U.S. Military HIV Research Program, Walter Reed Army Institute of Research, Silver Spring, Maryland, United States of America.
U.S. Military HIV Research Program, Henry M. Jackson Foundation for the Advancement of Military Medicine, Bethesda, Maryland, United States of America.
PLoS One. 2020 Feb 28;15(2):e0229424. doi: 10.1371/journal.pone.0229424. eCollection 2020.
Management of Human Immunodeficiency Virus Type 2 (HIV-2) infections present unique challenges due to low viral titers, slow disease progression, and poor response to standard antiviral therapies. The need for a nucleic acid assay to detect and quantify HIV-2 virus has led to the development of a number of molecular-based assays for detection and/or quantification of HIV-2 viral RNA in plasma in order to provide laboratory evidence of HIV-2 infection and viral loads for use in treatment decisions. As HIV-2 is less pathogenic and transmissible than HIV-1 and has resistance to several of the antiretroviral drugs, delay of treatment is common. Cross sero-reactivity between HIV-1 and HIV-2 makes it difficult to distinguish between the two viruses based upon serological tests. As such we developed a quantitative reverse transcription PCR (qRT-PCR) assay targeting the 5' long terminal repeat of HIV-2 for detection and quantification of HIV-2 viral RNA in plasma to identify HIV-2 infection and for use in viral load monitoring. Serial dilutions of cultured HIV-2 virus demonstrated a wide dynamic range (10 to 100,000 copies/ml) with excellent reproducibility (standard deviation from 0.12-0.19), linearity (R2 = 0.9994), and a lower limit of detection at 79 copies/ml (NIH-Z). The assay is highly specific for HIV-2 Groups A and B and exhibits no cross reactivity to HIV-1, HBV or HCV. Precision of the assay was demonstrated for the High (Mean = 6.41; SD = 0.12) and Medium (Mean = 4.46; SD = 0.13) HIV-2 positive controls. Replicate testing of clinical specimens showed good reproducibility above 1,000 copies/ml, with higher variability under 1,000 copies/ml. Analysis of 220 plasma samples from HIV-2 infected West African individuals demonstrated significantly lower viral loads than those observed in HIV-1 infections, consistent with results of previous studies. Slightly more than seven percent of clinical samples (7.3%) demonstrated viral loads above 100,000 copies/ml, while 37.3% of samples were undetectable. The high sensitivity, specificity, precision, and linearity of the WRAIR qRT-PCR assay makes it well suited for detection and monitoring of HIV-2 RNA levels in plasma of infected individuals.
由于 HIV-2 的病毒载量低、疾病进展缓慢以及对标准抗病毒疗法的反应不佳，管理 HIV-2 感染带来了独特的挑战。为了检测和定量 HIV-2 病毒，需要一种核酸检测方法，这导致了许多基于分子的检测方法的发展，用于检测和/或定量血浆中的 HIV-2 病毒 RNA，以便为 HIV-2 感染和病毒载量提供实验室证据，用于治疗决策。由于 HIV-2 的致病性和传染性低于 HIV-1，并且对几种抗逆转录病毒药物具有耐药性，因此延迟治疗很常见。HIV-1 和 HIV-2 之间的交叉血清学反应使得基于血清学检测难以区分这两种病毒。因此，我们开发了一种针对 HIV-2 5'长末端重复序列的定量逆转录 PCR（qRT-PCR）检测方法，用于检测和定量血浆中的 HIV-2 病毒 RNA，以识别 HIV-2 感染并用于病毒载量监测。培养的 HIV-2 病毒的系列稀释显示出广泛的动态范围（10 至 100,000 拷贝/ml），具有极好的重现性（标准偏差为 0.12-0.19）、线性（R2 = 0.9994）和检测下限为 79 拷贝/ml（NIH-Z）。该检测方法对 HIV-2 组 A 和 B 高度特异性，对 HIV-1、HBV 或 HCV 无交叉反应。高（Mean = 6.41；SD = 0.12）和中（Mean = 4.46；SD = 0.13）HIV-2 阳性对照的检测精密度得到了证明。对临床标本的重复检测显示，在 1,000 拷贝/ml 以上的复制性良好，在 1,000 拷贝/ml 以下的复制性较差。对来自感染 HIV-2 的西非个体的 220 份血浆样本的分析表明，病毒载量明显低于 HIV-1 感染观察到的病毒载量，这与之前的研究结果一致。略多于 7%的临床样本（7.3%）的病毒载量超过 100,000 拷贝/ml，而 37.3%的样本无法检测到。WRAIR qRT-PCR 检测方法的高灵敏度、特异性、精密度和线性使其非常适合于检测和监测感染个体血浆中的 HIV-2 RNA 水平。
