National Virus Reference Laboratory (NVRL), University College Dublin, Belfield, Dublin 4, Ireland.
J Clin Virol. 2011 Sep;52(1):38-44. doi: 10.1016/j.jcv.2011.05.022. Epub 2011 Jun 24.
Routine diagnosis of Human T Lymphotropic virus (HTLV) infection is primarily serologically based; however the proportion of unresolved and indeterminate Western blot results range from 0.02% to 50% in endemic areas.
To validate a sensitive in-house quantitative multiplex real-time assay (mqRT-PCR), capable of detecting and quantifying HTLV-1 and HTLV-2, and use it to differentiate unresolved serological profiles, and monitor infection in HTLV-1 infected patients.
The mqRT-PCR was designed as a single-tube assay. Quantitative results were reported as copy number of HTLV provirus per 10(6) cells and the numbers of cells were calculated based on the quantitative result for albumin, of which there are 2 copies/cell. Assay standards were amplified from HTLV-1 infected MT-2 cells and HTLV-2 transfected CEM cells. Blood samples were obtained from HTLV seropositive former blood donors.
The mqRT-PCR assay was efficient (98.8-101.2%), reproducible (coefficient of variance<5%) and sensitive to 1 copy for HTLV-1, HTLV-2 and Albumin. The assay resolved the infection profile in 16/17 patients, with undetermined subtype, all of which were reassigned as HTLV-1 infections. In addition, the average PVL detected in patients suffering from HTLV-1 associated HAM/TSP (n=23, 13,450 copies/10(6) cells) was significantly higher than those detected in asymptomatic carriers (n=21, 6665 copies/10(6) cells).
We propose a new testing algorithm for the laboratory diagnosis of HTLV infection, which includes HTLV specific mqRT-PCR for resolving HTLV serological results. Furthermore, quantitation of PVL load by real-time PCR may be useful in assessing the link between infection and disease, and in monitoring patients undergoing therapy.
人类嗜 T 淋巴细胞病毒（HTLV）感染的常规诊断主要基于血清学；然而，在流行地区，未解决和不确定的 Western blot 结果比例在 0.02%至 50%之间。
验证一种灵敏的内部定量多重实时聚合酶链反应（mqRT-PCR），能够检测和定量 HTLV-1 和 HTLV-2，并将其用于区分未解决的血清学特征，并监测 HTLV-1 感染患者的感染情况。
mqRT-PCR 设计为单管检测。定量结果以每 10(6)个细胞中的 HTLV 原病毒拷贝数报告，细胞数量根据白蛋白的定量结果计算，每个细胞有 2 个拷贝。该检测标准从 HTLV-1 感染的 MT-2 细胞和 HTLV-2 转染的 CEM 细胞中扩增。血液样本来自 HTLV 血清阳性的前献血者。
mqRT-PCR 检测具有高效（98.8-101.2%）、重复性好（变异系数<5%）和对 1 拷贝 HTLV-1、HTLV-2 和白蛋白的灵敏度。该检测方法解决了 17 名未确定亚型的患者的感染状况，所有患者均被重新归类为 HTLV-1 感染。此外，HTLV-1 相关 HAM/TSP 患者（n=23，13450 拷贝/10(6)个细胞）的平均 PVL 显著高于无症状携带者（n=21，6665 拷贝/10(6)个细胞）。
我们提出了一种新的 HTLV 感染实验室诊断检测算法，其中包括用于解决 HTLV 血清学结果的 HTLV 特异性 mqRT-PCR。此外，通过实时 PCR 定量 PVL 载量可能有助于评估感染与疾病之间的联系，并监测接受治疗的患者。
