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应用 PCR 联合质谱分析技术检测和半定量分析特发性扩张型心肌病成年患者的移植心脏组织中的病毒。

Virus detection and semiquantitation in explanted heart tissues of idiopathic dilated cardiomyopathy adult patients by use of PCR coupled with mass spectrometry analysis.

机构信息

Clinical and Molecular Virology Unit, University Hospital, and EA-4684 Cardiovir SFR-CAP Sante, Faculty of Medicine, Reims, France.

出版信息

J Clin Microbiol. 2013 Jul;51(7):2288-94. doi: 10.1128/JCM.00820-13. Epub 2013 May 8.

Abstract

Viral detection in heart tissues has become a central issue for the diagnosis and exploration of the pathogenesis of idiopathic dilated cardiomyopathy (IDCM). In the present study, common cardiotropic viruses in 67 explanted heart samples of 31 IDCM adult patients were detected and semiquantified by using for the first time a new technology based on PCR assay coupled to electrospray ionization-time of flight mass spectrometry analysis (PCR-MS), with comparison to reference quantitative real-time PCR (RT-qPCR) assay. PCR-MS identified single or mixed enterovirus (EV) and parvovirus B19 (PVB19) infections in 27 (40.2%) of 67 samples, corresponding to 15 (48.3%) of the 31 patients, whereas RT-qPCR identified viral infections in 26 (38.8%) samples, corresponding to 16 (51.6%) of the patients. The PCR-MS results correlated well with EV and PVB19 detection by RT-qPCR (kappa = 0.85 [95% confidence interval {CI}, 0.72 to 1.00] and kappa = 0.82 [95% CI, 0.66 to 0.99], respectively). The levels of EV RNA (median, 550 [range, 178 to 3,200] copies/μg of total extracted nucleic acids) and of PVB19 DNA (median, 486 [range, 80 to 1,157] copies/μg of total extracted nucleic acids) were measured using PCR-MS and correlated with those obtained by RT-qPCR (r(2) = 0.57, P = 0.002 and r(2) = 0.64, P < 0.001 for EV and PVB19, respectively). No viruses other than EV and PVB19 strains were detected using the new PCR-MS technology, which is capable of simultaneously identifying 84 known human viruses in one assay. In conclusion, we identified single or mixed EV and PVB19 cardiac infections as potential causes of IDCM. The PCR-MS analysis appeared to be a valuable tool to rapidly detect and semiquantify common viruses in cardiac tissues and may be of major interest to better understand the role of viruses in unexplained cardiomyopathies.

摘要

病毒在心脏组织中的检测已成为诊断和探讨特发性扩张型心肌病(IDCM)发病机制的核心问题。本研究首次采用基于 PCR 分析与电喷雾电离-飞行时间质谱分析(PCR-MS)相结合的新技术,检测了 31 例 IDCM 成人患者 67 例心脏标本中的常见心脏病毒,并进行了半定量分析。与参考定量实时 PCR(RT-qPCR)检测相比,PCR-MS 鉴定出 67 例样本中有 27 例(40.2%)存在单纯或混合肠道病毒(EV)和微小病毒 B19(PVB19)感染,31 例患者中有 15 例(48.3%)符合上述情况,而 RT-qPCR 鉴定出 26 例(38.8%)样本存在病毒感染,31 例患者中有 16 例(51.6%)符合上述情况。PCR-MS 结果与 EV 和 PVB19 的 RT-qPCR 检测结果相关性良好(kappa = 0.85 [95%置信区间{CI},0.72 至 1.00]和 kappa = 0.82 [95% CI,0.66 至 0.99])。使用 PCR-MS 测量了 EV RNA(中位数,550 [范围,178 至 3200]拷贝/μg 总提取核酸)和 PVB19 DNA(中位数,486 [范围,80 至 1157]拷贝/μg 总提取核酸)的水平,并与 RT-qPCR 测量结果相关(EV 和 PVB19 的 r(2)分别为 0.57,P = 0.002 和 r(2)为 0.64,P < 0.001)。除 EV 和 PVB19 株外,使用新的 PCR-MS 技术未检测到其他病毒,该技术能够在一次检测中同时鉴定出 84 种已知的人类病毒。总之,我们鉴定出单纯或混合 EV 和 PVB19 心脏感染可能是 IDCM 的潜在原因。PCR-MS 分析似乎是一种快速检测和半定量分析心脏组织中常见病毒的有价值工具,可能对更好地了解病毒在不明原因的心肌病中的作用具有重要意义。

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