采用干血斑进行单次聚合酶链反应检测诊断非洲环境中婴儿 HIV-1 感染的评估。
Evaluation of a single round polymerase chain reaction assay using dried blood spots for diagnosis of HIV-1 infection in infants in an African setting.
机构信息
Department of Medical Microbiology, University of Nairobi-College of Health Sciences, off Ngong Road, Nairobi, Box 19767-00202, Kenya.
出版信息
BMC Pediatr. 2011 Feb 18;11:18. doi: 10.1186/1471-2431-11-18.
BACKGROUND
The aim of this study was to develop an economical 'in-house' single round polymerase chain reaction (PCR) assay using filter paper-dried blood spots (FP-DBS) for early infant HIV-1 diagnosis and to evaluate its performance in an African setting.
METHODS
An 'in-house' single round PCR assay that targets conserved regions in the HIV-1 polymerase (pol) gene was validated for use with FP-DBS; first we validated this assay using FP-DBS spiked with cell standards of known HIV-1 copy numbers. Next, we validated the assay by testing the archived FP-DBS (N=115) from infants of known HIV-1 infection status. Subsequently this 'in-house' HIV-1 pol PCR FP-DBS assay was then established in Nairobi, Kenya for further evaluation on freshly collected FP-DBS (N=186) from infants, and compared with findings from a reference laboratory using the Roche Amplicor® HIV-1 DNA Test, version 1.5 assay.
RESULTS
The HIV-1 pol PCR FP-DBS assay could detect one HIV-1 proviral copy in 38.7% of tests, 2 copies in 46.9% of tests, 5 copies in 72.5% of tests and 10 copies in 98.1% of tests performed with spiked samples. Using the archived FP-DBS samples from infants of known infection status, this assay was 92.8% sensitive and 98.3% specific for HIV-1 infant diagnosis. Using 186 FP-DBS collected from infants recently defined as HIV-1 positive using the commercially available Roche Amplicor v1.5 assay, 178 FP-DBS tested positive by this 'in-house' single-round HIV-1 pol PCR FP-DBS PCR assay. Upon subsequent retesting, the 8 infant FP-DBS samples that were discordant were confirmed as HIV-1 negative by both assays using a second blood sample.
CONCLUSIONS
HIV-1 was detected with high sensitivity and specificity using both archived and more recently collected samples. This suggests that this 'in-house' HIV-1 pol FP-DBS PCR assay can provide an alternative cost-effective, reliable and rapid method for early detection of HIV-1 infection in infants.
背景
本研究旨在开发一种经济的“内部”单轮聚合酶链反应（PCR）检测方法，使用滤纸干血斑（FP-DBS）进行早期婴儿 HIV-1 诊断，并在非洲环境中评估其性能。
方法
针对 HIV-1 聚合酶（pol）基因保守区域的“内部”单轮 PCR 检测方法在 FP-DBS 上进行了验证；首先，我们使用含有已知 HIV-1 拷贝数的细胞标准品的 FP-DBS 对该检测方法进行了验证。接下来，我们通过检测已知 HIV-1 感染状态的婴儿的存档 FP-DBS（N=115）验证了该检测方法。随后，在肯尼亚内罗毕建立了这种“内部”HIV-1 pol PCR FP-DBS 检测方法，用于进一步评估新鲜采集的 FP-DBS（N=186），并与使用罗氏 Amplicor®HIV-1 DNA Test，version 1.5 检测方法的参考实验室的结果进行比较。
结果
HIV-1 pol PCR FP-DBS 检测方法可在 38.7%的检测中检测到 1 个 HIV-1 前病毒拷贝，在 46.9%的检测中检测到 2 个拷贝，在 72.5%的检测中检测到 5 个拷贝，在 98.1%的检测中检测到 10 个拷贝。使用已知感染状态的婴儿存档的 FP-DBS 样本，该检测方法对 HIV-1 婴儿诊断的灵敏度为 92.8%，特异性为 98.3%。使用最近通过市售罗氏 Amplicor v1.5 检测方法定义为 HIV-1 阳性的 186 份 FP-DBS 样本，178 份 FP-DBS 样本通过这种“内部”单轮 HIV-1 pol PCR FP-DBS PCR 检测方法呈阳性。随后的重新检测显示，8 份 FP-DBS 样本的结果不一致，在使用第二份血样后，这 8 份样本均被两种检测方法确认为 HIV-1 阴性。
结论
使用存档和最近采集的样本均能高度灵敏和特异性地检测到 HIV-1。这表明，这种“内部”HIV-1 pol FP-DBS PCR 检测方法可以为早期发现婴儿 HIV-1 感染提供一种替代的经济、可靠和快速的方法。
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