Human noroviruses are among the main causes of acute gastroenteritis worldwide. Frozen raspberries have been linked to several norovirus food-related outbreaks. However, the extraction of norovirus RNA from frozen raspberries remains challenging. Recovery yields are low and PCR inhibitors limit the sensitivity of the detection methodologies. In 2017, 724 people from various regions of the Province of Quebec, Canada, were infected by noroviruses and the outbreak investigation pointed to frozen raspberries as a putative source. A new magnetic silica bead approach was used for the extraction of viruses from different outbreak samples. The RNA extracts were tested by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and five samples were confirmed positive for norovirus by RT-qPCR amplicon sequencing. A multiplex long-range two-step RT-PCR approach was developed to amplify norovirus ORF2 and ORF3 capsid genes from the positive frozen raspberry RNA extracts and other sequencing strategies were also explored. These capsid genes were sequenced by Next-Generation Sequencing. Phylogenetic analyses confirmed the presence of multiple genotypes (GI.3, GI.6, and GII.17) and intra-genotype variants in some of the frozen raspberry samples. Variants of genotype GI.3 and GI.6 had 100% homology with sequences from patient samples. Similar strains were also reported in previous outbreaks. Confirmation approaches based on sequencing the norovirus capsid genes using Next-Generation Sequencing can be applied at trace level contaminations and could be useful to assess risk and assist in source tracking.

人类诺如病毒是全球急性胃肠炎的主要原因之一。冷冻覆盆子与几次诺如病毒食品相关的暴发有关。然而，从冷冻覆盆子中提取诺如病毒 RNA 仍然具有挑战性。回收率很低，PCR 抑制剂限制了检测方法的灵敏度。2017 年，来自加拿大魁北克省不同地区的 724 人感染了诺如病毒，疫情调查指出冷冻覆盆子是推定的来源。一种新的磁性硅珠方法被用于从不同的暴发样本中提取病毒。通过逆转录-定量聚合酶链反应 (RT-qPCR) 对 RNA 提取物进行检测，通过 RT-qPCR 扩增子测序确认五个样本对诺如病毒呈阳性。开发了一种多重长程两步 RT-PCR 方法来从阳性冷冻覆盆子 RNA 提取物中扩增诺如病毒 ORF2 和 ORF3 衣壳基因，还探索了其他测序策略。这些衣壳基因通过下一代测序进行测序。系统发育分析证实在一些冷冻覆盆子样本中存在多种基因型（GI.3、GI.6 和 GII.17）和基因型内变异。基因型 GI.3 和 GI.6 的变体与来自患者样本的序列具有 100% 的同源性。在以前的暴发中也报告了类似的菌株。基于使用下一代测序对诺如病毒衣壳基因进行测序的确认方法可应用于痕量污染，并可用于评估风险和协助源头追踪。