Unité VAE (Virologie des Aliments et de l'Eau), Agence Française de Sécurité Sanitaire des Aliments (AFSSA), Laboratoire d'Etudes et de Recherches sur la Qualité des Aliments et sur les Procédés Agroalimentaires (LERQAP), 23 Avenue du Général de Gaulle, 94706 Maisons-Alfort Cedex, France.
J Virol Methods. 2010 Jun;166(1-2):48-53. doi: 10.1016/j.jviromet.2010.02.017. Epub 2010 Feb 25.
Hepatitis A virus (HAV) infection is the leading worldwide cause of acute viral hepatitis. An important aspect of viral control is rapid diagnosis. Epidemiological studies have linked hepatitis A outbreaks to the consumption of drinking water or soft fruits exposed to faecal contamination. Real-time reverse transcriptase PCR (qRT-PCR) is now widely used for detecting RNA viruses in food samples. Efficiency of viral concentration, nucleic acid extraction and the presence of potential inhibitors of the RT-PCR reaction must be monitored to prevent false negative results. In this study, the MS2 bacteriophage used as a process control was detected simultaneously with HAV in a one-step duplex real-time qRT-PCR. The assay was developed for testing water and raspberries. Adding MS2 showed no loss of sensitivity for HAV detection in water and raspberry samples. The limit of detection of HAV with this new approach was 10PFU for 1.5L of bottled water, 100PFU for 1.5L of tap water, 50PFU for 25g of fresh raspberries and 100PFU for 25g of frozen raspberries. The data show that the MS2 offers a very reliable and simple way to monitor false-negative results, making it a valuable tool in the routine diagnostics laboratory.
甲型肝炎病毒（HAV）感染是全球导致急性病毒性肝炎的主要原因。病毒控制的一个重要方面是快速诊断。流行病学研究将甲型肝炎爆发与饮用水或暴露于粪便污染的软水果的消费联系起来。实时逆转录聚合酶链反应（qRT-PCR）现在广泛用于检测食品样本中的 RNA 病毒。必须监测病毒浓缩、核酸提取的效率以及 RT-PCR 反应的潜在抑制剂的存在，以防止假阴性结果。在这项研究中，MS2 噬菌体被用作过程控制，与 HAV 一起在一步法双重实时 qRT-PCR 中同时检测。该检测方法用于测试水和覆盆子。在水中和覆盆子样本中添加 MS2 并未降低 HAV 检测的灵敏度。使用这种新方法，HAV 的检测限为 1.5L 瓶装水中的 10PFU、1.5L 自来水中的 100PFU、25g 新鲜覆盆子中的 50PFU 和 25g 冷冻覆盆子中的 100PFU。数据表明，MS2 提供了一种非常可靠和简单的方法来监测假阴性结果，使其成为常规诊断实验室的有价值工具。
