Multicentric evaluation of a new commercial cytomegalovirus real-time PCR quantitation assay

Automated real-time PCR systems have become the most common method in the quantitation of viral load during cytomegalovirus (CMV) infection in immuno-compromised patients. In order to evaluate a new commercially available CMV real-time PCR assay (CMV R-gene (TM), Argene, France), a pp65 antigenemia assay and four different "in-house" real-time PCR assays were compared to the CMV R-gene (TM) for the detection and the quantitation of CMV load in 506 specimens of whole blood from transplant patients in four French hospital laboratories. The CMV R-gene (TM) was more sensitive than the pp65 antigenemia: there were 18% antigenemia-negative versus CMV R-gene (TM) -positive samples. A significant correlation was found between DNA quantitation by CMV R-gene (TM) and the number of positive cells detected by the pp65 a test (Spearman's rank test, r= 0.63, p < 0.0001). A CMV DNA load equivalent to 50 pp65-positive cells/200 000 polymorphonuclear leukocytes was 5.26 log(10) copies/mL of whole blood. When the CMV R-gene (TM) kit was compared to the four other "in-house" real-time PCR assays, there were few discordant results (6.7% total for the four laboratories), all detected with a weak positive CMV DNA viral load. Spearman's coefficients showed a good (r = 0.82 for laboratory 1, r = 0.66 for laboratory 3) to excellent (r = 0.99 for laboratory 2, r = 0.94 for laboratory 4) correlation between CMV R-gene (TM) and the four real-time "in-house" PCR assays. However, the results of CMV DNA viral load generated by CMV R-gene (TM) test were constantly higher than those generated by three out of four "in-house" PCR assays. This mean variation in CMV DNA viral load measured by CMV R-gene (TM) and "in-house" PCRs was of 0.77 log(10), 0.04 log(10), 0.77 log(10) and 0.97 log(10), for laboratories 1, 2, 3 and 4, respectively. We concluded that there was variability between results of different real-time PCR assays for CMV DNA quantitation. This observation emphasized the need of a standardised commercial assay to allow an "inter-laboratory" comparison of results. Our study showed that CMV R-gene (TM) is an accurate, efficient, reliable and versatile too] for rapid diagnosis and monitoring of CMV disease in transplantation recipients. (c) 2007 Elsevier B.V. All rights reserved.