Institute of Nutraceuticals and Functional Foods (INAF), Université Laval, Québec, Canada.
Foodborne Pathog Dis. 2013 Jan;10(1):47-54. doi: 10.1089/fpd.2011.1105.
Four nucleic acid extraction methods were evaluated for the purpose of quantifying a norovirus surrogate (murine norovirus [MNV-1]) concentrated from different food samples. Simple (strawberries and lettuce) and complex (sliced turkey breast, soft-shell clams, and potato salad) food matrices were inoculated with a viral suspension containing high (4×10(5) PFU) or low (4×10(3) PFU) numbers of viral particles. MNV-1 was eluted using either the Pulsifier™ or repetitive pipetting. The four methods were based on using magnetic silica (MiniMAG), non-magnetic silica (bioMérieux Basic kit), silica membrane (Qiagen kit), and phenol (TriReagent) for RNA extraction. The greatest recovery of viral RNA from simple matrices was obtained using magnetic silica for both inoculation levels. For strawberries, the addition of pectinase during the elution step improved RNA recovery when the Pulsifier was used with silica membrane extraction and when repetitive pipetting was used with magnetic silica extraction. In the case of complex matrices, the extraction of high or low numbers of MNV-1 was highest overall using magnetic silica. The exception was soft-shell clams with a high viral load, in which the greatest recovery was obtained with the phenol-based method. In general, magnetic silica was the most effective for extracting both high and low numbers of MNV-1 particles from a wide range of foods.
评估了四种核酸提取方法，目的是定量检测从不同食品样本中浓缩的诺如病毒替代物（鼠诺如病毒[MNV-1]）。用含有高（4×10(5)PFU）或低（4×10(3)PFU）病毒颗粒数的病毒悬浮液接种简单（草莓和生菜）和复杂（切片火鸡肉、软壳蛤和土豆沙拉）食品基质。使用 Pulsifier™或重复吸液法洗脱 MNV-1。这四种方法基于使用磁性硅（MiniMAG）、非磁性硅（bioMérieux Basic 试剂盒）、硅膜（Qiagen 试剂盒）和苯酚（TriReagent）提取 RNA。对于两种接种水平，从简单基质中回收病毒 RNA 的最佳方法是使用磁性硅。对于草莓，在用硅膜提取时，当使用 Pulsifier 时，在用磁性硅提取时重复吸液，添加果胶酶可提高 RNA 回收效率。对于复杂基质，总体而言，使用磁性硅可最高程度地提取高或低数量的 MNV-1。例外的是高病毒载量的软壳蛤，其中基于苯酚的方法可获得最大的回收率。总的来说，磁性硅是从各种食品中提取高数量和低数量 MNV-1 颗粒的最有效方法。
