Institute of Agrochemistry and Food Technology-IATA, Spanish Council for Scientific Research-CSIC, Spain.
Int J Food Microbiol. 2012 Jan 3;152(1-2):9-13. doi: 10.1016/j.ijfoodmicro.2011.10.002. Epub 2011 Oct 8.
Fresh-cut vegetables are prone to be contaminated with foodborne pathogens during growth, harvest, transport and further processing and handling. As most of these products are generally eaten raw or mildly treated, there is an increase in the number of outbreaks caused by viruses and bacteria associated with fresh vegetables. Foodborne pathogens are usually present at very low levels and have to be concentrated (i.e. viruses) or enriched (i.e. bacteria) to enhance their detection. With this aim, a rapid concentration method has been developed for the simultaneous recovery of hepatitis A virus (HAV), norovirus (NV), murine norovirus (MNV) as a surrogate for NV, Escherichia coli O157:H7, Listeria monocytogenes and Salmonella enterica. Initial experiments focused on evaluating the elution conditions suitable for virus release from vegetables. Finally, elution with buffered peptone water (BPW), using a Pulsifier, and concentration by polyethylene glycol (PEG) precipitation were the methods selected for the elution and concentration of both, enteric viruses and bacteria, from three different types of fresh-cut vegetables by quantitative PCR (qPCR) using specific primers. The average recoveries from inoculated parsley, spinach and salad, were ca. 9.2%, 43.5%, and 20.7% for NV, MNV, and HAV, respectively. Detection limits were 132 RT-PCR units (PCRU), 1.5 50% tissue culture infectious dose (TCID₅₀), and 6.6 TCID₅₀ for NV, MNV, and HAV, respectively. This protocol resulted in average recoveries of 57.4%, 64.5% and 64.6% in three vegetables for E. coli O157:H7, L. monocytogenes and Salmonella with corresponding detection limits of 10³, 10² and 10³ CFU/g, respectively. Based on these results, it can be concluded that the procedure herein is suitable to recover, detect and quantify enteric viruses and foodborne pathogenic bacteria within 5 h and can be applied for the simultaneous detection of both types of foodborne pathogens in fresh-cut vegetables.
新鲜切割的蔬菜在生长、收获、运输以及进一步加工和处理过程中容易受到食源性病原体的污染。由于这些产品大多数通常是生吃或轻度处理的，因此与新鲜蔬菜相关的病毒和细菌引起的暴发数量有所增加。食源性病原体通常存在于非常低的水平，必须进行浓缩（即病毒）或富集（即细菌）以提高其检测率。为此，开发了一种快速浓缩方法，用于同时回收甲型肝炎病毒 (HAV)、诺如病毒 (NV)、鼠诺如病毒 (MNV)（作为 NV 的替代品）、大肠杆菌 O157:H7、李斯特菌单核细胞增生李斯特菌和肠炎沙门氏菌。最初的实验侧重于评估适合从蔬菜中释放病毒的洗脱条件。最后，使用脉动器用缓冲蛋白胨水 (BPW) 洗脱，并通过聚乙二醇 (PEG) 沉淀浓缩，是从三种不同类型的新鲜切割蔬菜中通过使用特定引物的定量 PCR (qPCR) 对肠道病毒和细菌进行洗脱和浓缩的方法。从接种的欧芹、菠菜和沙拉中回收的平均回收率分别为 NV、MNV 和 HAV 的 9.2%、43.5%和 20.7%。NV、MNV 和 HAV 的检测限分别为 132 RT-PCR 单位 (PCRU)、1.5 50%组织培养感染剂量 (TCID₅₀) 和 6.6 TCID₅₀。该方案导致三种蔬菜中大肠杆菌 O157:H7、单核细胞增生李斯特菌和肠炎沙门氏菌的平均回收率分别为 57.4%、64.5%和 64.6%，相应的检测限分别为 10³、10²和 10³ CFU/g。基于这些结果，可以得出结论，该程序适用于在 5 小时内回收、检测和定量肠道病毒和食源性致病菌，并可应用于新鲜切割蔬菜中这两种食源性致病菌的同时检测。
