Department of Pathology and Diagnostics, Section of Microbiology, University of Verona, Verona, Italy.
J Clin Virol. 2013 Feb;56(2):124-8. doi: 10.1016/j.jcv.2012.10.015. Epub 2012 Nov 22.
Quantification of cytomegalovirus (CMV) DNA by real-time PCR is currently considered an alternative diagnostic approach for the evaluation of active infection in transplant patients. The pp65 antigenemia assay has been used as reference test for monitoring active CMV infection and guiding preemptive therapy in transplant recipients. However, this assay suffers from some limitations: need for immediate processing of the samples, labour-intensive process, lack of standardization and subjective result interpretation.
The aim of this study was to evaluate the performance of a new commercially available real-time PCR assay coupled with a fully automated DNA extraction system (COBAS Ampliprep/COBAS Taqman CMV Test, Roche Diagnostics) for the detection of CMV-DNA in plasma comparing it with pp65 antigenemia assay for monitoring active CMV infection in solid organ transplant recipients (SOTRs).
A total of 266 consecutive samples from 45 SOTRs were monitored with pp65 antigenemia and in parallel with CMV-DNA quantitation by real-time PCR assay.
Fifty-eight samples resulted PCR-positive, 163 negative and for 45 samples the CMV-DNA values obtained were below the lower limit of quantification (<150 copies/ml); pp65 antigen was detected in 47 samples and resulted negative in 219 specimens. Concordance between the two evaluations was 76.7%; also a good correlation was observed (r=0.718). Considering the existing treatment criteria based on pp65 antigenemia evaluation corresponding to pp65 levels≥20 positive cells/200,000, preemptive therapy was administered to four asymptomatically infected patients. The corresponding cut-off value of CMV-DNA load calculated for discrimination between self-clearing infections and those requiring therapy was 2500 copies/ml (or 2275 IU/ml).
The fully automated real-time PCR from Roche provided specific and sensitive results and represented a rapid and simple assay for the evaluation and monitoring of CMV infection in SOTRs. Further studies are required to validate the threshold level for the initiation of preemptive therapy.
实时聚合酶链反应（PCR）定量检测巨细胞病毒（CMV）DNA 目前被认为是评估移植患者活动性感染的一种替代诊断方法。pp65 抗原血症检测已被用作监测移植受者 CMV 感染和指导抢先治疗的参考检测。然而，该检测存在一些局限性：需要立即处理样本、劳动强度大、缺乏标准化和主观结果解释。
本研究旨在评估一种新的商业化实时 PCR 检测方法（罗氏诊断公司的 COBAS Ampliprep/COBAS Taqman CMV 检测）与全自动 DNA 提取系统相结合，用于检测血浆中的 CMV-DNA，与 pp65 抗原血症检测相比，评估实体器官移植受者（SOTR）中 CMV 感染的活性。
对 45 例 SOTR 的 266 例连续样本进行 pp65 抗原血症和实时 PCR 检测 CMV-DNA 定量监测。
58 例样本 PCR 阳性，163 例阴性，45 例样本的 CMV-DNA 值低于定量下限（<150 拷贝/ml）；47 例检测到 pp65 抗原，219 例结果阴性。两种评估方法的一致性为 76.7%；还观察到良好的相关性（r=0.718）。根据基于 pp65 抗原血症评估的现有治疗标准（对应于≥20 个阳性细胞/200,000 的 pp65 水平），对 4 例无症状感染患者进行了抢先治疗。计算出区分自限性感染和需要治疗的感染的 CMV-DNA 负荷的临界值为 2500 拷贝/ml（或 2275 IU/ml）。
罗氏全自动实时 PCR 提供了特异性和敏感性结果，是评估和监测 SOTR 中 CMV 感染的快速简便方法。需要进一步研究来验证抢先治疗开始的阈值水平。
