Permanent URL to this publication: http://dx.doi.org/10.5167/uzh-34540
Menassa, N; Bosshard, P P; Kaufmann, C; Grimm, C; Auffarth, G U; Thiel, M A (2010). Rapid detection of fungal keratitis using DNA-stabilizing FTA® filter paper. Investigative Ophthalmology and Visual Science, 51(4):1905-1910.
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Purpose. Polymerase chain reaction (PCR) is increasingly important for the rapid detection of fungal keratitis. However, techniques of specimen collection and DNA extraction before PCR may interfere with test sensitivity. The purpose of this study was to investigate the use of DNA-stabilizing FTA filter paper (Indicating FTA filter paper; Whatman International, Ltd., Maidstone, UK) for specimen collection without DNA extraction in a single-step, nonnested PCR for fungal keratitis. Methods. Specimens were collected from ocular surfaces with FTA filter discs, which automatically lyse collected cells and stabilize nucleic acids. Filter discs were directly used in single-step PCR reactions to detect fungal DNA. Test sensitivity was evaluated with serial dilutions of Candida albicans, Fusarium oxysporum, and Aspergillus fumigatus cultures. Test specificity was analyzed by comparing 196 and 155 healthy individuals from Switzerland and Egypt, respectively, with 15 patients with a diagnosis of microbial keratitis. Results. PCR with filter discs detected 3 C. albicans, 25 F. oxysporum, and 125 A. fumigatus organisms. In healthy volunteers, fungal PCR was positive in 1.0% and 8.4% of eyes from Switzerland and Egypt, respectively. Fungal PCR remained negative in 10 cases of culture-proven bacterial keratitis, became positive in 4 cases of fungal keratitis, but missed 1 case of culture-proven A. fumigatus keratitis. Conclusions. FTA filter paper for specimen collection together with direct PCR is a promising method of detecting fungal keratitis. The analytical sensitivity is high without the need for a semi-nested or nested second PCR, the clinical specificity is 91.7% to 99.0%, and the method is rapid and inexpensive.
|Item Type:||Journal Article, refereed, original work|
|Communities & Collections:||04 Faculty of Medicine > University Hospital Zurich > Ophthalmology Clinic|
04 Faculty of Medicine > University Hospital Zurich > Dermatology Clinic
|DDC:||610 Medicine & health|
|Deposited On:||05 Jul 2010 17:35|
|Last Modified:||23 Nov 2012 17:42|
|Publisher:||Association for Research in Vision and Ophthalmology|
|Funders:||Departmental Research Grant, Cantonal Hospital Lucerne|
|Free access at:||Publisher DOI. An embargo period may apply.|
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