Molecular identification and antifungal susceptibility of Curvularia australiensis,C. hawaiiensis and C. spicifera isolated from human eye infections

A reliable identification method was developed for three closely related Curvularia species, which are frequently isolated from human keratomycoses. Since the traditionally used morphological method and the increasingly used internal transcribed spacer (ITS)‐based molecular method proved to be insufficient to discern C. australiensis, C. hawaiiensis and C. spicifera, other molecular targets, such as β‐tubulin, translation elongation factor 1‐α and the nuclear ribosomal intergenic spacer (IGS), were tested. Among them, the use of the highly divergent IGS sequence is suggested and the species‐specific discriminating characters were determined in appropriate reference strains. It was also concluded that C. hawaiiensis and C. spicifera can be predominantly isolated from eye infections among the three species. The in vitro antifungal susceptibility of 10 currently used antifungal agents against 32 Curvularia isolates was also investigated. MICs were determined in each case. Isolates of C. spicifera proved to be less susceptible to the tested antifungals than those of C. hawaiiensis, which underline the importance of the correct identification of these species.     

Involvement of Fusarium spp. in fungal keratitis

Members of the filamentous fungal genus Fusarium are among the agents most frequently causing keratomycosis in humans. Fusarium keratitis is most common among agricultural workers in geographical regions with hot, humid, tropical or semi-tropical climates, but can occur more rarely in countries with temperate climates, such as Hungary. Keratitis is usually treated with a topical antifungal agent, sometimes in combination with sub-conjunctival injections and/or antimycotic agents, but therapeutic keratoplasty may be needed for patients whose corneal infection does not resolve. Early and accurate diagnosis, coupled with appropriate antifungal therapy, is crucial for improving the chances of complete recovery.     

EFFICACY OF MICONAZOLE IN EXPERIMENTAL KERATOMYCOSIS

For the first time comparative efficacy of 1 % miconazole in ointment base and drops has been evaluated against Candida, Aspergillus and Fusarium organisms In experimental heratomycosis. The findings of the present study reveal that miconazole in ointment drug delivery system has the advantage of being more effective in healing the fungal ulcers and associated lesions. It requires much less frequent application than drops, a parameter of great importance in improving patient compliance. Miconazole ointment was well tolerated by the rabbit eye and produced no ocular or systemic toxicity. The present study confirms that miconazole is a broad-spectrum antifungal agent and it could be a better alternative to presently available drugs to treat human heratomycosis .     

Immune Response and Mechanisms of IFN-γ in Administration for Keratomycosis

Purpose: To investigate the immune response and mechanisms of interferon-γ (IFN-γ) in the fungal keratitis in mice.

Methods: Mice were divided into two groups: group A, topical PBS four times daily post-infection; group B: topical IFN-γ four times daily post-infection. At1, 3, 5, and 7 days, the corneal lesions and inflammatory responses were observed by slit lamp, and immunofluorescence staining was performed to evaluate F4/80+ and CD4+ cells. Using ELISA, and RT–PCR to detect the expression levels of macrophage migration inhibitory factor (MIF), macrophage inflammatory protein-2 (MIP-2), IL-4, IL-10, IL-12, and IFN-γ.

Results: The treatment with IFN-γ decreased clinical scores and expression levels of IL-4, increased expression of F4/80+ and CD4+ cells, whereas IL-12, MIF, and MIP-2 were expressed highly, and the peaks of IL-10 and IFN-γ move forward.

Conclusion: This experiment showed that IFN-γ eye drops increase the accumulation of macrophages and shorten the duration of fungal keratitis.     

Antimycotic Activity of Different Chemicals, Chaksine Iodide and Garlic/Antimykotische Aktivität von verschiedenen Chemikalien, Chaksinjodit und Knoblauch

Summary: Thirteen different fungi reported to produce keratomycosis in man, were tested for their susceptibility to in vitro antifungal action of alum, sulphacetamide, copper sulphate, silver nitrate, chaksine iodide and garlic. Interestingly, 2% alum, 30% sodium sulphacetamide, 0.2% copper sulphate and 0.1% silver nitrate inhibited the growth of all 13 fungi viz. Alternaria, Aspergillus flavus, Asp. fumigatus, Asp. niger, Candida albicans, Curvularia, Drechslera, Fusarium, Mucor, Penicillium, Rhizopus oryzae, Scopulariopsis and Syncephalastrum. Chaksine iodide, an alkaloid from an Indian medicinal herb, Cassia absus, at a concentration of 0.5% inhibited all fungi. Lower concentrations were either ineffective or effective against a few fungi only. Garlic (up to 1%) could inhibit none of the fungi. Repeated experiments yielded consistent findings.
Zusammenfassung: 13 verschiedene Pilze, die als Erreger einer Keratomykose des Menschen bekannt sind, wurden in vitro gegen folgende Substanzen getestet: Alaun, Sulfazetamid, Kupfersulfat, Silbernitrat, Chaksinjodit und Knoblauch.
Alle 13 Pilze wurden durch 2% Alaun, 30% Natriumsulfazetamid, 0,2% Kupfersulfat und 0,1% Silbernitrat gehemmt (Alternaria, Aspergillus flavus, Aspergillus fumigatus, Aspergillus niger, Candida albicans, Curvularia, Drechslera, Fusarium, Mucor, Penicillium, Rhizopus oryzae, Scopulariopsis und Syncephalastrum). Chaksinjodit, ein Alkaloid aus einer indischen Heilpflanze, Cassia absus hemmte alle Pilze bei einer Konzentration von 0,5%. Niedrigere Konzentrationen waren entweder unwirksam oder nur gegen einige der Pilze wirksam. Knoblauch konnte in einer Konzentration bis zu 1% keine Pilze am Wachstum hindern. Die Wiederholung der Experimente führte zu entsprechenden Ergebnissen.     

真菌性角膜炎的诊断与治疗探讨

目的 探讨真菌性角膜炎的临床表现及治疗情况。方法 将我院附属医院及铜仁市医院五官科2003年10月～2005年10月收治的真菌性角膜炎患者21例就其发病情况、临床表现、治疗效果进行分析。结果 治愈17例，占81％，好转4例，占19％。结论 采用中西药联合、病灶切除、碘酊烧灼、手术等综合治疗取得了较好的效果。     

Therapeutic efficacy of intracameral amphotericin B injection for 60 patients with keratomycosis

? AIM: To evaluate the therapeutic efficacy of intracameral amphotericin B (ICAMB) injection in the treatment of keratomycosis. ? METHODS: The study design was a prospective controlled clinical trial. A total of 60 eyes of 60 patients were divided into two groups, 30 in the ICAMB injection group (group A) and 30 in the control group-topical application amphotericin B (group B). Serial measurements of the size of the keratomycosis-namely, two maximum linear dimensions perpendicular to each other, and the area and perimeter was done at start of therapy and follow up on day 3, 7, and 21. Rate of healing of the keratomycosis were measured as percentage decrease from the baseline parameter at each subsequent follow up. The data were analyzed by the non-parametric Wilcoxon rank sum test. ? RESULTS: The mean time to disappearance of hypopyon was 9.6±9.2 (range:1-26) days in group A and 26.8±20.8 (range:14-62) days in group B (P=0.03). The median percentage decrease in the size of the keratomycosis was significantly greater than that in the cord serum group at day 21(P＜0.05) when measured in terms of the area and perimeter. A greater number of patients showed complete re-epithelialization in group A (n=27) than in group B (n=14) (P＜0.05). None of the patients reported any side effects or discomfort with either treatment. ? CONCLUSION: ICAMB injection leads to faster healing of the keratomycosis refractory to all medical management and reducing time to disapperence of hypopyon compared to topical application amphotericin B.     

真菌性角膜炎的实验观察与治疗

在兔角膜板层注射镰刀菌，形成深层真菌性角膜炎，病灶向浅层发展形成角膜溃疡。另外，可见虹膜炎、前房积脓、新生血管和卫星病灶，未见角膜穿孔。这些真菌性角膜炎用萘替芬治疗，每天用药５次，共治疗１３天。结果表明萘替芬有一定治疗作用，但与对照组相比差异性不够显著（Ｐ〉０．０５）。     

角膜板层切除氟康唑角膜基质注射联合自体结膜移植治疗真菌性角膜炎

目的: 探讨角膜板层切除、氟康唑角膜基质注射、自体结膜移植三联术治疗中、浅层真菌性角膜炎的手术适应证及时机,并对其临床效果进行分析。方法: 对我院2005-03/2010-06药物治疗效果不显著、病灶位于角膜中、浅层的真菌性角膜炎患者168例,大于病灶边缘0.5mm角膜刀板层切除病变角膜至基本透明,角膜基质注射2g/L氟康唑至水肿区大于病变区切缘约0.5mm,依病灶大小自患眼就近取带蒂结膜瓣连续缝合覆盖植床。术后第1d常规开放点眼,并观察角膜感染有无复发,注射造成水肿消退情况,结膜植片血运及患眼刺激症状,及视力变化。结果: 患者157眼成功实施三联手术治疗,占拟实施此术式患者93.5%;术后7d好转率96.2%,术后1mo,复发率3.8%,痊愈率95.5%,好转率0.6%;注射药物后角膜基质水肿平均消退时间为13.4h;术后1mo视力较术前提高者占24.8%,无明显变化者占58.0%,下降者占17.2%;角膜内皮损失率0～8%,平均2.9%;术后刺激症状:轻度87%,中度10%,重度3%;住院天数平均7.3d;住院费用平均2160元;术后3mo发现4例角膜轻微膨隆。结论: 角膜板层切除、氟康唑角膜基质注射、自体结膜移植三联术治疗早、中期真菌性角膜炎经济实用、疗效好、痊愈快,二期常规安全行光学性角膜移植术,对于经济条件差患者和角膜供体短缺地区,三联手术可作为首要选择。     

真菌毒素与角膜真菌感染

真菌毒素（Mycotoxin）是由产毒真菌在适宜的环境条件下产生的具有毒性的次级代谢产物,主要毒性有致癌作用、遗传毒性、致畸作用、肝细胞毒性和免疫抑制等.真菌感染角膜后可能释放真菌毒素导致角膜损害,但至今对真菌毒素在真菌性角膜炎中究竟扮演什么角色,仍不明确.真菌毒素的生物合成受到一系列相关功能基因的调控;此外,pH值、碳氮比等环境条件也能影响其产量.本文就常见的真菌毒素的危害、调控因素及其可能在真菌性角膜炎发生发展中的作用进行综述.