Specific PCR and Quantitative Real-Time PCR in Ocular Samples from Acute and Delayed-Onset Postoperative Endophthalmitis

Abstract

Rapid identification of virulent pathogens is essential to strengthen the therapeutic strategy of acute endophthalmitis. This study sought to compare the contribution of a combination of polymerase chain reaction (PCR)-based tests to culture methods, in patients with postoperative endophthalmitis. Prospective multicenter study diagnostic evaluation. Setting: university referral centers. 153 consecutive patients presenting with acute or delayed-onset postoperative endophthalmitis, between 2008 and 2015. There were a total of 284 aqueous humor (AH) and/or vitreous fluid (VF) samples. Outcomes and measurements: microbiological tests of intraocular samples included bacterial culturing of pediatric blood culture bottles; 16SrDNA amplification and sequencing (panbacterial PCR) for detection and identification of all bacterial species; real-time PCR (qPCR) assays targeting the femA or lytA gene for detection of Staphylococcus aureus (S. aureus) or Streptococcus pneumoniae (S. pneumoniae), respectively; and a qPCR assay targeting the tuf gene for detection and quantification of Staphylococcus epidermidis (S. epidermidis). At the time of admission, the rate of detection of microorganisms by PCR-based tests was not significantly different than that by culturing (38% versus 30% in AH samples [n= 69]; 66% versus 63% in VF samples [n= 82], respectively). In contrast, after 1 intravitreal injection (IVI) of antibiotics, the identification rate by PCR-based tests was higher than that in VF by culturing (62% vs 48%, respectively; n= 94; P= 0.05). Bacteria were identified in 70% of patients, with a predominance of Gram-positive bacteria (93%). Specific qPCR tests targeting S. aureus and S. pneumoniae did not provide additional diagnoses but provided earlier results. The S. epidermidis load in vitreous at the time of patients' admission was higher in cases of final visual acuity (VA) of <20/40 (127,118 ± 125,848 DNA copies/mL) in patients with a VA of ≥20/40 (40350,000 ± 46,912 DNA copies/mL; P= 0.09). No significant changes in S. epidermidis load was found after one IVI. Patients with acute or delayed-onset endophthalmitis should benefit from microbiological identification in vitreous samples by combined analysis using bacterial cultures in pediatric blood culture bottles and panbacterial PCR. The last test was more effective than cultures in vitreous samples collected after an IVI of antibiotics. The qPCR tests targeting S. aureus and S. pneumoniae gave earlier results than culture and panbacterial PCR but did not provide additional diagnoses. As for S. epidermidis infections, determination of bacterial load using the qPCR test targeting the tuf gene could help evaluation of the visual prognosis of patients. Its role in the follow-up of patients after antibiotic treatment needs further investigation.