Method Development for Detection of Human Norovirus in Produce Samples during an Outbreak Investigation

Introduction: Noroviruses are the leading causes of outbreaks as well as sporadic cases of acute gastroenteritis worldwide. Although epidemiological evidence has indicated that norovirus is responsible for the majority of produce related outbreaks in both Europe and the United States there are still few occasions where it has been successfully detected from foods associated in outbreaks.

Purpose: Fruit salad was implicated in a recent norovirus outbreak in Wisconsin. An experimental investigation was undertaken to develop a method and test the implicated food vehicle.

Methods: Fifty grams of fruit salad (watermelon, cantaloupe, pineapple, honeydew and grapes) were artificially inoculated with norovirus. The protocol involved eluting the virus with  a 0.1 M Tris-HCl, 0.05 M glycine, 1% beef extract, pH 9.2 (TGBE) buffer containing 2% Polyvinyl Pyrrolidone (PVP) and pectinase. After a brief spin, the eluate containing the virus particles was concentrated by pelleting virus via ultracentrifugation.  Virus RNA was subsequently isolated from the pellet using a commercial kit, and viral genome detection accomplished using real-time RT-PCR.

 Results: When fruit salad was artificially seeded, norovirus could be consistently detected at a level of at least 1,000 RNA copies per 50 g of sample. When the same experimental method was used to process samples from the outbreak, they all tested positive and more specifically contamination ranged from 10-1,000 RNA copies/sample.

Significance: Developing a robust method to detect norovirus in food samples associated with foodborne illness allows for a confirmatory tool for validation and application in response to outbreak management.