R Bagyalakshmi1, H N Madhavan, K L Therese. 1. Larsen and Toubro Microbiology Research Centre, Vision Research Foundation, No.18, College Road, Sankara Nethralaya, Chennai-600006, Tamil Nadu, India.
Abstract
BACKGROUND: Uniplex polymerase chain reaction (PCR) for detection of bacterial and panfungal genome has been applied onto a large number of intraocular fluids facilitating management of infective endophthalmitis. AIM: To develop and apply a novel, rapid multiplex polymerase chain reaction (mPCR) to detect the presence of eubacterial, Propionibacterium acnes and panfungal genomes in intraocular fluids from patients clinically diagnosed to have infective endophthalmitis. SETTINGS AND DESIGN: Prospective study. MATERIALS AND METHODS: Conventional methods of direct microscopy by KOH/calcofluor mount, Gram's staining and culture were done on 30 (19 Aqueous humor-AH and 11 Vitreous fluid-VF) intraocular specimens and mPCR done for simultaneous detection of eubacterial, P. acnes and panfungal genomes. RESULTS: mPCR detected an infectious etiology in 18 (60%) of 30 intraocular specimens. Eubacterial genome was detected in 12 (40%) specimens, P. acnes genome in 4 (13.3%) specimens and panfungal genome in 2 (6.6%) specimens. mPCR results correlated with those of uniplex PCR. mPCR results were available within 5-6 hours after receipt of specimen, as against 8 hours required for each uniplex PCR with three separate thermalcyclers for their completion. Consumption of Taq polymerase was reduced considerably for mPCR. CONCLUSION: mPCR is a cost effective, single tube method for the simultaneous detection of eubacterial, P. acnes and panfungal genomes in intraocular specimens from patients with infective endophthalmitis. It is a more rapid procedure than uniplex PCRs and requires only a single thermalcycler.
BACKGROUND: Uniplex polymerase chain reaction (PCR) for detection of bacterial and panfungal genome has been applied onto a large number of intraocular fluids facilitating management of infective endophthalmitis. AIM: To develop and apply a novel, rapid multiplex polymerase chain reaction (mPCR) to detect the presence of eubacterial, Propionibacterium acnes and panfungal genomes in intraocular fluids from patients clinically diagnosed to have infective endophthalmitis. SETTINGS AND DESIGN: Prospective study. MATERIALS AND METHODS: Conventional methods of direct microscopy by KOH/calcofluor mount, Gram's staining and culture were done on 30 (19 Aqueous humor-AH and 11 Vitreous fluid-VF) intraocular specimens and mPCR done for simultaneous detection of eubacterial, P. acnes and panfungal genomes. RESULTS: mPCR detected an infectious etiology in 18 (60%) of 30 intraocular specimens. Eubacterial genome was detected in 12 (40%) specimens, P. acnes genome in 4 (13.3%) specimens and panfungal genome in 2 (6.6%) specimens. mPCR results correlated with those of uniplex PCR. mPCR results were available within 5-6 hours after receipt of specimen, as against 8 hours required for each uniplex PCR with three separate thermalcyclers for their completion. Consumption of Taq polymerase was reduced considerably for mPCR. CONCLUSION: mPCR is a cost effective, single tube method for the simultaneous detection of eubacterial, P. acnes and panfungal genomes in intraocular specimens from patients with infective endophthalmitis. It is a more rapid procedure than uniplex PCRs and requires only a single thermalcycler.
Authors: Rubens C Vilela; Luiza Vilela; Priscila Vilela; Raquel Vilela; Roberta Motta; Ana Paula Pôssa; Cirênio de Almeida; Leonel Mendoza Journal: Int Ophthalmol Date: 2013-10-01 Impact factor: 2.031
Authors: Silvana Maria de Morais Cavalcanti; Emmanuel Rodrigues de França; Marcelo Magalhães; Ana Kelly Lins; Laura Costa Brandão; Vera Magalhães Journal: Braz J Microbiol Date: 2011-06-01 Impact factor: 2.476