Evaluation of antifungal susceptibility and clinical characteristics in fungal keratitis in a tertiary care center in North India

Purpose: To study the antifungal susceptibility of common corneal pathogenic fungi to antifungal agents in the North Indian population. Methods: Prospective study of the antifungal sensitivity testing (natamycin, amphotericin B, voriconazole, itraconazole, fluconazole, posaconazole, caspofungin, micafungin) of fungal isolates from 50 cases of culture positive fungal keratitis by using E test method. Details noted included demographic data, visual acuity, clinical details, grade of keratitis, healing time, and success in medical management. Results: Of 50 patients with fungal keratitis (mean age: 40.28 ± 16.77 years), 12 eyes healed within 3 weeks, 14 had a delayed healing response, and 24 had chronic keratitis. Among the 15 cases of Fusarium isolates, 93.3% were sensitive to natamycin, while 40% to amphotericin B; 66.6% to voriconazole, 13.4% to itraconazole and fluconazole each. 80% of Fusarium cases (n = 12) showed susceptibility to posaconazole. Among Aspergillus flavus isolates, 53.4% (n = 8) were sensitive to natamycin, with only 40% (n = 7) showing sensitivity to amphotericin B and good susceptibility to azoles. MIC against susceptible Fusarium spp. for natamycin was 3–16 ?g/mL, amphotericin B: 1–8 ?g/mL, voriconazole: 0.5–1.5 ?g/ mL, itraconazole: 0.5–12 ?g/mL, posaconazole: 0.094–1.5 ?g/mL. MIC against Aspergillus flavus was natamycin: 8–32 ?g/mL, amphotericin B: 0.5–16 ?g/mL, voriconazole: 0.025–4 ?g/mL, itraconazole: 0.125–8 ?g/mL, posaconazole: 0.047–0.25 ?g/mL; against Aspergillus niger isolates, to natamycin was 6 ?g/mL (n=1), amphotericin B 8–12 ?g/mL (n = 3), voriconazole: 0.125–0.19 ?g/mL (n = 3), itraconazole: 0.38–0.75 ?g/mL, posaconazole: 0.064–0.19 ?g/mL and against Aspergillus fumigatus (n = 1), was natamycin4 ?g/ mL, amphotericin B ? 8 ?g/mL, voriconazole 0.25 ?g/mL, itraconazole 1 ?g/mL, and posaconazole 0.19 ?g/mL. MIC against susceptible Acremonium spp. for natamycin was 1.5–16 ?g/mL, amphotericin B: 0.5–8 ?g/mL, voriconazole: 0.19–3 ?g/mL, itraconazole: 0.125 ?g/mL, posaconazole: 0.125–0.5 ?g/mL and against susceptible Curvularia was natamycin 0.75–4 ?g/mL, amphotericin B 0.5–1 ?g/mL, voriconazole 0.125–0.19 ?g/mL, itraconazole 0.047–0.094 ?g/mL, posaconazole 0.047–0.094 ?g/mL. MIC against Mucor spp.+ Rhizopus spp. (n = 1) was natamycin: 8 ?g/mL, amphotericin B: 0.75 ?g/mL, posaconazole: 1.5 ?g/ mL. MIC against of Alternaria (n = 1) was voriconazole: 0.19 ?g/mL, posaconazole: 0.094 ?g/mL. MIC against Penicillium (n=1) was natamycin: 8 ?g/mL, voriconazole: 0.25 ?g/mL, itraconazole: 0.5 ?g/mL, and Posaconazole: 0.125 ?g/mL. Conclusion: Our observations highlight the variations in susceptibility to antifungal agents. Posaconazole seems to be effective with low MIC against common corneal pathogenic fungal isolates

Engineering the precursor supply pathway in Streptomyces gilvosporeus for overproduction of natamycin / 生物工程学报

Natamycin is a safe and efficient antimycotics which is widely used in food and medicine industry. The polyene macrolide compound, produced by several bacterial species of the genus Streptomyces, is synthesized by type Ⅰ polyketide synthases using acetyl-CoA, malonyl-CoA, and methylmalonyl-CoA as substrates. In this study, four pathways potentially responsible for the supply of the three precursors were evaluated to identify the effective precursor supply pathway which can support the overproduction of natamycin in Streptomyces gilvosporeus, a natamycin-producing wild-type strain. The results showed that over-expressing acetyl-CoA synthetase and methylmalonyl-CoA mutase increased the yield of natamycin by 44.19% and 20.51%, respectively, compared with the wild type strain under shake flask fermentation. Moreover, the yield of natamycin was increased by 66.29% compared with the wild-type strain by co-overexpression of acetyl-CoA synthetase and methylmalonyl-CoA mutase. The above findings will facilitate natamycin strain improvement as well as development of strains for producing other polyketide compounds.

Advances in the biosynthesis of natamycin and its regulatory mechanisms / 生物工程学报

Natamycin is a polyene macrolide antibiotics with strong and broad spectrum antifungal activity. It not only effectively inhibits the growth and reproduction of fungi, but also prevents the formation of some mycotoxins. Consequently, it has been approved for use as an antifungal food preservative in most countries, and is also widely used in agriculture and healthcare. Streptomyces natalensis and Streptomyces chatanoogensis are the main producers of natamycin. This review summarizes the biosynthesis and regulatory mechanism of natamycin, as well as the strategies for improving natamycin production. Moreover, the future perspectives on natamycin research are discussed.

A Novel and Rapid HPLC Method for Determination of Natamycin in Turkish Cheese Samples

Nicotine is a potential inducer of oxidative stress, which can damage the numerous tissues and biological molecules.Nicotine is first metabolized in liver, and the lung is the main target organ susceptible to smoking; therefore, the purpose ofthe current examination was scrutinizing the impacts of nicotine on these organs using the histological method, and studyingthe ameliorative (additive or synergistic) effects of quercetin and luteolin supplements in target tissues against nicotinetoxicity in young albino rats. Fifty animals were involved in this study which were divided into five groups of ten rats, theywere treated as: (1) untreated Control (Cont); (2) Nicotine (Ni) treated (0.75 mg/kg b.w./d, i.p.); (3) Nicotine as above withintragastric administration of quercetin (Ni+Qu) (50mg/kg b.w) or (4) with luteolin (50mg/kg b.w) in Group (Ni+Lu), and(5) nicotine with combination of quercetin and luteolin (Ni+Qu+Lu) as described above. All groups were treated for 8weeks, then their liver and lung tissues were dissected out. Several histological damages were noticed in (Ni) rats such asdegeneration changes in hepatocytes, slight congestion of central vein (CV) and cellular infiltration. Nicotine caused drasticchanges in lung tissues such as, inflammatory cells aggregation inside and outside the bronchiol artery with mild increase inthe thickness of bronchial muscular wall and marked thickening in the alveolar wall. All supplemented groups amelioratedthe damage that induced by nicotine injection in both liver and lung tissues. In liver tissues, the quercetin (Ni+Qu)supplemented rats showed the better improvement nearby normal hepatocytes than the other groups, while in lung tissues,the luteolin supplemented rats had better recovery than the other treatments. These observations suggested that the intake ofquercetin and luteolin as supplements may be useful in combating tissue injury that is a result of nicotine toxicity

Antifungal effect of combined application of corneal collagen cross-linking and natamycin / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To Study the anti-fungal effect of corneal collagen cross-linking combined with natamycin <i>in vivo</i> and <i>in vitro</i>, so as to provide the treatment and experimental basis for the treatment of clinical fungal keratitis. <p>METHODS: Three common pathogenic fungi(<i>Aspergillus flavus</i>, <i>Fusarium Solani</i> and <i>Candida albicans</i>)were used. The experimental group was divided into cross-linking combined natamycin group, natamycin combined riboflavin group, natamycin combined UVA irradiation group, cross-linking group and natamycin group as the control group. The drug was added to the center of the Sabouraud dextrose agar(SDA)plate coated with liquid with each fungal spores with the same maid turbidity of 1.5. Ten minutes later, it was irradiated with collagen cross-linking instrument for 10min and cultured at 28℃ for 36h, and then the inhibition zone size was measured and analyzed statistically. The rabbit model of <i>Fusarium Solani</i> corneal infection was prepared. The model rabbits were randomly divided into model control group, cross-linking treatment group, natamycin treatment group, cross-linking combined natamycin group, 5 rabbits in each group. And another 5 normal rabbits were taken as control, and five rabbits were irradiated in accordance with corneal collagen cross-linking therapy. The results were observed by anterior segment photography, corneal scraping and confocal microscopy, and the ultra micro structural changes of the corneas were observed by electron microscope after the treatment.<p>RESULTS: Corneal collagen cross-linking alone had shown no effect on each fungus <i>in vitro</i>. Corneal collagen cross-linking combined with natamycin produced significant anti-fungal effect(<i>P</i><0.05). However, the anti-fungal effect of natamycin combined riboflavin group and natamycin combined ultraviolet light group showed no statistical difference(<i>P</i>>0.05)comparing with the control group. For the model of rabbit fungal infection, the course of disease was about 14d in the natamycin group and CXL combined with natamycin group, and it was about 21d in CXL group. After the treatment, all the groups healed. There were no defects in the corneal epithelium, no mycelium in the corneas, except for more corneal neovascularization. The results of the anterior segment photography showed that the treatment effect of the cross-linking combined natamycin group was better than other groups, with fewer scar tissue, better corneal healing and relatively short course of disease.<p>CONCLUSION: Corneal collagen cross-linking combined with natamycin treatment is able to enhance anti-fungal effect, promote corneal healing, and shorten the course of disease. So it is a promising therapeutic technique for the clinical treatment of fungal keratitis.

Natamicina versus voriconazol para queratitis fúngica / Natamycin versus voriconazole for fungal keratitis

INTRODUCCIÓN: La queratitis infecciosa de origen fúngico afecta principalmente a personas de países tropicales y subtropicales, y constituye una importante causa de ceguera prevenible. Los antifúngicos tópicos, en particular la natamicina y el voriconazol, se consideran efectivos, pero no está claro cuál de ellos constituye la mejor alternativa de tratamiento. MÉTODOS: Realizamos una búsqueda en Epistemonikos, la mayor base de datos de revisiones sistemáticas en salud, la cual es mantenida mediante el cribado de múltiples fuentes de información, incluyendo MEDLINE, EMBASE, Cochrane, entre otras. Extrajimos los datos desde las revisiones identificadas, analizamos los datos de los estudios primarios, realizamos un metanálisis y preparamos una tabla de resumen de los resultados utilizando el método GRADE. RESULTADOS Y CONCLUSIONES: Identificamos tres revisiones sistemáticas que en conjunto incluyeron tres estudios primarios, todos correspondientes a ensayos aleatorizados. Concluimos que natamicina probablemente se asocia a mejor agudeza visual tras la infección, y que previene la perforación corneal y/o la necesidad de realizar queratoplastia terapéutica en comparación a voriconazol en queratitis fúngica.

INTRODUCTION: Infectious keratitis of fungal origin mainly affects people in tropical and subtropical countries, and is an important cause of preventable blindness. Topical antifungals, particularly natamycin and voriconazole, are considered effective, but it is not clear which one is the best treatment alternative. METHODS: We searched in Epistemonikos, the largest database of systematic reviews in health, which is maintained by screening multiple information sources, including MEDLINE, EMBASE, Cochrane, among others. We extracted data from the systematic reviews, reanalyzed data of primary studies, conducted a meta-analysis and generated a summary of findings table using the GRADE approach. RESULTS AND CONCLUSIONS: We identified three systematic reviews including three studies overall,all of which were randomized trials. We concluded natamycin probably is associated with better visual acuity after infection, and it prevents corneal perforation and/or need to perform therapeutic keratoplasty compared to voriconazole in fungal keratitis.

In Vitro Assessment of Antileishmanial Activity of Natamycin and Nystatin

ABSTRACT The present study was aimed to evaluate the in vitro antileishmanial activity of four different concentrations of natamycin and nystatin by using MTT 3-(4.5-dimethylthiazol-2-yl)-2.5-diphenyl tetrazolium bromide reduction assay. In vitro antileishmanial activity revealed that the IC50 of natamycin (80.49 μg/ml) and nystatin (105.7 μg/ml) was less than that of sodium stibogluconate (127.9 μg/ml), and more than amphotericin B (18.91 μg/ml).

Natamycin in the treatment of keratomycosis: Correlation of treatment outcome and in vitro susceptibility of fungal isolates.

In this study, 60 fungal isolates from 60 patients with fungal keratitis were tested in vitro for their susceptibility to natamycin and the mean minimum inhibitory concentrations of natamycin (MICn) were correlated with clinical outcome. The mean MICn for various groups of fungi from patients with either early (<10 days) or late (≥10 days) presentation was correlated with the outcome. Aspergillus flavus showed resistance to natamycin with a high mean MICn (>16 μg/ml). While the clinical response in all patients with early A. flavus keratitis was good it was poor in late cases (5/8 patients, 62.5%). Fusarium species, Acremonium species and dematiaceous fungi were sensitive with low mean MICn (Fusarium: 5.7-7.2 μg/ml, Acremonium: 5.7-6.8 μg/ml, dematiaceous: (1.6-4 μg/ml). However, 46.6% (7/15) patients in Fusarium and 57.1% (4/7) in Acremonium group needed keratoplasty. We conclude that despite susceptibility of most fungal species causing keratitis to natamycin, the treatment outcome is poor in advanced fungal keratitis.

A Case of Fungal Keratitis Caused by Drechslera Species

PURPOSE: To report a case of fungal keratitis caused by Drechslera species. CASE SUMMARY: A 77-year-old man visited our clinic complaining of left ocular pain and decrease of visual acuity for 1 week On slit-lamp examination, epithelial defect and stromal infiltration on the corneal center with numerous inflammatory cells in the anterior chamber were found. There was no improvement after routine antibiotic treatment. A corneal biopsy and culture were performed on the corneal lesion. The KOH smear study reported hyphae, thus the patient was treated with 0.25% amphotericin B, 0.2% fluconazole and 5% natamycin eye drops. A clinical improvement was observed on the corneal lesion and Drechslera species was identified by the culture study. CONCLUSIONS: The authors report an experience of fungal keratitis caused by Drechslera species.

Fungal keratitis in Lattice dystrophy.

We report a case of fungal keratitis occurring in a patient with lattice dystrophy. A 57-year-old farmer presented with a corneal ulcer following probable entry of paddy husk in the right eye, of one month duration. Corneal scraping revealed pigmented fungal filaments while culture grew Alternaria alternata. Treatment with 5% natamycin eye drops and 1% atropine healed the infection in four weeks. We would like to draw attention to the fact that the cornea in lattice dystrophy is prone to frequent erosions and is a compromised epithelial barrier to invasion by microorganisms. Patients must be made aware of this fact and should seek attention at the earliest following any trivial trauma. Management of minor corneal abrasions in them should be directed at healing the epithelium with adequate lubricants and preventing infection with topical antibiotic prophylaxis.

Identification of an antifungal metabolite produced by a potential biocontrol Actinomyces strain A01 / Identificação de um metabólito antifúngico produzido pela cepa Actinomyces A01

Actinomyces strain A01 was isolated from soil of a vegetable field in the suburb of Beijing, China. According to the morphological, cultural, physiological and biochemical characteristics, and 16S rDNA sequence analysis, strain A01 was identified as Streptomyces lydicus. In the antimicrobial spectrum test strain A01 presented a stable and strong inhibitory activity against several plant pathogenic fungi such as Fusarium oxysporum, Botrytis cinerea, Monilinia laxa, etc. However, no antibacterial activity was found. In pot experiments in greenhouse, the development of tomato gray mold was markedly suppressed by treatment with the fermentation broth of the strain A01, and the control efficacy was higher than those of Pyrimethanil and Polyoxin. A main antifungal compound (purity 99.503 percent) was obtained from the fermentation broth of strain A01 using column chromatography and HPLC. The chemical structural analysis with UV, IR, MS, and NMR confirmed that the compound produced by the strain A01 is natamycin, a polyene antibiotic produced by S. chattanovgensis, S. natalensis, and S. gilvosporeus, widely used as a natural biological preservative for food according to previous reports. The present study revealed a new producing strain of natamycin and its potential application as a biological control agent for fungal plant diseases.

A cepa Actinomyces A01 foi isolada do solo de um campo agrícola no subúrbio de Beijing, China. De acordo com as características morfológicas, culturais, fisiológicas e bioquímicas, e análise da sequência 16S rDNA , a cepa A01 foi identificada como Streptomyces lydicus. Nos testes de espectro antimicrobiano, a cepa A01 apresentou atividade inibitória intensa e estável contra vários fungos patogênicos para plantas, como Fusarium oxysporum, Botrytis cinerea, Monilia laxa, etc. Entretanto, não foi encontrada atividade antibacteriana. Em experimentos em estufas, o desenvolvimento do fungo cinza do tomate foi fortemente inibido pelo tratamento com o caldo de fermentação da cepa A01, com eficiência superior à do pyremethanil e polyoxin. Por cromatografia em coluna e HPLC, obteve-se um composto fúngico (pureza 99,503 por cento), cuja análise estrutural por UV, IR, MS e NMR revelou ser natamicina, um antibiótico polienico produzido por S. chattanovgensis, S. natalensis e S.gilvosporeus, empregado como conservador biológico natural em alimentos. O presente estudo relata a detecção de uma nova cepa produtora de natamicina e sua aplicação potencial como um agente de controle biológico de doenças fúngicas em plantas.

Effect of topical 0.5 percent povidone-iodine compared to 5 percent natamycin in fungal keratitis caused by Fusarium solani in a rabbit model: a pilot study / Efeito tópico de iodo-povidona 0, 5 por cento comparado com natamicina 5 por cento em ceratites fúngicas causadas por Fusarium solani em um modelo animal de coelhos: estudo piloto

PURPOSE: To evaluate the efficacy of topical administration of 0.5 percent povidone-iodine in experimental Fusarium solani keratitis in rabbits. METHODS: Fungal keratitis caused by Fusarium solani was induced in the right eye of 24 New Zealand rabbits. The rabbits were randomly divided into 3 different treatment groups: Group I (povidone-iodine) - treated with topical 0.5 percent povidone-iodine; Group II (natamycin) - treated with topical 5 percent natamycin; and Group III (control) - treated with topical saline solution. In all groups the rabbits were treated for three days and then sacrificed. The corneas were excised, macerated and immersed in 10 mL BHI. Culture samples were plated daily on Sabouraud's agar for 7 days, and the number of colony-forming units (CFU) was counted. The rabbits were clinically evaluated during the treatment period. RESULTS: The povidone-iodine and natamycin groups demonstrated better efficacy than the control group based on the number of rabbits with no colonies growing. However, there were no statistically significant differences between the three groups when the number of CFU was analyzed (p>0.05). CONCLUSIONS: Our study demonstrates important methodological considerations in the use of in vivo animal models for the testing of antifungal agents. Using this sample size and methodology of counting CFU, topical 0.5 percent povidone-iodine demonstrated no benefit in the treatment of experimental Fusarium solani when compared with topical 5 percent natamycin.

OBJETIVO: Avaliar a eficácia do uso tópico de iodo-povidona 0,5 por cento em ceratite experimental por Fusarium solani em coelhos. MÉTODOS: Ceratite fúngica por Fusarium solani foi induzida no olho direito de 24 coelhos da raça New Zealand. Os coelhos foram divididos aleatoriamente em 3 diferentes grupos de tratamento: Grupo I (iodo-povidona) - tratados com iodo-povidona 0,5 por cento; Grupo II (natamicina) - tratados com natamicina 5 por cento; Grupo III (controle) - tratados com solução salina. Os coelhos dos 3 grupos foram tratados por 3 dias e sacrificados em seguida. As córneas foram removidas cirurgicamente, maceradas e incubadas em meio BHI. Semeou-se culturas em placas de ágar Sabouraud, diariamente, durante 7 dias, e contou-se o número de unidades formadoras de colônias (UFC). Os coelhos foram avaliados clinicamente durante o período de tratamento. RESULTADOS: Os grupos iodo-povidona e natamicina demonstraram melhor eficácia do que o grupo controle considerando-se o número de coelhos nos quais não houve crescimento de colônias. Entretanto, não houve diferença estatística significante entre os 3 grupos quando se analizou o número de UFC (p>0,05). CONCLUSÃO: Este estudo demonstrou considerações metodológicas importantes na utilização de modelos animais para o teste de agentes antifúngicos. Usando a metodologia de contar UFC e com este tamanho amostral, administração tópica de iodo-povidona 0,5 por cento não demonstrou benefício do tratamento de ceratite fúngica experimental causada por Fusarium solani quando comparado com a administração tópica de natamicina 5 por cento.

A Case of Fungal Keratitis as a Complication of Orthokeratology Contact Lens

PURPOSE: To present a case report of fungal keratitis related to prolonged overnight use of orthokeratology contact lenses. METHODS: A 13 year-old girl presented with a corneal ulcer in her left eye refractory to antibacterial medication. She had a history of wearing orthokeratology contact lenses overnight for seven months. RESULTS: The organism Aspergillus was isolated by corneal scraping, the contact lens itself, and from the storage case. The patient was treated with topical fluconazole and Natamycin pimaricin in addition to oral itraconazole, resulting in a resolution of the ocular lesion. CONCLUSIONS: The risk of fungal infection as a potential complication of the use of overnight orthkeratology contact lenses should be considered when using these lenses.

Toxicity of Topical Antifungal Agents on Corneal Epithelium of Rabbits

Though topical administration of antifungal agents for fungal keratitis is effective, it results various toxic effect on cornea depending on the kinds of agents. So, the authors applied several kinds of antifungal agents on the cornea of rabbit and made a comparative study for corneal epithelial damage of which is related to each agents and instillation time using scanning electron microscope. Forty eyes from twenty rabbits were instilled with 0.15% amphotericin B, 5% natamycin, 1% clotrimazole, 0.2% fluconazole and BSS(as a control) in single drop in each eight eyes and then each two corneas resected and examined after 30 minutes, 1 hours, 3 hours and 6 hours. All antifungals showed toxic effects on the corneal epithelium. Significant loss, elongation or swelling in microvilli, especially at the cell periphery, were noted. The top layer of epithelial cells showed moth-eaten appearance or punched-out lesions, and localized destruction. The surface cells were shrunken with wrinkling of plasma membrane and bulging of nucleous. Part of surface cells lost intercellular attachment leaving retraction fibrils. Rarely first and second layer cells of the epithelium were destructed, exposing third layer cells. Dark cells and premature desquamating cells increased significantly with the lapse of time. In general, 5% natamycin and 1% clotrimazole showed relatively severe epithelial toxicity, on the other hand, 0.15% amphotericin B and 0.2% fluconazole shoed mild epithelial toxicity. Localized damages in the epithelial cells were developed in the early stage, and an extent and the depth of the damage in the cells increased in the course of time after instillation.

PROGRESS IN MOLECULAR BIOLOGY OF NATAMYCIN / 微生物学通报

Natamycin is a polyene macrolide antifungal antibiotic ,which is wide ly used in the food industry in order to prevent mould contamination .Biosynthe s is gene cluster of natamycin is discovered by the overall of progress in molecul ar biology of natamycin, including 16 open reading frames which includes the gen e for 26-member ring formation of natamycin (pimS0-pimS4 ) and the modifying gene s, and the function of the protein including polyketide synthases(PKSs)、PimD、P imJ、PimK were studied

STREPTOMYCIN RESISTANCE MUTATION——A STUDY ON THE BREEDING OF HIGH NATAMYCIN-PRODUCING STRAIN / 微生物学通报

A method of streptomycin resistance screening was applied to improve t he productivity of Natamycin by Streptomyces gilvosporeus(ATCC13326) The sp ores treated with UV light were regenerated on agar plates containing 0 6?g/mL stre p tomycin 122 streptomycin resistant(str) mutants were obtained The Natamycin y iel ds of 13 mutants were higher than the original strain The mutants with high Na t amycin productivity were screened at a high frequency(10 6%) The highest one that demonstrated 1 46 times that of the original strain in Natamy cin productivity was obtained