Polymerase Chain Reaction-guided Diagnosis of Infective Keratitis – A Hospital based Study

Abstract

PURPOSE: To compare polymerase chain reaction (PCR) to microbial culture and smear for detection and identification of bacterial and fungal pathogens in suspected microbial keratitis. MATERIALS AND METHODS: Corneal scrapings from 88 patients with suspected infectious keratitis were subjected to routine bacterial culture and sensitivity, Gram's stain, fungal culture; potassium hydroxide (KOH) wet mount, and PCR. PCR was performed with primer pairs targeted to the 16S and 18S r RNA gene. The result of the PCR was compared with conventional culture and Gram staining method. RESULTS: By broad-range PCR, 40 (45.45%) cases were positive for fungi (90.9% sensitivity), 26 (29.5%) were culture positive (59.09% sensitivity), 29 (33%) of all patients were positive for bacteria by broad-range PCR (87.9% sensitivity) and 19 (21.6%) were culture positive (57.58% sensitivity). The time taken for PCR assay was 4-8 h whereas positive fungal cultures took 2-10 days and bacterial culture from 2 to 4 days. Smears were positive for fungi in 29 eyes (33% of cases, 65.91% sensitivity) and for bacteria in 11 eyes (12.5% of cases, 33.33% sensitivity). CONCLUSIONS: DNA amplification with universal primers is a promising diagnostic tool in cases of infectious keratitis where routine laboratory culture failed to identify the pathogen. PCR may be performed in cases where the results of corneal scraping stains are negative without waiting for the results of the culture.