Abstract
The increase of foodborne viral outbreaks highlights the need for a rapid and sensitive method for the prediction of viral infectivity in food samples. This study assesses the use of propidium monoazide (PMA) coupled with real-time PCR methods (RT-qPCR or qPCR for RNA or DNA viruses, respectively) in the determination of viral infectivity in complex animal-related food matrices. Clam and Spanish fermented sausage (“chorizo”) samples were spiked with infectious and heat-inactivated human adenovirus-2 (HAdV-2) and mengovirus (vMC0). PMA-qPCR/RT-qPCR discriminated infective virus particles, with significant reductions (>2.7 log10 or 99.7%). Additionally, infectious HAdV-2 and vMC0 were quantified by plaque assay (in plaque forming units, PFU), and compared with those in virus genomes copies (GCs) quantified by PMA-qPCR/RT-qPCR. A consistent correlation (R2 > 0.92) was showed between PFU and GCs along serial 10-fold dilutions in both DNA and RNA virus and in both food matrices. This study shows the use of PMA coupled to qPCR/RT-qPCR as a promising alternative for prediction of viral infectivity in food samples in comparison to more expensive and time-consuming methods and for those viruses that are not able to grow under available cell culture techniques.
Highlights
Food and food related environments are a major source of viral transmission to humans (Koopmans et al, 2002; Rodríguez-Lázaro et al, 2012)
In order to assess the interference produced by propidium monoazide (PMA), food samples were firstly spiked with infectious virus particles and treated with PMA prior to real-time polymerase chain reaction (PCR)
Clam and “chorizo” samples were spiked with decreasing amounts of thermally inactivated viral particles (HAdV-2 and vMC0) that were submitted to a treatment with PMA prior to real-time PCR and the ability of the method for distinguishing between infectious and inactivated viruses was evaluated
Summary
Food and food related environments are a major source of viral transmission to humans (Koopmans et al, 2002; Rodríguez-Lázaro et al, 2012). Human NoV are responsible of more than five millions of gastroenteritis illnesses in USA annually, which represents more than 50% of the infectious cases (Scallan et al, 2011). These agents are recognized as major causes of foodborne illnesses, which can vary from gastroenteritis to hepatitis, paralysis or aseptic meningitis (Bosch et al, 2011). Pork meat products are a significant route of zoonotic transmission, as enteric viruses can infect humans from the consumption of contaminated raw or undercooked pork meat (Martínez-Martínez et al, 2011)
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