Development and application of multiplex polymerase chain reaction for the etiological diagnosis of infectious endophthalmitis.
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.


OBJECTIVE
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.


METHODS
Prospective study.


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.


CONCLUSIONS
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.
Bagyalakshmi
R
R
Larsen and Toubro Microbiology Research Centre, Vision Research Foundation, No.18, College Road, Sankara Nethralaya, Chennai-600006, Tamil Nadu, India.
Madhavan
H N
HN
Therese
K L
KL
D002176:Candida albicans / Q000235:genetics / Q000302:isolation & purification*; D003362:Cost-Benefit Analysis; D009877:Endophthalmitis / Q000175:diagnosis* / Q000382:microbiology*; D016680:Genome, Bacterial; D016681:Genome, Fungal; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011425:Propionibacterium acnes / Q000235:genetics / Q000302:isolation & purification*; D011446:Prospective Studies; D012680:Sensitivity and Specificity