Evaluation of viral extraction methods on a broad range of Ready-To-Eat foods with conventional and real-time RT-PCR for Norovirus GII detection
Evaluation of viral extraction methods on a broad range of Ready-To-Eat foods with conventional and real-time RT-PCR for Norovirus GII detection
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DOI：
10.1016/j.ijfoodmicro.2007.12.020
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发表时间：
2008-03-31
影响因子：
5.4
通讯作者：
Debevere, Johan
中科院分区：
文献类型：
--
作者：
Baert, Leen;Uyttendaele, Micke;Debevere, Johan
诺如病毒 (NoV) 是食源性疫情的常见原因。尽管如此，目前还没有适用于食品的标准病毒检测方法。因此，我们对多种即食 (RTE) 食品（混合生菜、水果沙拉、覆盆子和两种 RTE 菜肴）中的三种病毒洗脱浓缩方法和一种直接 RNA 分离方法进行了评估，这些食品人工接种了受 NoV 基因组 II 污染的稀释粪便样本。这些播种实验揭示了两类 RTE 产品，即水果和蔬菜，而富含蛋白质和脂肪的 RTE 菜肴（通心粉和意大利面条沙拉）则形成了另一类。使用两种传统 RT-PCR 系统（Booster 和半巢式 GII）和一种实时 RT-PCR（实时 GII）测定法对 RNA 提取物进行扩增和检测。快速直接 RNA 分离方法通过传统 RT-PCR 检测在 10 g 通心粉和意大利面条沙拉中检测到 10(2) RT-PCRU。然而，实时 RTPCR 对通心粉沙拉的敏感性较低。一种病毒洗脱浓缩方法，包括用于洗脱步骤的缓冲溶液和一个聚乙二醇 (PEG) 沉淀步骤，能够通过常规和实时 RT-PCR 检测在 50 g 冷冻覆盆子上检测到 102 RT-PCRU。而且后一种提取方法不使用对环境有害的化学试剂并且易于操作。 (C) 2007 Elsevier B.V. 保留所有权利。
Noroviruses (NoV) are a common cause of foodborne Outbreaks. In spite of that, no standard viral detection method is available for food products. Therefore, three viral elution-concentration methods and one direct RNA isolation method were evaluated on a broad range of Ready-To-Eat (RTE) food products (mixed lettuce, fruit salad, raspberries and two RTE dishes) artificially seeded with a diluted stool sample contaminated with NoV genogroup II. These seeding experiments revealed two categories of RTE products, fruits and vegetables grouped together and RTE dishes (penne and tagliatelle salads) which are rich in proteins and fat formed another category. The RNA extracts were amplified and detected with two conventional RT-PCR systems (Booster and Semi-nested GII) and one real-time RT-PCR (Real-time GII) assay. A fast direct RNA isolation method detected 10(2) RT-PCRU on 10 g penne and tagliatelle salads with the conventional RT-PCR assays. However real-time RTPCR was less sensitive for penne salad. A viral elution-concentration method, including a buffer solution for the elution step and one polyethylene glycol (PEG) precipitation step, was able to detect 102 RT-PCRU on 50 g frozen raspberries with conventional and real-time RT-PCR assays. Moreover the latter extraction method used no environmental hazardous chemical reagents and was easy to perform. (C) 2007 Elsevier B.V. All rights reserved.