Polymerase chain reaction in the diagnosis of bacterial endophthalmitis

Abstract

Background: Microbiological investigations of vitreous fluid (VF) and aqueous humour (AH) specimens have often failed to detect the infecting agent in infectious endophthalmitis, resulting in a clinical dilemma regarding therapy. In this study, the polymerase chain reaction (PCR) was evaluated in the diagnosis of bacterial and Propionibacterium acnes endophthalmitis.

Methods: 58 intraocular specimens (30 VF and 28 AH) from 55 cases of endophthalmitis and 20 specimens (14 VF and 6 AH) as controls from non-infective disorders were processed for microbiological investigations. Nested PCR directed at the 16S rDNA using universal primers for eubacterial genome was done. PCR for P acnes was performed on specimens microbiologically negative by conventional techniques but eubacterial genome positive.

Results: Of the 20 controls from non-infective cases, one (5%) was positive using eubacterial primers and none with P acnes primers. PCR for eubacterial genome showed 100% correlation with 20 (34.5%) bacteriologically positive specimens. Eubacterial genome, was detected in 17 (44.7%) of 38 bacteriologically negative specimens and nine (52.9%) out of the 17 were positive for P acnes genome. Among the 21 eubacterial PCR negative specimens, seven were fungus positive. By inclusion of PCR, microbiologically positive specimens increased from 46.5% to 75.8%. PCR on AH was as sensitive as that on VF for the detection of both eubacterial and the P acnes genome.

Conclusion: PCR performed on AH and VF is a reliable tool for the diagnosis of bacterial and P acnes endophthalmitis particularly in smear and culture negative specimens.

Figure 1

Nested PCR results with universal eubacterial primers U2,rU3. Lanes 1, negative control (reagents); 2, negative control (sample extraction); 3, M* 1162/96 (VF) negative; 4, M 1111/96 (AH) positive; 5, M 1537/96 (VF) negative; 6, M 59/96 (VF) positive; 7, positive control; 8, molecular weight marker (Phi X 174/Hinf 1 Digest). *M denotes the laboratory identification numbers.

Figure 2

Nested PCR results with primers Pa1, rPa2. Lanes 1, negative control (reagents); 2, negative control (sample extraction); 3, M*1189/96 (AH) positive; 4, M 2512/95 negative (VF); 5, M 59/96 (VF) positive; 6, M 2964/96 (AH) negative; 7, positive control; 8, molecular weight marker (Phi X 174/Hinf 1 Digest). *M denote the laboratory identification numbers.