Clearance of low-level viremia that persists in most HIV-1-positive individuals on antiretroviral therapy (ART) is an important milestone for efforts to cure HIV-1 infection. The level of persistent viremia on ART is generally below the lower limit of quantification (LOQ) of current FDA-cleared plasma HIV-1 RNA assays (20 to 40 copies/ml) but can be quantified by reverse transcriptase PCR (RT-PCR) assays with single-copy sensitivity. Such assays require multistep manual methods, and their low throughput limits the capacity to monitor the effects of interventions on persistent viremia. Recently, S. Bakkour, X. Deng, P. Bacchetti, E. Grebe, et al. (J Clin Microbiol 58:e01400-20, 2020, https://doi.org/10.1128/JCM.01400-20), reported the use of multiple replicates and Poisson statistics to infer HIV-1 RNA concentrations below the commercial LOQ of an automated platform (Hologic Panther Aptima). Here, we evaluate the detection and quantitation of low-level viremia using the following two adaptions of the automated platform: a multireplicate strategy (9×) and a concentrated single-replicate strategy in which 5 ml of plasma is concentrated by centrifugation (1×, concentrated). We compare these new methods to a recently reported manual integrase-targeting single-copy assay version 2 (iSCA v2). Using laboratory-generated HIV-1 RNA plasma samples at known concentrations, all three methods had similar sensitivity for HIV-1 RNA detection, although iSCA v2 was most sensitive (95% LOD, 2.3 copies/ml), 9× was marginally less sensitive (95% LOD, 3.0 copies/ml), and 1×, concentrated was least sensitive (95% LOD, 3.9 copies/ml). In contrast, for clinical plasma samples, 9× had greater sensitivity than iSCA v2 (82% of samples were quantifiable compared with 62% of samples by iSCA v2). These results support 9× as an acceptable high-throughput alternative to iSCA v2 for quantifying low-level viremia in individuals on ART.

中文翻译：

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进行抗逆转录病毒疗法治疗的个体中持久性HIV-1病毒血症的自动化，多重复定量分析。

在大多数接受抗逆转录病毒疗法（ART）的HIV-1阳性个体中清除低水平病毒血症是努力治疗HIV-1感染的重要里程碑。ART上的持续病毒血症水平通常低于当前FDA批准的血浆HIV-1 RNA检测的定量下限（LOQ）（20至40拷贝/ ml），但可以通过逆转录酶PCR（RT-PCR）进行定量单拷贝灵敏度的检测。这样的测定需要多步手动方法，并且它们的低通量限制了监测干预对持续病毒血症的影响的能力。最近，S。Bakkour，X。Deng，P。Bacchetti，E。Grebe等人。（J Clin Microbiol 58：e01400-20，2020，https://doi.org/10.1128/JCM.01400-20），报道了使用多次重复和Poisson统计来推断HIV-1 RNA浓度低于自动化平台（Hologic Panther Aptima）的商业LOQ。在这里，我们使用以下两种自动平台改型来评估低水平病毒血症的检测和定量：多重复制策略（9x）和浓缩的单一复制策略，其中5 ml血浆通过离心浓缩（1x ， 集中）。我们将这些新方法与最近报告的手册进行了比较整合酶靶向单拷贝测定法版本2（iSCA v2）。尽管iSCA v2最敏感（95％LOD，2.3拷贝/ ml），9×灵敏度稍低，但使用实验室已知浓度的HIV-1 RNA血浆样品，这三种方法对HIV-1 RNA的检测灵敏度均相似。 （95％LOD，3.0拷贝/ ml）和1x浓缩最不敏感（95％LOD，3.9拷贝/ ml）。相反，对于临床血浆样品，9x的灵敏度高于iSCA v2（82％的样品可定量，而iSCA v2的样品为62％）。这些结果支持9x作为iSCA v2可接受的高通量替代品，用于量化ART个体中的低水平病毒血症。