AUTHOR=Quijada Narciso M. , Fongaro Gislaine , Barardi Célia R. M. , Hernández Marta , Rodríguez-Lázaro David 

TITLE=Propidium Monoazide Integrated with qPCR Enables the Detection and Enumeration of Infectious Enteric RNA and DNA Viruses in Clam and Fermented Sausages

JOURNAL=Frontiers in Microbiology

VOLUME=7

YEAR=2016

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2016.02008

DOI=10.3389/fmicb.2016.02008

ISSN=1664-302X

ABSTRACT=<p>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 (vMC<sub>0</sub>). PMA-qPCR/RT-qPCR discriminated infective virus particles, with significant reductions (&gt;2.7 log<sub>10</sub> or 99.7%). Additionally, infectious HAdV-2 and vMC<sub>0</sub> 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 (<italic>R</italic><sup>2</sup> &gt; 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.</p>