Development and Evaluation of a Real-Time TaqMan-PCR for the Detection of Human Cytomegalovirus DNA in Bone Marrow Transplant Recipients

Introduction: Cytomegalovirus (CMV) has been recognized as the most important viral pathogen in persons undergoing bone marrow transplantation (BMT). In this study, we present the development of a TaqMan-based real-time PCR assay to quantify human cytomegalovirus (CMV) DNA in peripheral blood leukocytes (PBLs) of bone marrow transplantation patients. Materials and Methods: A plasmid containing the target sequence from the pp65 region (UL83) of CMV was constructed as a positive control template. Serial dilutions of 107 to 101 plasmids per assay were prepared. Peripheral blood samples were collected from patients after transplantation. CMV DNA was quantified by RQ-PCR in parallel with the pp65 antigenemia assay in PBL samples. Results: The real-time PCR assay could detect CMV DNA in patient's samples with a wide linear range, from 10 to over 107 copies of CMV. Real-time PCR assay results correlated with those of the CMV pp65 antigenemia assay (P < 0.0001). Discussion: The TaqMan assay may be a useful tool for rapid quantification of CMV infection and for monitoring of CMV reactivation in bone marrow transplantation recipients. The results of both quantitative assays were significantly correlated; however, the RQ-PCR assay was more sensitive than the pp65 antigenemia assay.