Is inclusion of Sabouraud dextrose agar essential for the laboratory diagnosis of fungal keratitis?

Purpose:

To determine whether the inclusion of Sabouraud dextrose agar (SDA) is essential in the diagnosis of fungal keratitis.

Materials and Methods:

Corneal scrapings of 141 patients with microbial keratitis were smeared and cultured. Sheep blood agar (BA), chocolate agar (CA), SDA, non-nutrient agar (NNA) with Escherichia coli overlay, and brain heart infusion broth (BHI) were evaluated for time taken for growth and cost. The media were also evaluated experimentally for rate of growth and time taken for identification.

Results:

Twenty-six of 39 patients positive for fungus in corneal scrapings by microscopy were culture-positive. Fungus grew on BA in 22/39, on CA in 18/39, on SDA in 17/39, on NNA in 17/39, and on BHI in 13/39 cases. Growth on SDA was higher in ulcers with larger infiltrate (6/18 versus 9/13, P = 0.04). Estimated saving with inclusion of only BA/CA was Rs. 600 per patient. Performance of all media was similar in in vitro experiment although the characteristic spores and color were seen earlier on SDA.

Conclusion:

Fungal keratitis can be reliably confirmed on BA or CA, which support growth of both bacteria and fungus.     

Incidence and microbiological profile of mycotic keratitis in a tertiary care eye hospital: A retrospective analysis

Purpose

To determine the incidence and microbiological profile of mycotic keratitis seen at a tertiary care eye hospital.

Materials and methods

A retrospective review of microbiology records of patients presenting with suspected microbial keratitis seen between January 2006 and December 2009 was performed. Patients with positive fungal cultures were further analyzed for the type of fungus isolated and associated bacterial pathogens.

Results

Microbiology records of 2300 patients with suspected microbial keratitis were reviewed. A microbiological diagnosis of mycotic keratitis was established in 87 (3.8%) patients over a four year period based on positive fungal cultures. The yearly incidence of mycotic keratitis was 3.2% (2006), 4.9% (2007), 3.3% (2008) and 3.6% (2009). Filamentous fungi were isolated more often than yeasts. Aspergillus species followed by Fusarium species and Trichophyton species were the commonest filamentous fungi isolated while Candida albicans was the most frequently encountered yeast. Mixed infections due to fungal and bacterial pathogens were seen in 25/87 (28.7%) patients.

Conclusion

Cumulative incidence of mycotic keratitis was 3.8% over a four year period. Aspergillus species and Candida albicans were the most frequent pathogenic organisms causing mycotic keratitis in this part of the world. Mixed infections were seen in 28.7% of the patients. Knowledge of the “local” etiology within a region may be valuable in the management of mycotic keratitis in instituting an empirical therapy, especially when facilities for microscopy, cultures and antifungal susceptibility are not readily available. The baseline information presented will also be helpful in the planning of a corneal ulcer management strategy and for future studies on mycotic keratitis.     

Evaluation of agent and host factors in progression of mycotic keratitis: A histologic and microbiologic study of 167 corneal buttons

PURPOSE: To evaluate the host and agent factors in the progression of mycotic keratitis through the microbiologic evaluation and histologic study of human corneal buttons obtained at the time of therapeutic keratoplasty. DESIGN: Retrospective noncomparative consecutive case series. MATERIALS: One hundred sixty-seven corneal buttons from 148 patients of microbiologically diagnosed and treated cases of mycotic keratitis who underwent therapeutic keratoplasty between January 1995 and May 1998. METHODS: Therapeutic penetrating keratoplasty, review of microbiologic results, histopathologic and microbiologic evaluation of the corneal buttons of mycotic keratitis MAIN OUTCOME MEASURES: Histologic evaluation of the buttons for morphologic changes, degree and distribution of inflammatory cells, presence or absence of fungal filaments, and their degree and distribution within the corneal buttons. RESULTS: The diagnosis of fungal infection was made on corneal scrapings in 36 cases; whereas in 131 (78%), the fungus was grown in cultures and identified as Aspergillus in 55 (42%), Fusarium in 42 (32%), unidentified hyaline fungi in 22 (17%), dematiaceous (unidentified) in 4 (3%), and others in 8 (6%). The mean interval between diagnosis and keratoplasty was 19 (+/-40) days. From the keratoplasty specimen, the fungus was identified at histologic examination in 127 of 167 (76%) buttons and grown by culture techniques in 76 of 115 (66%) buttons. The fungal species identified in the corneal button were Fusarium in 30 (39%); Aspergillus in 25 (33%); unidentified hyaline in 19 (25%), and others in 2 (3%). Fungus-positive corneal buttons had early surgery (mean, 15 days) compared with fungus-negative (39 days) corneal buttons (P = 0.0005), with 93% fungus positivity in the buttons removed within 2 weeks and 42% after 2 months. In the fungus-positive buttons, there was an inverse correlation between the degree, distribution of inflammatory cells, and fungal filaments (r = -0.255, P = 0.024; r = -0.199, P = 0.027), respectively. The factors necessitating an early keratoplasty were heavy fungal load, deeper penetration of fungus, and possibly insufficient inflammation to combat infection. A granulomatous reaction was noted in the posterior stroma and around the fragmented Descemet's membrane in 23 buttons (13.8%), independent of fungal species. Inflammation was unaffected by elimination of fungus and increasing interval between diagnosis and treatment. CONCLUSIONS: Rapid progression of mycotic keratitis in the early phases is by agent factors such as heavy load and deeper penetration of the fungus, insufficient inflammatory response, and possibly relative ineffectiveness of antifungal agents. Progression in the later phase of mycotic keratitis need not necessarily be agent mediated; it could be either host-modulated, species-related, or drug resistance, thereby suggesting that ideal treatment regimens should include sensitivity-based antifungal therapy aided by in vivo monitoring of fungal filaments.     

Polymicrobial keratomycosis in a three-year-old child

PURPOSE: To describe a previously unreported case of polymicrobial mycotic keratitis caused by an association between Candida lusitaniae, C. parapsilosis, and Geotrichum candidum. METHODS: A three-year-old child with an antecedent trauma with vegetable matter and a prolonged use of corticosteroid eyedrops developed fungal keratitis. RESULTS: The isolates of the corneal scraping using Sabaraud dextrose agar grew C. lusitaniae, C. parapsilosis, and G. candidum. After topical 0.2% and systemic fluconazole treatment, the corneal lesion resolved with no recurrence. CONCLUSIONS: Corneal trauma with vegetables and the indiscriminate use of corticosteroids are important risk factors for mycotic keratitis. A combination of topical 0.2% and systemic fluconazole therapy was effective in the treatment of this mycotic association. This is the first report of fungal keratitis caused by C. lusitaniae and G. candidum.     

Polymicrobial Keratomycosis in a Three-Year-Old Child

Purpose: To describe a previously unreported case of polymicrobial mycotic keratitis caused by an association between Candida lusitaniae, C. parapsilosis, and Geotrichum candidum. Methods: A three-year-old child with an antecedent trauma with vegetable matter and a prolonged use of corticosteroid eyedrops developed fungal keratitis. Results: The isolates of the corneal scraping using Sabaraud dextrose agar grew C. lusitaniae, C. parapsilosis, and G. candidum. After topical 0.2% and systemic fluconazole treatment, the corneal lesion resolved with no recurrence. Conclusions: Corneal trauma with vegetables and the indiscriminate use of corticosteroids are important risk factors for mycotic keratitis. A combination of topical 0.2% and systemic fluconazole therapy was effective in the treatment of this mycotic association. This is the first report of fungal keratitis caused by C. lusitaniae and G. candidum.     

巴基斯坦南部84例真菌性角膜炎的研究

目的:确定真菌性角膜炎的易感因素、致病真菌,以及提高相应的实验室诊断能力。方法:240例240眼临床疑似微生物感染的角膜溃疡患者进入本研究。通过病史回溯及裂隙灯检查获得病例资料。采用标准技术完成角膜刮片检查。取部分刮片样本加入10%的氢氧化钾液后,以及采用革兰氏和吉姆萨染色,在直接在显微镜下观察是否存在真菌、细菌和棘阿米巴。另一部分刮片样本直接接种在血琼脂、麦康基琼脂、巧克力琼脂和萨布罗琼脂表面。结果:在纳入本研究的240例伴有脓性分泌物的角膜溃疡患者中,84例(35%)被确认为真菌感染。其中48例(57.1%)为男性,其余36例(42.9%)为女性。年龄分布从22-80岁。其中60例(71.4%)为农村患者,其余24例(28.6%)为城市居民。18例(21.4%)曾有植物外伤史。病变高发于10月至11月间。在84例确诊为真菌性角膜炎的患眼,致病原因为单纯真菌者74眼(80.10%),其余10眼(11.90%)为真菌合并细菌感染。分离得到的最常见致病微生物为白色念珠菌,在66眼(78.6%)中被发现。结论:在巴基斯坦南部,感染性角膜溃疡中,最常见的是真菌性角膜炎,而白色念珠菌是最常见的致病原。采用加入10%的氢氧化钾液后直接显微镜下观察是此类感染原简单、迅速、价廉及可信的诊断方法。     

Fungal keratitis:84 cases report in Southern Pakistan

AIM: To identify the predisposing factors, causative fungi and to improve the facilities for the laboratory diagnosis of fungal keratitis. METHODS: Two hundred and forty eyes of 240 patients of clinically suspected microbial corneal ulcer were included in the study. Data was collected through history and slit lamp examination. Using standard techniques, corneal scraping was performed.A portion of each scraping was examined by direct microscopy for the presence of fungi, bacteria and acanthamoeba by using 100g/L potassium hydroxide and also by Gram and Giemsa stainings. Another portion was inoculated directly on the surface of solid media such as blood agar, Mac-Conkey agar, chocolate agar and Sabouraud agar in C-shaped streaks for culture. RESULTS: A total of 240 patients with suppurative corneal ulcer were enrolled in the study, out of which fungi were identified in 84 (35%) patients. Of these, 48 (57.1%) were males and 36 (42.9%) females. The age ranged between 22-80 years. Sixty patients (71.4%) belonged to rural population and twenty four (28.6%) to urban population. Trauma with vegetative material was the most frequent cause noted in eighteen (21.4%) patients. Peak incidence was in the months of October-November. Out of 84 eyes with fungal keratitis, fungi alone were the etiologic agents in 74 (80.10%) cases and bacteria with fungi were identified in 10 (11.90%). The most frequently isolated organism was Candida albicans which was found in 66 (78.6%) patients. CONCLUSION: Fungal keratitis is the leading cause of infective corneal ulcer and Candida albicans being the most commonly isolated pathogen in the patients belonging to Southern Pakistan. The direct microscopic examination with potassium hydroxide 100g/L method is a simple, rapid, inexpensive and reliable method in the diagnosis of this infection.     

Single culture media in infectious keratitis.

PURPOSE: To evaluate the usefulness of the various culture media used in the traditional workup in infectious keratitis. METHODS: Microbiology data sheets from all corneal cultures performed at the University of California Davis Medical Center over a 1-year period were reviewed retrospectively. RESULTS: Bacterial cultures were sent in 76 cases. In 19 cases, culture specimens from ulcers were plated onto blood, chocolate, and inhibitory mold agar and were inoculated into an anaerobic medium. In 58 cases, blood and chocolate agar were sent. In 70% of cases, blood and chocolate agar provided identical information. Inhibitory mold agar was positive twice in 39 plates sent. A fungal pathogen had been identified on chocolate agar plates sent for these cases. CONCLUSION: In the evaluation of infectious keratitis, plating onto chocolate agar or blood agar alone is a reasonable alternative to sending multiple cultures.     

聚合酶链反应技术对真菌性角膜炎快速诊断的实验研究及临床应用

目的：探讨聚合酶链反应（PCR）技术与真菌性角膜炎快速诊断的可行性和优越性及其在临床应用中的价值。方法：以一段真菌特异性通用纸上谈兵物并配合热启动聚合酶链反应技术来检测临床上常见的病原性真菌。结果：8种真菌均扩增出了一种310bp的阳性条带,而其它细菌、病毒和人体角膜组织均为阴性。在动物造型中,PCR技术的敏感性为86.7%,特异性为100%;真菌培养的敏感性为56.7%,特异性为72.7%。在对临床标本的研究中,PCR技术的敏感性为66.6%,真菌培养的敏感性为33.3%,角膜刮片的敏感性为40%。结论：采用通用性特异引物并配合热启动PCR技术来检测实验室和临床标本中是否有真菌存在有重要的应用价值。     

90例真菌性角膜炎菌种鉴定

目的 分析2000年河南省的真菌性角膜炎致病菌种类。方法 收集真菌性角膜炎患者的标本，Saboraud‘s琼脂培养基分离后，选择合适培养基培养，10%KOH湿封片观察鉴定。结果 共分离出真菌菌株10属90株，致病真菌以镰孢菌属最多（73.3%），尤以紫二孢镰孢菌最多见（18.9%）。结论 2000年河南省的真菌性角膜炎以镰孢菌属最多，镰孢菌属的基础研究需进一步加强。     

Spectrum of fungal keratitis in North India

PURPOSE: To report the epidemiologic features and laboratory results of 191 consecutive cases of fungal keratitis presenting to a tertiary level superspecialty teaching hospital of North India. METHODS: A prospective hospital-based study was carried out on 485 consecutive patients presenting with corneal ulcers to the outpatient department of Guru Nanak Eye Center, Maulana Azad Medical College, New Delhi, from January 1999 to June 2001. The sociodemographic data, predisposing risk factors, clinical details, prior treatment modalities, laboratory results, and visual outcomes were analyzed. RESULTS: Diagnosis of mycotic keratitis was established in 191 (39%) out of the total study group of 485 cases. Direct microscopic examination of KOH mounts and Gram-stained smears revealed presence of fungal elements in the corneal scrapings in 119 (62.3%) and 114 (60%) of the subsequently fungal culture-positive cases, respectively. Men (68%) were more commonly affected by fungal keratitis than women (32%). Young adults 31-40 years of age were the most common age group to be involved (36%). Predisposing risk factors were noted in 79%, with corneal trauma 42%, contact lens wear 25%, and topical corticosteroids in 21% patients. The spectrum of fungi isolated were Aspergillus species in 78 (41%) followed by Curvularia species in 55 (29%). CONCLUSIONS: In contrast to other studies from our subcontinent, we found Aspergillus niger to be the most common fungal isolate, followed by Curvularia species in culture-proven cases of fungal keratitis. Direct microscopic examination of KOH mounts emerged as a rapid, reliable, and inexpensive diagnostic modality, with a sensitivity of 62%, which would facilitate the institution of early antifungal therapy before the culture results became available, thus proving to be sight saving.     

海南省81例真菌性角膜溃疡病原学分析

目的:探讨中国海南省真菌性角膜溃疡的病原学特点及流行病学特征.方法:收集2014-01/2016-12在中山大学中山眼科中心海南眼科医院住院治疗的真菌性角膜溃疡确诊病例81例81眼的基本资料.分析致病真菌种群分布特点、患者年龄分布特点、患病季节分布构成比以及疾病危险因素.回顾分析真菌性角膜溃疡治疗方式,计算角膜溃疡坏死组织的真菌培养及鉴定、真菌涂片结果阳性率,卡方检验比较两种实验室检查方法阳性检出率.结果:选取临床诊断真菌性角膜溃疡患者81例81眼,其中男54眼,女27眼,年龄中位数51岁.角膜外伤是最常见的危险因素.≥50岁患病人数45眼(55%),占比最大;其次为>30~<50岁年龄段人群组28眼(35%);≤30岁人群组8眼(10%),占比最小.第一季度是全年真菌性角膜溃疡的高发时段,患病人数占36%.81份真菌培养样本阳性例数71眼,阳性率88%.80份角膜坏死物涂片样本,阳性样本共41眼,阳性率51%.真菌涂片阳性检出率低于真菌培养,差异有统计学意义(x2=23.730, P<0.01).主要致病性真菌依次为镰刀菌属34眼(48%),不产孢子菌11眼(15%).治疗上均采用了多种抗真菌药物治疗,23眼(28%)联合角膜清创+结膜瓣遮盖术治疗;9眼(11%)联合角膜移植术;7眼(9%)联合眼内容物剜除术.结论:中国海南省本岛真菌性角膜溃疡的最主要致病菌是镰刀菌属、不产孢子菌,高发季节为1~3mo.患者多有角膜外伤史.抗真菌药物及手术治疗是目前真菌性角膜溃疡的主要治疗手段.与真菌涂片相比较,真菌培养具有更高的阳性检出率.     

Efficacy of topical and systemic itraconazole as a broad-spectrum antifungal agent in mycotic corneal ulcer. A preliminary study

PURPOSE: To evaluate the efficacy of topical (1%) and systemic itraconazole against common fungi such as Aspergillus and other filamentous fungi that cause mycotic corneal ulcer. METHODS: A prospective randomised, controlled study was done in 54 clinically suspected cases of fungal keratitis of which 44 were culture proven. Half the cases (n=27) with superficial involvement were treated with only topical itraconazole (1%) and the other half were treated with both topical and systemic itraconazole. RESULTS: Aspergillus, Penicillium and Fusarium were the most common fungi isolated. The ulcer resolved in 42 eyes (77%) and 12 eyes (23%) did not respond well to treatment. Four of 12 non-responding eyes were caused by Fusarium species. CONCLUSION: Itraconazole, given either topically or systemically, is effective in treating mycotic corneal ulcers.     

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.     

Increased level of (1,3)-β-D-glucan in tear fluid of mycotic keratitis

Purpose  Increased concentration of (1,3)-β-D-glucan, one of the major components of fungal cell walls, is detected in the serum of systemic fungal infection. In our study, the concentration of (1,3)-β-D-glucan was measured in the tear fluid of patients with mycotic keratitis. Methods  Tear fluid was collected from patients with fungal keratitis (n = 4) and bacterial corneal ulcers (n = 4) with or without corneal scraping. In addition, tear fluid was collected from patients without corneal diseases. Results  The concentration of (1,3)-β-D-glucan in tear fluid collected without corneal scraping was 4.0 ± 3.5, 5.8 ± 2.6, 184 ± 128 pg/ml in the control, bacterial corneal ulcer, and mycotic keratitis samples respectively. The concentration of (1,3)-β-D-glucan in tear fluid collected after scraping the corneal lesions with a tip of glass capillary was 4.4 ± 1.3, 8.2 ± 5.2 and >1,000 pg/ml in the control, bacterial ulcer, and mycotic keratitis samples respectively. Conclusions  A significant increase in (1,3)-β-D-glucan was detected in tear samples from patients with mycotic keratitis. Measuring the concentration of (1,3)-β-D-glucan in tear fluid might be helpful in the diagnosis of mycotic keratitis. Financial Interest: None     

Predisposing factors and etiologic diagnosis of ulcerative keratitis

PURPOSE: To determine the predisposing factors, demographic characteristics, and etiology of ulcerative keratitis in a referral center in Bangkok, Thailand. METHODS: The medical records of admitted patients with positive-culture ulcerative keratitis were retrospectively reviewed for demographic data, predisposing factors, and microbial culture results. Predisposing factors were compared between bacterial and fungal keratitis. RESULTS: From January 2001 to December 2004, there were 127 positive-culture ulcerative keratitis cases. The most frequent microbiological diagnosis was bacterial keratitis (76 eyes, 60%), followed by fungal (48 eyes, 38%) and Acanthamoeba keratitis (3 eyes, 2%). The most common organisms isolated were Pseudomonas spp. for bacteria and Fusarium spp. for fungus. Compared with bacterial keratitis, fungal keratitis was more likely to be associated with ocular trauma (odds ratio = 11.20; 95% confidence interval, 3.62-34.66) but less likely to be associated with contact lens wear (odds ratio = 0.02; 95% confidence interval, 0.01-0.08). CONCLUSIONS: In our study, Pseudomonas and Fusarium species are the most common causes of bacterial and fungal keratitis, respectively. Fungal keratitis was more likely than bacterial keratitis to be associated with ocular trauma, whereas fungal keratitis was less likely to be associated with contact lens wear.     

互隔交链孢霉菌性角膜炎的临床与病原学研究

目的探讨互隔交链孢霉菌性角膜溃疡的临床和实验室特征。方法对真菌性角膜溃疡患者,裂隙灯显微镜下观察临床表现后,角膜刮片作10％KOH湿片、Giemsa染色显微镜检查,真菌菌丝阳性者作真菌培养鉴定,并根据美国国家临床实验室标准化委员会（NCCLS）M38-A方案进行体外药敏试验,同时对患者给予两性霉素B、酮康唑、氟康唑滴眼液治疗,并观察抗真菌药物的疗效。以分离的互隔交链孢霉菌株制作兔真菌性角膜溃疡模型,5d和14d后处死兔,取角膜行真菌培养和组织学检查。结果互隔交链孢霉菌性角膜炎患者均为单眼患病,患者和兔模型的角膜溃疡灶均较浅薄,呈淡灰色,边界模糊。显微镜下角膜刮片可见直且粗大的无色隔膜菌丝,并有大量球形厚垣孢子位于菌丝顶端或相连。分离培养基中该种真菌菌丝为棕色,壁砖墙分隔的深棕色孢子链生于棕色分生孢子梗顶端。体外药物敏感试验显示该种真菌对两性霉素B、酮康唑、氟康唑、伏立康唑、咪康唑、益康唑和特比萘芬敏感,对克霉唑、氟胞嘧啶及伊曲康唑耐药。23例患者除1例手术治愈外,其余22例用两性霉素B、酮康唑、氟康唑联合频繁滴眼治愈,平均治愈时间34d。结论互隔交链孢霉菌性角膜溃疡具有独特的临床和实验室特征,对角膜毒力较低,抗真菌药物联合滴眼可完全治愈。     

Evaluation of corneal scraping smear examination methods in the diagnosis of bacterial and fungal keratitis: a survey of eight years of laboratory experience

PURPOSE: To determine the sensitivity, specificity, and predictive values of Gram and potassium hydroxide with calcofluor white (KOH+CFW) stains in the diagnosis of early and advanced microbial keratitis, a retrospective analysis of comparative data from a prospectively collected database was done. METHODS: Patients with nonviral microbial keratitis seen at L.V. Prasad Eye Institute between February 1991 and December 1998 were included in the study. The type of bacteria seen on Gram stain was determined from 251 corneal scrapings from patients with early keratitis and 841 corneal scrapings from patients with advanced keratitis. The presence of fungi in corneal scrapings was determined by KOH+CFW stain of 114 and 363 scrapings from patients with early and advanced keratitis, respectively. The smear findings were compared with culture results to analyze specificity, sensitivity, and predictive values of the staining techniques. RESULTS: The sensitivity of Gram stain in the detection of bacteria was 36.0% in early and 40.9% in advanced keratitis cases; however, the specificity was higher in both groups (84.9% and 87.1%, respectively). Comparatively, the sensitivity and specificity of fungal detection were higher using KOH+CFW in early (61.1% and 99.0%, respectively) as well as advanced keratitis (87.7% and 83.7%, respectively). Predictive values were high for KOH+CFW in fungus detection, while they were poor for Gram stain in bacteria detection. In advanced keratitis cases, the false positives were higher in fungal detection (16.3%) than in bacterial detection (10.3%), while the false negatives were significantly higher in bacterial detection compared with fungal detection (59.1% versus 12.3%, p< 0.0001). In early keratitis, on the other hand, both false positives and false negatives for bacterial detection were significantly higher than fungal detection. CONCLUSIONS: Decisions can reliably be based on KOH+CFW stain of corneal scrapings for initiation of antifungal therapy in mycotic keratitis. The results of Gram stain, on the other hand, have limited value in therapeutic decisions for bacterial keratitis. Therefore, the search for a better modality for early and efficient diagnosis of bacterial keratitis needs to continue.     

Mycotic keratitis in non-steroid exposed vernal keratoconjunctivitis.

PURPOSE: To report a patient with vernal keratoconjunctivitis who developed mycotic keratitis in absence of known risk factors. METHODS: A 17-year-old male suffering from vernal keratoconjunctivitis presented with infective keratitis. The patient had been treated in the past with topical antihistaminics and vasoconstrictors. The patient had not been exposed to topical steroids in 2 years of follow-up. He did not have dry eye or corneal micro or macroerosions prior to the development of infective keratitis. Corneal scrapings were obtained and subjected to KOH wet mount smear, calcofluor and Grams stain as well as bacterial culture sensitivity and fungal culture. RESULTS: Clinical diagnosis of mycotic keratitis in association with vernal conjunctivitis was supported by microbiological investigations. KOH wet mount and calcofluor staining showed presence of filamentous septate hyphae while fungal culture showed growth of aspergillus fumigatus. Antifungal therapy was initiated in the form of topical natamycin 5% suspension to which the patient responded and recovered 6/6 final visual acuity. CONCLUSION: The authors wish to conclude that patients suffering from vernal keratoconjunctivitis, even in the absence of corneal involvement, steroid exposure and trauma, may be at increased risk of developing keratomycosis.     

Fungal spectrum identified by a new slide culture and in vitro drug susceptibility using Etest in fungal keratitis

PURPOSE: To investigate the spectrum of fungal species causing keratitis and to test antifungal drug susceptibility to each isolate using Etest. METHODS: Microbial cultures were performed for patients who were clinically diagnosed with fungal keratitis between September 2002 and July 2004. Modified slide culture was established to identify the fungal species of the isolates. Etest (AB BIODISK, Solna, Sweden) was applied to determine the antifungal agent susceptibility of each isolate to itraconazole, fluconazole, and amphotericin B in vitro, respectively. RESULTS: Among 73 eyes of 73 patients with clinical diagnosis of fungal keratitis, 61 strains of fungi were isolated from 61 eyes. The rate of positive culture was 81.3% of all cases. The spectrum of fungal species involved: 58 (95.1%) isolates of filamentous fungi, including the two most common genera-Fusarium (n = 33, 54.1%) and Aspergillus (n = 9, 14.8%),-followed by 16 (26.2%) isolates of other genera of filamentous fungi such as Alternaria (n = 3, 4.9%), Trichophyton (n = 3, 4.9%), Curvularia (n = 2, 3.3%), Chrysosporium (n = 2, 3.3%), Acremonium (n = 2, 3.3%), and Scedosporium (n = 1, 1.6%), 1 (1.6%) yeast of Candida, as well as two (3.3%) dimorphic fungi of Blastomyces and Sporothrix isolate each. Three filamentous fungi of the isolates failed to be identified according to the information provided by slide culture. The results of Etest showed that 20 (60.6%) isolates of Fusarium were susceptible to amphotericin B, whereas all of them were resistant to itraconazole and fluconazole. All nine (100%) isolates of Aspergillus were sensitive to itraconazole, whereas four (44.4%) of them were sensitive to amphotericin B, and only two (22.2%) of them were sensitive to fluconazole. Seventeen (89.5%), 13 (68.4%), and 10 (52.6%) isolates of the remaining 19 organisms were sensitive to amphotericin B, itraconazole, and fluconazole, respectively. CONCLUSIONS: Fusarium and Aspergillus are the most frequent pathogenic organisms in causing fungal keratitis, whereas other species of fungi can also cause corneal infection. In vitro Etest for assessing antifungal drug susceptibility is a simple and practical method and may provide referential information for clinical consideration of choosing antifungal agents to treat fungal keratitis.