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Quantification of Pathogenic Viruses in Ready-To-Eat (RTE) Foods


Data from the Centre for Disease Control and Prevention (CDC) published in 1999 suggests that almost 80% of the estimated 38.5 million illnesses due to foodborne pathogens annually in the US are attributable to viruses. Of these the vast majority (23 million cases) are caused by noroviruses with hepatitis A accounting for a further 83,400 cases. Very little is known about the prevalence of viruses in our food supply although the major sources of these viruses appear to include foods such as shellfish, fruits and vegetables coming into contact with contaminated water in their growing area before harvest; and foods that become contaminated during preparation through contact with fecally contaminated surfaces or infected food handlers. Despite these high figures, methods to detect foodborne viruses are still in their infancy but with the advent of molecular techniques such as PCR, reliable and rapid assays are emerging. However, one of the major problems in the application of PCR protocols to foods is the removal of target genetic material in a form that can be amplified. To remove and concentrate targets from food, methods such as immunomagnetic separation and filtration have been applied but work to date has mainly focused on bacterial pathogens. Also PCR assays provide an opportunity to detect more than one organism in a single sample, but, again, this multiplexing approach has not been widely used in the field of food virology.
This study will build on research that has been completed in the Canadian Research Institute for Food Safety (CRIFS) laboratories in which has developed a real-time RT-PCR assay for the detection of hepatitis A in fresh produce. The aim of this project is to develop a real-time PCR methodology for detection and quantification of pathogenic viruses in Ready-to-Eat (RTE) foods, wherein the main focus will not only be the development of a multiplex quantitative real-time PCR assay but especially the development of a sample treatment method capable of concentrating very low amounts of virus from the food sample. To achieve this we will study immunomagnetic separation, filtration using charged membranes and a proprietary product produced by a Canadian company, EcoVu Analytics, which uses novel chemistry to capture particles from suspension, to capture and concentrate virus particles from a variety of foods.

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Expected Impact of Project Outcomes on Food Safety in Ontario:
Noroviruses (NV) and hepatitis A virus (HAV) are the most commonly reported food-borne viruses worldwide with NV estimated as the leading cause of food-borne illness in the US. In Ontario about 50% of the hepatitis A reported cases are associated with food- or waterborne transmission. It has been estimated that the costs of hepatitis A infections may be as much as $3,000 per adult case and $1,750 for infant cases. Health-care associated outbreaks of gastroenteritis are also becoming an increasing problem. A study in the UK showed that NV was responsible for 63% of such outbreaks and gastroenteritis outbreaks as a whole were estimated to cost the English National Health Service a staggering $2.5 million each year.
Common-source outbreaks are frequently due to contamination of the food by an infected food-handler, just prior to consumption. However, reports of contamination of foods before retail distribution are increasing and may be prevented by appropriate testing of high-risk food commodities. The aim of this project is to use real-time RT-PCR for rapid simultaneous detection and possible quantification of NV and HAV in RTE foods. To obtain efficient detection, it is crucial to integrate a real-time PCR method with a suitable sample preparation method. Therefore, methods to concentrate possibly very low amounts of virus before PCR detection and remove PCR inhibitors have to be developed or improved. Finally, by developing a quantitative method, much needed data on infectious dose and virus contamination levels in RTE foods will provide information to OMAFRA which will be invaluable in setting priorities for provincial food safety programs. <P> For more information, please visit the <a href="; target="_blank">Ontario Ministry of Agriculture, Food & Rural Affairs (OMAFRA) Food Safety Research Program</a>.

Griffiths, Mansel
University of Guelph
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