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Persistence and Inactivation of Norovirus in Fresh Produce Chains

Persistence and Inactivation of Norovirus in Fresh Produce Chains. Katharina Verhaelen. Introduction. Human Noroviruses (hNoV) are the leading cause of food-borne viral gastroenteritis worldwide Number of reported outbreaks attributed to fresh produce increasing

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Persistence and Inactivation of Norovirus in Fresh Produce Chains

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  1. Persistence and Inactivation of Norovirus in Fresh Produce Chains Katharina Verhaelen

  2. Introduction • Human Noroviruses (hNoV) are the leading cause of food-borne viral gastroenteritis worldwide • Number of reported outbreaks attributed to fresh produce increasing • hNoV is the most important cause of fresh produce associated outbreaks DeWaal and Buhiya, 2007

  3. Work package 6 Objectives To determine the persistence of NoV under conditions encountered in food supply chains To asses the efficacy of current and novel intervention measures in inactivating NoV To verify the disinfectant efficacy of interventions at pilot plant scale and in field experiments for hNoV Gathered quantitative data on NoV reduction will feed back into QVRA

  4. Life of a berry Fresh Processed Dried Puree Juice Frozen • Conditions/Preservation Measures • Freeze drying: • Freezing, Heat + reduced pressure  sublimation of ice • Literature data: (Butot, 2009) • Raspberries/Strawberries: reduction hNoV GI ~0.7 log, hNoV GII ~1.3 log

  5. Life of a berry Fresh Processed Dried Puree Juice Frozen • Conditions/Preservation Measures • Pasteurisation: • Scenario a)88°C, 2min; Scenario b1) 65°C, 30s or b2)75 °C, 15s • Literature data: (Baert, 2008) • MNV-1 log (PFU) reduction in buffer at 80°C, 2.5min: a) > 6 log • MNV-1 log (PFU) reduction in rasp. puree: b1) 1.8 log b2) 2.8 log

  6. Life of a berry Fresh Processed Dried Puree Juice Frozen • Conditions/Preservation Measures • Freezing • Literature data: (Butot, 2008) • FCV log (TCID50) reduction, 2 days: raspb. 1.1 log; strawb. 2.7 log • hNoV GI/GII (PCRU) reduction, 2 days, raspberry: no reduction • hNoV GI/GII (PCRU) reduction, 2 days, strawberry: < 0.4 log

  7. Life of a berry Fresh Processed Dried Puree Juice Frozen • Conditions/Preservation Measures • Chilling • Literature data: (Mattison, 2007) • FCV reduction (PFU), strawberries, 6 days, 4°C: > 3.5 log reduction • No data on hNoV/MNV-1 persistence on raspberries/strawberries

  8. Life of a berry Fresh Processed Dried Puree Juice Frozen Conclusion • If data available often inconclusive/incomplete • Study persistence on fresh and frozen berries • Highest risk (short shelf life, conditions preserve virus infectivity) • Highest consumption

  9. Viruses under Study • hNoV GI.4 & GII.4  target pathogen • Murine norovirus (MNV-1)  surrogate to study hNoV infectivity • Human adenovirus (hAdV)  indicator for hNoV • Low and high virus titer

  10. Approach Spike food with virus Virus inactivation • Controls • Process control: • Mengovirus • Plaque assay • hAdV: Virobathe protocol • MNV: Wobus, 2004 Sample taking during time Elution of virus particles from food surfaceusing TGBE buffer pH 9.5 Centrifugation10000 rpm Precipitation of virus particles using polyethylene glycol pH 7.2 Centrifugation10000 rpm Centrifugation10000 rpm Chloroform:Butanol extraction Filtration Nucleic acid extraction using magnetic silica Quantitative PCR Plaque assay

  11. Pre-Experiment Aim: • Determination of experimental set up • Initial virus concentration  low recovery • Sampling intervals • Testing the whole procedure Approach: • 10g raspberry spiked with 5000 PFU of MNV-1 and hAdV • Duplicates • Storage of raspberries at 4°C for 1 week and at room temperature for 2 days Preliminary results: • No decrease in Cp values for hAdV and MNV-1 after 7 days of storage at 4°C and after 2 days at RT compared to T0 control • Cp (hAdV): 31-32 • Cp (MNV-1) 10x dilution: 33-34 • Infectivity data not yet available  difficulties in culturing RAW 264.7 cells

  12. Conditions Mimic storage conditions of fresh and frozen raspberries/strawberries: Temperature profile: Harvest Distribution Point of sale Harvest Distribution Point of sale RT RT 10 °C 10 °C 14 d 4 °C 4 °C 7 d

  13. Expected Outcome • Knowledge on persistence of hNoV on soft berries under commonly applied storage conditions will provide data for: • Setting health targets • Assessing public health risk • Redesigning food processing practices • hAdV useful indicator of hNoV contamination of food commodities in respect to its persistence • Suitability of MNV-1 as a surrogate of hNoV removal • Differences in persistence of hNoV GI and GII • Impact of food matrix on hNoV persistence

  14. Outlook for this year • Persistence of hNoV in agrochemicals • Application of agrochemicals possible introduction source for hNoV on fresh produce? • In the raspberry production insecticides are applied prior to mechanical harvesting to avoid contamination of produce • Direct contact with produce • Inactivation of hNoV in raspberry puree by high pressure processing on industrial scale (ITACyL) • Efficacy of lettuce washing with and without inclusion of sanitizers in inactivating hNoV on pilot plant scale (ITACyL)

  15. Acknowledgement: Ana Maria de Roda Husman Saskia Rutjes Martijn Bouwknegt Froukje Lodder Mohammed Alsady Marjolijn Schalk Willemijn Lodder Erwin Duizer & Era Tuladhar Arie Havelaar Frans van Knapen Thanks for your attention!

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