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首页> 外文期刊>Journal of Clinical Microbiology >Reverse transcription-PCR detection of hepatitis G virus.
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Reverse transcription-PCR detection of hepatitis G virus.

机译：逆转录PCR检测丙型肝炎病毒。

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Hepatitis G virus (HGV) was recently identified as a new member of the Flaviviridae, but its clinical significance is still unclear. Since no immunoassay for the diagnosis of HGV is available, we developed a sensitive reverse transcription-PCR (RT-PCR) assay to facilitate the detection of the viral genome by mass screening in the clinical laboratory. Sequences within the 5'-noncoding region and within the putative NS5a region are independently amplified in the presence of digoxigenin-11-dUTP and are detected by hybridization with biotinylated capture probes binding to a streptavidin-coated matrix. Semiquantitative Enzymun-Test DNA detection via chemiluminescence can be performed either in a microtiter plate format or on fully automated ES 300 machines. We were able to detect at least 8 x 10(2) genome equivalents per ml of serum using both primer pairs. HGV was shown to be present in 43 of 130 (33%) serum samples from intravenous drug abusers with a high risk of parenteral exposure. However, only two of the patients were positive when the NS5a primers only were used, and only one patient was positive when only the 5'-noncoding region primers were used, demonstrating the increased sensitivity of HGV detection with two sets of primers. Among these patients, there was no obvious correlation with other viral infections like hepatitis B virus, hepatitis C virus, or human immunodeficiency virus. Within a blood donor panel, 3 of 92 (3%) samples were found to be HGV positive, suggesting that donated blood may need to be screened for HGV.
机译：最近鉴定为丙型肝炎病毒（HGV）是黄病毒科的新成员，但其临床意义仍不清楚。由于尚无用于诊断HGV的免疫测定法，因此我们开发了一种灵敏的逆转录PCR（RT-PCR）测定法，以利于在临床实验室中通过大规模筛选来检测病毒基因组。在存在洋地黄毒苷-11-dUTP的情况下，可独立扩增5'-非编码区内和推定NS5a区内的序列，并通过与与链霉亲和素包被的基质结合的生物素化捕获探针杂交来检测。通过化学发光法进行的半定量酶检测DNA检测可以微量滴定板形式或在全自动ES 300机器上进行。使用这两个引物对，我们能够检测到每毫升血清至少8 x 10（2）个基因组当量。结果表明，在静脉注射药物滥用者的130份血清样品中，有43份（33％）存在HGV，具有很高的非肠道暴露风险。但是，仅使用NS5a引物时，只有两名患者为阳性，而仅使用5'-非编码区引物时，只有一名患者为阳性，这表明使用两组引物可提高HGV检测的灵敏度。在这些患者中，与其他病毒感染（如乙型肝炎病毒，丙型肝炎病毒或人类免疫缺陷病毒）没有明显的相关性。在献血者小组中，发现92个样本中有3个（3％）为HGV阳性，这表明可能需要对献血进行HGV筛查。

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