens. MATERIAL AND METHODS: Minimum inhibitory concentration of fluconazole, itraconazole, voriconazole, nystatin and natamycin was assessed against the 61 isolated corneal fungal pathogens as per CLSI guidelines. RESULTS: MIC value of different antifungal agents varies as per fungal strains. Voriconazole showed the lowest MIC against the isolated fungi, in comparison to fluconazole and itraconazole. In comparison to other fungi, higher natamycin MIC was observed against Aspergillus species. Itraconazole is poorly effective against Fusarium sp. CONCLUSION: Identification of causative fungi is necessary before antifungal treatment. Lowest voriconazole MIC promotes its use as 1st line drug. Comparative higher natamycin MIC, especially against Aspergillus species, warns clinician to have MIC in each case of a non-responding fungal corneal ulcer.

Antifungal Effect of Essential Oils against Fusarium Keratitis Isolates.

The present study was carried out to investigate the antifungal effects of Cinnamomum zeylanicum, Citrus limon, Juniperus communis, Eucalyptus citriodora, Gaultheria procumbens, Melaleuca alternifolia, Origanum majorana, Salvia sclarea, and Thymus vulgaris essential oils against Fusarium species, the most common etiologic agents of filamentous fungal keratitis in South India. C. zeylanicum essential oil showed strong anti-Fusarium activity, whereas all the other tested essential oils proved to be less effective. The main component of C. zeylanicum essential oil, trans-cinnamaldehyde, was also tested and showed a similar effect as the oil. The in vitro interaction between trans-cinnamaldehyde and natamycin, the first-line therapeutic agent of Fusarium keratitis, was also investigated; an enhanced fungal growth inhibition was observed when these agents were applied in combination. Light and fluorescent microscopic observations revealed that C. zeylanicum essential oil/trans-cinnamaldehyde reduces the cellular metabolism and inhibits the conidia germination. Furthermore, necrotic events were significantly more frequent in the presence of these two compounds. According to our results, C. zeylanicum essential oil/trans-cinnamaldehyde provides a promising basis to develop a novel strategy for the treatment of Fusarium keratitis.

Corneal Cross-linking as an Adjuvant Therapy in the Management of Recalcitrant Deep Stromal Fungal Keratitis: A Randomized Trial.

PURPOSE: To assess the efficacy of corneal cross-linking (CXL) as an adjuvant to appropriate antifungal therapy in nonresolving deep stromal fungal keratitis. DESIGN: Randomized clinical trial. METHODS: Eyes with culture-positive deep stromal fungal keratitis not responding to appropriate medical therapy for a period of 2 weeks were randomized to receive either adjuvant CXL or no additional treatment. Antifungal medical therapy was continued in both groups. The prespecified primary outcome was treatment failure at 6 weeks after enrollment, defined as perforation and/or increase in ulcer size by ≥2 mm. RESULTS: The trial was stopped before full enrollment because of a marked difference in the rate of perforation between the 2 groups. Of the 13 cases enrolled in the study, 6 were randomized to the CXL group and 7 to the non-CXL group. Five eyes in the CXL group and 3 eyes in the non-CXL group experienced treatment failure by 6 weeks (P = .56). In a secondary analysis, the CXL group experienced more perforations than the non-CXL group (4 vs 0, respectively; P = .02). CONCLUSION: CXL used as adjuvant therapy for recalcitrant deep stromal fungal keratitis did not improve outcomes.

Aspergillus terreus infection in a sutureless self-sealing incision made during cataract surgery.

Here, we describe a case of keratitis caused by Aspergillus terreus in an 80-year-old immunocompetent woman 1 month after uneventful cataract surgery. The patient presented with decreased visual acuity (20/50) and severe pain in her right eye. Examination revealed a 3.5 × 4.5 mm white-colored deep stromal infiltration located on the temporal corneal tunnel incision. Corneal scraping samples were obtained for cytological and culture examinations. The cinnamon-brown colonies consisting of a dense felt of conidiophores were identified as A. terreus using molecular data. Using CLSI M38-A2 microdilution method, minimum inhibitory concentration values of amphotericin B, itraconazole, voriconazole, and posaconazole were determined to be 2, 1, 0.25, and 1 µg/ml, respectively, and minimum effective concentration values of caspofungin and anidulafungin were ≤0.03 and ≤0.03, respectively, at 48 h for the A. terreus strain. Antifungal therapy was started as topical 1 % voriconazole drops hourly and 5 % natamycin ointment five times a day; however, corneal infection and melting progressed despite the ongoing intensive treatment and visual acuity dropped to the 20/200 level at the end of the first week. Amniotic membrane transplantation was performed to promote corneal healing. Topical medication was tapered and discontinued within 2 months based on the clinical features. The ulcer healed with scarring and vascularization, and visual acuity improved to 20/50. In conclusion, A. terreus is a very uncommon cause of mycotic keratitis and is especially rare after cataract surgery. We suggest that early and accurate diagnosis and appropriate treatment of A. terreus keratitis may have a major impact on preventing severe disease complications.

A Case of Alternaria alternata Keratitis Isolated from Corneal Tissue / 대한의진균학회지

A 71-year-old man presented with pain in the left eye that revealed a 3x3 mm deep corneal stromal infiltrate, with a 2x2 mm epithelial defect. The patient started topical moxifloxacin, voriconazole 2%, and natamycin for 2 weeks. However, the treatment was not effective and the corneal infiltration worsened. Subsequently, the patient underwent therapeutic penetrating keratoplasty. Thick brown/gray mold colonies on Potato Corn Meal Tween 80 agar was isolated from excised corneal tissue and on slide culture many septated, and club-shaped ascospores were revealed. Histological findings also showed numerous hyphae scattered in corneal tissue. A. alternata colonies were confirmed by 18S rRNA sequencing. Intracameral voriconazole was injected every other day for 2 weeks to eliminate remaining fungi on the deep corneal stroma. The remaining corneal infiltration was improved one month after the injection. During 5 months postoperative follow up, the infection did not recurred. In conclusion, deep corneal infection of A. alternata was effectively treated with intracameral voriconazole injection.