A self-assembled clavanin A-coated amniotic membrane scaffold for the prevention of biofilm formation by ocular surface fungal pathogens.

Amniotic membrane (AM) is frequently used in ophthalmologic surgery for rapid ocular surface reconstruction. Sometimes it may create a major problem with associated infections after biofilm formation over the membrane. To overcome this problem, AM was coated with the antimicrobial peptide clavanin A. The antifungal activity of clavanin A in the native and self-assembled form was determined against the common ocular surface pathogens Candida albicans, Aspergillus fumigatus, Alternaria sp. and Fusarium sp. Biofilm formation over the coated surface was significantly reduced in comparison with the uncoated membrane. The coated membrane revealed effectiveness in terms of biocompatibility, cell attachment colonization when tested in non-cancerous 3T3 and human embryonic kidney (HEK)-293 cell lines. Clavanin A-coated AM also exhibited excellent physical, morphological and antifungal characteristics, indicating potential applicability for ocular surface infection control.

Rhodotorula mucilaginosa Keratitis: a rare fungus from Eastern India.

Rhodotorula mucilaginosa rarely cause keratitis in immunocompromised individuals. A 30 year old male with history of minor trauma presented with cotton wool like stromal infiltration and hypopyon in left eye. Microbiological examination of corneal scraping showed fungal hyphae and yeast cells in direct smear. Molecular identification of the organism was performed which showed 100% homology with Rhodotorula mucilaginosa. Management of these cases is difficult often necessitating surgical procedures. However further reports are necessary to understand the disease and establish a treatment protocol.

Kaajal fights against eye pathogens and is safe for eye make-up: a reinvestigation of an ancient practice.

The present report includes a scientific view of the traditional use of 'Kaajal' from the leaf of Euphorbia neriifolia, an Indian spurge tree locally called the 'Monosha' plant. A thorough analysis of the material demonstrates that it contains carbon nanoparticles having two uniform sizes of 2-3 nm and 35-45 nm. Further study revealed that Kaajal has antimicrobial properties against eye pathogens and is biocompatible for cosmetic use.

N, N'-Olefin functionalized bis-imidazolium gold(I) salt is an efficient candidate to control keratitis-associated eye infection.

Keratitis treatment has become more complicated due to the emergence of bacterial or fungal pathogens with enhanced antibiotic resistance. The pharmaceutical applications of N-heterocyclic carbene complexes have received remarkable attention due to their antimicrobial properties. In this paper, the new precursor, 3,3'-(p-phenylenedimethylene) bis{1-(2- methyl-allyl)imidazolium} bromide (1a) and its analogous PF6 salt (1b) were synthesized. Furthermore, silver(I) and gold(I) -N-heterocyclic carbene (NHC) complexes [Ag2LBr2/Au2LBr2; 2a/3a], [(Ag2L2)(PF6)2/(Au2L2)(PF6)2; 2b/3b] were developed from their corresponding ligands. All compounds were screened for their antimicrobial activities against multiple keratitis-associated human eye pathogens, including bacteria and fungi. Complexes 2a and 3a showed highest activity, and the effectiveness of 3a was also studied, focusing eradication of pathogen biofilm. Furthermore, the structures of 1a, 2a and 3b were determined using single crystal X-ray analysis, 2b and 3a were optimized theoretically. The mechanism of action of 3a was evaluated by scanning electron microscopy and docking experiments, suggesting that its target is the cell membrane. In summary, 3a may be helpful in developing antimicrobial therapies in patients suffering from keratitis-associated eye infections caused by multidrug-resistant pathogens.

Schizophyllum commune: a new organism in eye infection.

PURPOSE: We report a case of mycotic keratitis caused by a rare fungus Schizophyllum commune. METHODS: Clinical examination, slit-lamp examination, and microbiological evaluation of the corneal ulcer were done, and its treatment outcome was studied. The fungal etiology was established by conventional microbiological techniques, polymerase chain reaction and speciation by DNA sequencing. RESULTS: Corneal scraping showed the presence of fungal filaments. The fungus was identified as S. commune based on DNA sequence analysis of the internal transcribed spacer region. The organism was susceptible to amphotericin B and voriconazole and demonstrated resistance to anidulafungin, itraconazole, and fluconazole. Therapeutic keratoplasty was performed but there was recurrence of the infection in the graft, which was controlled with topical voriconazole and intracameral amphotericin B. At the end of 3 months, the affected eye had developed phthisis bulbi. CONCLUSION: The best of our knowledge, this is the first reported case of keratitis caused by the rare fungus S. commune. Management of these cases is difficult, and surgical procedures may be needed.

Candida fermentati: a rare yeast involved in fungal keratitis.

PURPOSE: The aim of this study was to report a rare case of fungal keratitis from Eastern India. METHODS: This is a case report. RESULTS: A 52-year-old man with a history of minor trauma presented with a total corneal ulcer and hypopyon in the left eye. Microbiologic examination of corneal scrapings showed yeast cells in direct smear and typical yeast colonies on multiple solid agar media. Identification of the organism isolated in the culture was performed using the D1/D2 region of the large subunit (LSU 28S rDNA)-based molecular technique. Polymerase chain reaction amplified a band with a sequence that was 100% homologous with that of Candida fermentati. The organism was susceptible to amphotericin B and anidulafungin and demonstrated resistance to voriconazole, itraconazole, and fluconazole. Therapeutic keratoplasty was performed, followed by the recurrence of the infection in the graft, which was controlled with topical and intracameral amphotericin B. At the end of 3 months, the affected eye had developed phthisis bulbi. CONCLUSIONS: This is the first report of isolation of C. fermentati, a species closely related to Candida guilliermondii, from keratitis. Molecular diagnostic techniques are helpful in the accurate identification of this organism, which is clinically important in view of an antifungal susceptibility pattern that differs from that of other yeasts and for selection of appropriate therapy.

Comparison of Goldmann Tonometry and Dynamic Contour Tonometry in Normal and Descemet Stripping Endothelial Keratoplasty Eyes.

PURPOSE: This study aimed to compare intraocular pressures (IOP) measured by dynamic contour tonometry (DCT) with Goldmann tonometry (GAT) in Descemet stripping endothelial keratoplasty (DSEK) eyes. The agreeability between the 2 established methods and their relation to central corneal thickness (CCT) was also studied. DESIGN: A comparative case series. METHODS: Forty-six eyes of DSEK were enrolled at least 3 months after the procedure. Thirty-five other eyes of the same patient cohort served as control. Intraocular pressures were measured by DCT and GAT by 2 different examiners in a randomized manner. Central corneal thickness was measured by ultrasonic pachymetry. Statistical analysis was used for the calculation of means, SD, coefficient of determination, linear regression, and Bland and Altman plot. Statistical significance was considered as P < 0.05. RESULTS: Mean CCT was 624.49 µm and 518.41 µm in DSEK and control eyes, respectively. Mean IOP by GAT and DCT, respectively, was 19.4 and 21.5 mm Hg in DSEK eyes and 15.12 and 17.6 mm Hg in controls. Descemet stripping endothelial keratoplasty eyes showed universally higher IOP by either method in comparison with normal controls. Dynamic contour tonometry reading was significantly higher than GAT in both (P < 0.05) groups. The correlation between IOP and CCT was not significant in DSEK eyes (P > 0.05). CONCLUSIONS: A higher baseline IOP is to be expected in DSEK eyes, which is independent of corneal thickness. Both GAT and DCT can be used as a standard method to measure IOP in DSEK eyes, but their readings cannot be used interchangeably.

Lasiodiplodia theobromae keratitis: a case report and review of literature.

A case report and review of literature is reported of a rare case of fungal keratitis from eastern India. A 32-year-old woman with a history of vegetative trauma presented with keratitis in left eye. Microbiological examination of corneal scraping showed refractile hyphae with aseptate branching filaments and black pigmented colonies on multiple solid agar medium. Organism was identified from culture using D1/D2 region of LSU (Large Sub Unit: 28S rDNA)-based molecular technique. PCR amplified a band with a sequence that was 100 % homologous with Lasiodiplodia theobromae. The organism was susceptible to amphotericin B and voriconazole and demonstrated resistance to itraconazole and fluconazole. A therapeutic keratoplasty was performed following non-responsiveness to initial topical voriconazole (2 %) therapy. Recurrence in graft was controlled with topical voriconazole and intracameral amphotericin B. However, the graft failed at the end of 3 months. L. theobromae is a rare cause of fungal keratitis. Management of these cases is difficult, often involving surgical procedures.

Effects of lactoferricin B against keratitis-associated fungal biofilms.

Biofilms are considered as the most important developmental characteristics in ocular infections. Biofilm eradication is a major challenge today to overcome the incidence of drug resistance. This report demonstrates the in vitro ability of biofilm formation on contact lens by three common keratitis-associated fungal pathogens, namely, Aspergillus fumigatus, Fusarium solani, and Candida albicans. Antifungal sensitivity testing performed for both planktonic cells and biofilm revealed the sessile phenotype to be resistant at MIC levels for the planktonic cells and also at higher concentrations. A prototype lens care solution was also found to be partially effective in eradication of the mature biofilm from contact lenses. Lactoferricin B (Lacf, 64 µg/ml), an antimicrobial peptide, exhibited almost no effect on the sessile phenotype. However, the combinatory effect of Lacf with antifungals against planktonic cells and biofilms of three fungal strains that were isolated from keratitis patients exhibited a reduction of antifungal dose more than eightfold. Furthermore, the effect of Lacf in lens care solution against biofilms in which those strains formed was eradicated successfully. These results suggest that lactoferricin B could be a promising candidate for clinical use in improving biofilm susceptibility to antifungals and also as an antibiofilm-antifungal additive in lens care solution.

Candida keratitis: emerging problem in India.

PURPOSE: To determine the epidemiological characteristics and outcome of Candida keratitis in a Cornea Care Unit of Kolkata-based tertiary eye hospital. METHODS: A retrospective, noncomparative, observational case series involving patients of culture-proven fungal keratitis from January 2008 to December 2008. A total of 85 cases of culture-proven fungal keratitis were identified. Of these, 16 cases were caused by Candida sp and selected for the study. The records were analyzed for demographics, risk factors, mode of management (medical or surgical), indication of surgical therapy, and the response to treatment with final outcome. Medical therapy consisted of topical amphotericin B with or without intracameral application after obtaining culture reports. Surgical therapy included application of tissue adhesive with bandage contact lens and therapeutic keratoplasty. RESULTS: All cases of Candida keratitis were caused by Candida albicans accounting for 16 cases [18.81%; 95% confidence interval (CI), 11.8-28.5] of total culture-positive fungal keratitis. We found postsurgical steroid therapy in 8 cases as most important association, followed by diabetes and trauma (4 cases each) as next common comorbidities. All patients required therapeutic keratoplasty. Surgical indications were corneal melt in 10 cases (62.5%; 95% CI, 38.5-81.6), extension up to limbus in 2 cases (12.5%; 95% CI, 12.2-37.2) and nonresponse with worsening in 4 cases (25%; 95% CI, 19.7-49.9). Final outcome consists of phthisis bulbi in 3 cases (18.8%; 95% CI, 5.8-43.8), failed graft in 7 cases (43.7%; 95% CI, 23-66.8), and clear graft in 6 cases (37.5%; 95% CI, 18.4-61.5). CONCLUSIONS: Candida is a new concern in developing countries like India. We are concerned about the poorer outcome, probably resulting from our unpreparedness and failure of medical therapy leading to more complication and requiring surgical intervention in higher numbers.

Characteristics of microsporidial keratoconjunctivitis in an eastern Indian cohort: a case series.

BACKGROUND: Microsporidia are intracellular parasites responsible for human infections. Recently, there has been an increase in the incidence of microsporidial keratoconjunctivitis (MKC) affecting normal individuals worldwide. AIM: To determine the characteristics of MKC in an Indian cohort. MATERIALS AND METHODS: This is a retrospective, noncomparative, observational case series, involving patients with MKC between June and September 2009. Of the 24 patients identified, microbiological confirmation in direct smear was obtained in 22 cases and selected. Standard microbiological workup was performed in all the cases. We studied the demographics, predisposing conditions, antecedent treatment received before presentation, clinical characteristics, treatment offered, and resolution time with sequel. The management consisted of simple debridement and application of chloramphenicol ointment (1%) two times a day. RESULTS: Mean age of onset was 18.7 years (95% CI, 15.7-21.7; range, 11-36 s years). All patients gave history of prior outdoor activity and exposure to rain water/mud. Antecedent treatment comprised of Acyclovir eye ointment (45.4%) and antibiotic eye drop (27.3%) most commonly. Microsporidia were identified in Gram stain (81.8%), 10% potassium hydroxide mount (72.7%), modified Ziehl-Neelsen staining (36.4%), and Giemsa (18.2%). Majority presented as unilateral superficial keratoconjunctivitis with punctate epithelial keratitis. Mean resolution time was 9 days (95%CI, 7.9-10.2). CONCLUSIONS: MKC can occur in normal patients with exposure to rain and mud, related to outdoor activity often misdiagnosed as viral ocular infections. Strong clinical suspicion with proper microbiological evaluation helps to diagnose this commonly misdiagnosed condition.

Bacterial keratitis after manual descemet stripping endothelial keratoplasty--a different pathophysiology?

PURPOSE: To report two cases of infective keratitis caused by Pseudomonas aeruginosa after manual descemet stripping endothelial keratoplasty (DSEK). METHOD: Case report. RESULTS: Case 1, a 65-year-old woman, presented with a central corneal infiltrate after manual DSEK for pseudophakic bullous keratopathy, 6 weeks before this presentation. Case 2, a 55-year-old woman, developed a paracentral infiltrate after 7 weeks of undergoing DSEK. Both cases demonstrated Pseudomonas aeruginosa. The risk factors were analyzed. Both the eyes were treated with ciprofloxacin (0.3%) eye drop, while topical dexamethasone (0.1%) was used additionally in the second case. Resolution occurred after 48 days (case 1) and 21 days (case 2), respectively. Although the DSEK failed in case 1, the graft is clear in case 2 at 3 months. CONCLUSIONS: Microbial keratitis after DSEK is a new area of concern in the postoperative period, both in developed and in developing countries, with a poor impact on graft outcome. Surface-related factors may predispose to this condition, particularly in pseudophakic bullous keratopathy, countering the advantage gained from absence of suture-related infections common after conventional keratoplasty. The location of the ulcer may have an impact on outcome, as also the inflammatory status. Judicious use of topical steroids early in the course of the disease may improve the immediate graft survival.

Epidemiological profile of fungal keratitis in urban population of West Bengal, India.

BACKGROUND: Corneal diseases are one of the major causes of visual loss and blindness, second only to cataract. Amongst corneal diseases, microbial keratitis is a major blinding disease. In some countries, fungal keratitis accounts for almost 50% of patients with culture-proven microbial keratitis. AIM: This study was conducted to determine the epidemiological characteristics of fungal keratitis in an urban population of West Bengal and identify the specific pathogenic organisms. METHODS: The charts of patients with microbial keratitis who attended the Cornea Services of Priyamvada Birla Aravind Eye Hospital from January to December 2008 were retrospectively reviewed. Records of patients with 10% KOH mount and culture positive fungal keratitis were analyzed for epidemiological features, laboratory findings and treatment outcomes. RESULTS: Of the 289 patients of microbial keratitis included in the study, 110 patients (38.06%) were diagnosed with fungal keratitis (10% KOH mount positive). Of the 110 patients, 74 (67.27%) fitted the study inclusion criteria (10% KOH mount and culture positive). Forty five of 74 patients (60.81%) in the study group were in the older age group (>50 years). Ocular trauma in 35 cases (47.29%) was identified as a high risk factor and vegetative injuries in 17 cases (22.97%) were identified as a significant cause for fungal keratitis. Maximum organism source was from corneal scrapings in 41 cases (55%). The predominant fungal species isolated was Aspergillus sp (55.40%) followed by Candida albicans 14 cases (18.91%) and Fusarium sp. in 8 cases (10.81%). Agricultural activity related ocular trauma was the principal cause of mycotic keratitis and males were more commonly affected. Thirty of 74 cases (40.55%) of the culture positive patients healed with corneal scar formation with medical treatment whereas 44 cases (59.45%) required therapeutic keratoplasty. CONCLUSION: Fungal keratitis is an important cause of microbial keratitis with injury to the cornea being a leading predisposing factor. Although Aspergillus sp. was implicated in most of the patients in our study population, Candida sp. were found in higher numbers than previously reported. Keratitis caused by filamentous fungi responds adequately to medical management. Therapeutic keratoplasty continues to remain an important treatment modality in infections with Candida sp. Early diagnosis with prompt identification of the pathogenic organism is mandatory to initiate appropriate therapy and thereby reduce morbidity.