Novel Use of Fluorescence In Situ Hybridization for the Rapid Identification of Microorganisms in Endophthalmitis and Keratitis

BACKGROUND AND OBJECTIVE: To evaluate an application of fluorescence in situ hybridization (FISH) for the rapid identification of bacterial and fungal pathogens causing endophthalmitis and keratitis and compare time to detection with other laboratory methods. MATERIALS AND METHODS: Culture-positive isolates obtained from vitreous and corneal samples were tested. Organisms tested were Staphylococcus aureus, coagulase-negative staphylococci, Pseudomonas aeruginosa, Candida albicans, C. glabrata, and C. parapsilosis. Inoculi were prepared to a final concentration between 1 × 102 colony-forming units (CFU)/mL to 1 × 108 CFU/mL. Samples were hybridized with peptide nucleic acid probes for pathogens using the QuickFISH protocol (AdvanDx; OpGen, Gaithersburg, MD), and the slides were read with fluorescence microscopy. RESULTS: Of the 29 total isolates tested, 28 yielded positive identification. S. aureus was identified in four out of five vitreous samples, whereas coagulase-negative staphylococci were identified in all vitreous samples. Mixed staphylococci culture was identified in all samples. P. aeruginosa was identified in all six keratitis samples. C. albicans, C. glabrata, C. parapsilosis, and mixed fungal culture were identified respectively in eight of eight samples at minimal concentration of 1 × 104 CFU/mL. There were no false-negatives. Time to detection was 20 minutes after the 12- to 18-hour inoculation period and provided an identification 6 hours sooner than by polymerase chain reaction (PCR) and 1 to 2 days sooner than by routine culture. CONCLUSIONS: This small study demonstrates the sensitive, specific, and rapid detection of gram-positive bacteria, gram-negative bacteria and fungi using FISH probes in isolates from endophthalmitis and keratitis samples. This method decreases time to identification and reduces labor intensity compared with routine PCR and culture methods. [Ophthalmic Surg Lasers Imaging Retina. 2019;50:S9-S12.].