Quantification of genital human immunodeficiency virus type 1 (HIV-1) DNA in specimens from women with low plasma HIV-1 RNA levels typical of HIV-1 nontransmitters
Quantification of genital human immunodeficiency virus type 1 (HIV-1) DNA in specimens from women with low plasma HIV-1 RNA levels typical of HIV-1 nontransmitters
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DOI：
10.1128/jcm.01481-06
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发表时间：
2006-12-01
影响因子：
9.4
通讯作者：
Overbaugh, Julie
中科院分区：
文献类型：
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作者：
Benki, Sarah;McClelland, R. Scott;Overbaugh, Julie
人类免疫缺陷病毒1型（HIV-1）传播的研究表明，生殖器HIV-1 RNA和DNA可能都是HIV-1感染性的决定因素。尽管它在HIV-1传播中具有潜在作用，但关于生殖器HIV-1 DNA的定量数据有限。在这里，我们验证了一种内部实时PCR方法，用于定量生殖器标本中的HIV-1 DNA。在与从含有一个HIV-1前病毒/细胞的细胞系中分离的100个基因组对1个基因组的反应中，该实时PCR测定是线性的，并且与特异于细胞管家基因的市售实时PCR测定密切一致。在加标少量HIV-1感染细胞（预期HIV-1 DNA拷贝数/反应为100、10或5）的模拟生殖器样本中，平均拷贝数/反应分别为80.2（标准差[SD]，28.3）、9.1（SD，5.4）或3.1（SD，2.1）。我们用这种方法来检查生殖器HIV-1 DNA水平的标本，从妇女的血浆HIV-1 RNA水平低是典型的HIV-1非传播者。这些妇女宫颈分泌物中HIV-1 DNA拷贝数的中位数（1.8 HIV-1 DNA拷贝/10，000个细胞）低于血浆HIV-1 RNA水平较高的妇女（16.6 HIV-1 DNA拷贝/10 000个细胞）（P = 0.04），阴道分泌物中的HIV-1 DNA拷贝数中位数也是如此（不可检测vs 1.0 HIV-1 DNA拷贝/10，000个细胞）。这些数据表明，血浆HIV-1 RNA水平较低的妇女，因此预测HIV-1传播的风险较低，生殖器HIV-1细胞相关病毒水平较低。本文所述的检测方法可用于未来的研究，以检查细胞相关的HIV-1在传播中的作用。
Studies of human immunodeficiency virus type 1 (HIV-1) transmission suggest that genital HIV-1 RNA and DNA may both be determinants of HIV-1 infectivity. Despite its potential role in HIV-1 transmission, there are limited quantitative data on genital HIV-1 DNA. Here we validated an in-house real-time PCR method for quantification of HIV-1 DNA in genital specimens. In reactions with 100 genomes to 1 genome isolated from a cell line containing one HIV-1 provirus/cell, this real-time PCR assay is linear and agrees closely with a commercially available real-time PCR assay specific for a cellular housekeeping gene. In mock genital samples spiked with low numbers of HIV-1-infected cells such that the expected HIV-1 DNA copy number/reaction was 100, 10, or 5, the average copy number/reaction was 80.2 (standard deviation [SD], 28.3), 9.1 (SD, 5.4), or 3.1 (SD, 2.1), respectively. We used this method to examine genital HIV-1 DNA levels in specimens from women whose low plasma HIV-1 RNA levels are typical of HIV-1 nontransmitters. The median HIV-1 DNA copy number in endocervical secretions from these women (1.8 HIV-1 DNA copies/10,000 cells) was lower than that for women with higher plasma HIV-1 RNA levels (16.6 HIV-1 DNA copies/10,000 cells) (P = 0.04), as was the median HIV-1 DNA copy number in vaginal secretions (undetectable versus 1.0 HIV-1 DNA copies/10,000 cells). These data suggest that women with low plasma HIV-1 RNA and thus a predicted low risk of HIV-1 transmission have low levels of genital HIV-1 cell-associated virus. The assay described here can be utilized in future efforts to examine the role of cell-associated HIV-1 in transmission.