Cytomegalovirus Positivity Predicts Graft Failure And Repeat Dmek: Prospective Development And Clinical Validation Of A Rapid Real-Time Pcr Assay

This study aims to investigate the role of cytomegalovirus (CMV) endotheliitis in graft failure after Descemet membrane endothelial keratoplasty (DMEK) and repeat DMEK (RE-DMEK). We also aimed to develop a highly sensitive real-time PCR kit for rapid CMV DNA detection, enabling early diagnosis and timely antiviral therapy to reduce graft failure risk.

This prospective study was conducted at the Cornea Unit, Department of Ophthalmology, Ankara Bilkent City Hospital, a tertiary referral center. Clinical examinations, surgical procedures, and follow-up assessments were standardized. Patients were evaluated at 3, 6, and 12 months postoperatively. Molecular analyses were performed in collaboration with an academic laboratory using standardized real-time PCR protocols to ensure analytical reliability and to assess clinical and diagnostic outcomes.

This prospective study included 160 patients undergoing DMEK (n=100) or repeat DMEK (n=60). Standardized ophthalmic examinations were performed pre- and postoperatively. Follow-up at 3, 6, and 12 months included best-corrected visual acuity (logMAR), intraocular pressure, central corneal thickness, and endothelial cell density. Anterior chamber samples were obtained intraoperatively or upon clinical suspicion of CMV endotheliitis. Samples were analyzed using a commercial real-time PCR system or the newly developed in-house assay. Diagnostic performance was assessed using cycle threshold values to determine sensitivity, specificity, and accuracy. Multivariate regression analysis was performed to identify independent risk factors.

No significant differences were observed between DMEK and RE-DMEK groups regarding age or sex. Graft survival was significantly lower in CMV-positive cases (p<0.001), with mean survival of 23.0 months in CMV-negative and 12.5 months in CMV-positive patients. CMV positivity was an independent predictor of repeat DMEK, conferring a sixfold increased risk (p<0.001). Rebubbling history was an additional independent risk factor. Visual acuity and endothelial cell density remained significantly worse in CMV-positive eyes during follow-up. The in-house PCR assay demonstrated 100% sensitivity and specificity, with overall diagnostic accuracy of 85.7%. CMV-positive eyes showed significantly shorter graft survival throughout follow-up.

CMV corneal endotheliitis is a major and underrecognized cause of graft failure after DMEK. Early differentiation from immune-mediated rejection is essential for timely antiviral therapy and improved graft survival. The newly developed in-house real-time PCR assay provides a rapid, reliable, and cost-effective diagnostic tool with high sensitivity and specificity. Its clinical implementation may support evidence-based postoperative management, reduce unnecessary immunosuppression, and potentially decrease repeat DMEK rates.