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

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          Abstract

          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.

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          Multistate outbreak of Fusarium keratitis associated with use of a contact lens solution.

          Fusarium keratitis is a serious corneal infection, most commonly associated with corneal injury. Beginning in March 2006, the Centers for Disease Control and Prevention received multiple reports of Fusarium keratitis among contact lens wearers. To define the specific activities, contact lens hygiene practices, or products associated with this outbreak. Epidemiological investigation of Fusarium keratitis occurring in the United States. A confirmed case was defined as keratitis with illness onset after June 1, 2005, with no history of recent ocular trauma and a corneal culture growing Fusarium species. Data were obtained by patient and ophthalmologist interviews for case patients and neighborhood-matched controls by trained personnel. Available Fusarium isolates from patients' clinical and environmental specimens were genotyped by multilocus sequence typing. Environmental sampling for Fusarium was conducted at a contact lens solution manufacturing plant. Keratitis infection with Fusarium species. As of June 30, 2006, we identified 164 confirmed case patients in 33 states and 1 US territory. Median age was 41 years (range, 12-83 years). Corneal transplantation was required or planned in 55 (34%). One hundred fifty-four (94%) of the confirmed case patients wore soft contact lenses. Forty-five case patients and 78 controls were included in the case-control study. Case patients were significantly more likely than controls to report using a specific contact lens solution, ReNu with MoistureLoc (69% vs 15%; odds ratio, 13.3; 95% confidence interval, 3.1-119.5). The prevalence of reported use of ReNu MultiPlus solution was similar between case patients and controls (18% vs 20%; odds ratio, 0.7; 95% confidence interval, 0.2-2.8). Fusarium was not recovered from the factory, warehouse, solution filtrate, or unopened solution bottles; production of implicated lots was not clustered in time. Among 39 isolates tested, at least 10 different Fusarium species were identified, comprising 19 unique multilocus genotypes. The findings from this investigation indicate that this outbreak of Fusarium keratitis was associated with use of ReNu with MoistureLoc contact lens solution. Contact lens users should not use ReNu with MoistureLoc.
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            Current perspectives on ophthalmic mycoses.

            J. Thomas (2003)
            Fungi may infect the cornea, orbit and other ocular structures. Species of Fusarium, Aspergillus, Candida, dematiaceous fungi, and Scedosporium predominate. Diagnosis is aided by recognition of typical clinical features and by direct microscopic detection of fungi in scrapes, biopsy specimens, and other samples. Culture confirms the diagnosis. Histopathological, immunohistochemical, or DNA-based tests may also be needed. Pathogenesis involves agent (invasiveness, toxigenicity) and host factors. Specific antifungal therapy is instituted as soon as the diagnosis is made. Amphotericin B by various routes is the mainstay of treatment for life-threatening and severe ophthalmic mycoses. Topical natamycin is usually the first choice for filamentous fungal keratitis, and topical amphotericin B is the first choice for yeast keratitis. Increasingly, the triazoles itraconazole and fluconazole are being evaluated as therapeutic options in ophthalmic mycoses. Medical therapy alone does not usually suffice for invasive fungal orbital infections, scleritis, and keratitis due to Fusarium spp., Lasiodiplodia theobromae, and Pythium insidiosum. Surgical debridement is essential in orbital infections, while various surgical procedures may be required for other infections not responding to medical therapy. Corticosteroids are contraindicated in most ophthalmic mycoses; therefore, other methods are being sought to control inflammatory tissue damage. Fungal infections following ophthalmic surgical procedures, in patients with AIDS, and due to use of various ocular biomaterials are unique subsets of ophthalmic mycoses. Future research needs to focus on the development of rapid, species-specific diagnostic aids, broad-spectrum fungicidal compounds that are active by various routes, and therapeutic modalities which curtail the harmful effects of fungus- and host tissue-derived factors.
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              Detection and identification of fungal pathogens by PCR and by ITS2 and 5.8S ribosomal DNA typing in ocular infections.

              The goal of this study was to determine whether sequence analysis of internal transcribed spacer/5.8S ribosomal DNA (rDNA) can be used to detect fungal pathogens in patients with ocular infections (endophthalmitis and keratitis). Internal transcribed spacer 1 (ITS1) and ITS2 and 5.8S rDNA were amplified by PCR and seminested PCR to detect fungal DNA. Fifty strains of 12 fungal species (yeasts and molds) were used to test the selected primers and conditions of the PCR. PCR and seminested PCR of this region were carried out to evaluate the sensitivity and specificity of the method. It proved possible to amplify the ITS2/5.8S region of all the fungal strains by this PCR method. All negative controls (human and bacterial DNA) were PCR negative. The sensitivity of the seminested PCR amplification reaction by DNA dilutions was 1 organism per PCR, and the sensitivity by cell dilutions was fewer than 10 organisms per PCR. Intraocular sampling or corneal scraping was undertaken for all patients with suspected infectious endophthalmitis or keratitis (nonherpetic), respectively, between November 1999 and February 2001. PCRs were subsequently performed with 11 ocular samples. The amplified DNA was sequenced, and aligned against sequences in GenBank at the National Institutes of Health. The results were PCR positive for fungal primers for three corneal scrapings, one aqueous sample, and one vitreous sample; one of them was negative by culture. Molecular fungal identification was successful in all cases. Bacterial detection by PCR was positive for three aqueous samples and one vitreous sample; one of these was negative by culture. Amplification of ITS2/5.8S rDNA and molecular typing shows potential as a rapid technique for identifying fungi in ocular samples.
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                Author and article information

                Contributors
                Journal
                Biomed Res Int
                Biomed Res Int
                BMRI
                BioMed Research International
                Hindawi
                2314-6133
                2314-6141
                2019
                20 January 2019
                : 2019
                : 6395840
                Affiliations
                1Department of Medical Laboratory Sciences, College of Applied Medical Sciences, Majmaah University, Al Majmaah, Saudi Arabia
                2Aravind Eye Hospital and Postgraduate Institute of Ophthalmology, Coimbatore, India
                3Greenlink Analytical and Research Laboratory India Private Limited, Coimbatore, India
                4Botany and Microbiology Department, Faculty of Science, Al-Azhar University, Assiut Branch, Egypt
                5Microbial Resources Division, Institute of Bioresources and Sustainable Development, Takyel, Imphal, Manipur, India
                6Department of Microbiology, PSG College of Arts and Science, Coimbatore, India
                7Department of Microbiology, M.R Government Arts College, Mannargudi, Tamilnadu, India
                Author notes

                Academic Editor: Macit Ilkit

                Author information
                http://orcid.org/0000-0002-9636-4006
                http://orcid.org/0000-0002-1933-8197
                http://orcid.org/0000-0002-4423-8892
                Article
                10.1155/2019/6395840
                6360544
                30800674
                cb5bbe79-ff12-461c-92de-0f4e130d8fd6
                Copyright © 2019 Palanisamy Manikandan et al.

                This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 31 October 2018
                : 10 December 2018
                Funding
                Funded by: Indian Council of Medical Research
                Award ID: 2008-03140
                Funded by: Kingdom of Saudi Arabia
                Award ID: 38/128
                Categories
                Research Article

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