UMI233, TransVIHMI, Institut de Recherche pour le Développement (IRD) and Université Montpellier 1, Montpellier, France.
J Clin Microbiol. 2013 Mar;51(3):787-98. doi: 10.1128/JCM.02792-12. Epub 2012 Dec 19.
Although antiretroviral treatment availability has improved, the virological monitoring of patients remains largely uneven across regions. In addition, viral quantification tests are suffering from human immunodeficiency virus type 1 (HIV-1) genetic diversity, fueled by the emergence of new recombinants and of lentiviruses from nonhuman primates. We developed a real-time reverse transcription-PCR (RT-PCR) assay that is relatively inexpensive and able to detect and quantify all circulating forms of HIV-1 and its simian immunodeficiency virus (SIV) precursors, SIVcpz and SIVgor. Primers and a probe were designed to detect all variants of the HIV-1/SIVcpz/SIVgor lineage. HIV-1 M plasma (n = 190; 1.68 to 7.78 log(10) copies/ml) representing eight subtypes, nine circulating recombinant forms, and 21 unique recombinant forms were tested. The mean PCR efficiency was 99%, with low coefficients of intra- and interassay variation (<5%) and a limit of quantification of <2.50 log(10) copies/ml, with a 200-μl plasma volume. On the studied range, the specificity and the analytical sensitivity were 100 and 97.4%, respectively. The viral loads were highly correlated (r = 0.95, P < 0.0001) with the reference method (generic HIV assay; Biocentric) and had no systematic difference, irrespective of genotype. Furthermore, 22 HIV-1 O plasmas were screened and were better quantified compared to the gold-standard RealTime HIV-1 assay (Abbott), including four samples that were only quantified by our assay. Finally, we could quantify SIVcpzPtt and SIVcpzPts from chimpanzee plasma (n = 5) and amplify SIVcpz and SIVgor from feces. Thus, the newly developed real-time RT-PCR assay detects and quantifies strains from the HIV-1/SIVcpz/SIVgor lineage, including a wide diversity of group M strains and HIV-1 O. It can therefore be useful in geographical areas of high HIV diversity and at risk for the emergence of new HIV variants.
尽管抗逆转录病毒治疗的可及性有所提高，但患者的病毒学监测在各地区仍存在很大差异。此外，由于新的重组体和来自非人类灵长类动物的慢病毒的出现，病毒定量检测受到人类免疫缺陷病毒 1 型（HIV-1）遗传多样性的影响。我们开发了一种实时逆转录 PCR（RT-PCR）检测方法，该方法相对便宜，能够检测和定量所有循环形式的 HIV-1 及其猴免疫缺陷病毒（SIV）前体，SIVcpz 和 SIVgor。设计了引物和探针来检测 HIV-1/SIVcpz/SIVgor 谱系的所有变体。检测了代表 8 种亚型、9 种循环重组形式和 21 种独特重组形式的 HIV-1 M 血浆（n=190；1.68 至 7.78 log(10) 拷贝/ml）。PCR 效率的平均值为 99%，内和间试验变异系数低（<5%），定量下限<2.50 log(10) 拷贝/ml，检测体积为 200 μl。在研究范围内，特异性和分析灵敏度分别为 100%和 97.4%。病毒载量与参考方法（通用 HIV 检测；Biocentric）高度相关（r=0.95，P<0.0001），且不论基因型如何，均无系统差异。此外，筛选了 22 份 HIV-1 O 血浆，与金标准 RealTime HIV-1 检测（Abbott）相比，定量效果更好，包括我们的检测方法唯一定量的 4 个样本。最后，我们可以从黑猩猩血浆（n=5）中定量检测 SIVcpzPtt 和 SIVcpzPts，并从粪便中扩增 SIVcpz 和 SIVgor。因此，新开发的实时 RT-PCR 检测方法可检测和定量 HIV-1/SIVcpz/SIVgor 谱系的病毒株，包括广泛的 M 株和 HIV-1 O 株。因此，它可用于 HIV 多样性高的地区和出现新 HIV 变体的风险地区。
