Impacts of phenolic compounds on RT-qPCR detection of hepatitis A virus in berries.
Berries are frequently implicated in outbreaks of foodborne illness due to viruses, particularly norovirus and hepatitis A virus. Compounds naturally present in berries can compromise the reliability of RT-qPCR methods, such as ISO 15216-1:2017, for detecting and quantifying viruses in foods. The aim of this study was to evaluate the inhibitory impact of seven phenolic compounds (ellagic acid, hydroxybenzoic acid, caffeic acid, coumaric acid, ferulic acid, quercetin, and cyanidine-3-glucoside) found naturally in raspberries as well as batch effects due to different concentrations of inhibitors (e.g., associated with ripeness) when using RT-qPCR to detect HAV in raspberries, blackberries, strawberries, blueberries, cranberries, and mixed berries. To assess the impact of dilution on RT-qPCR inhibition, samples were diluted at four levels (1/2, 1/5, 1/10, 1/100). Spiking the RT-qPCR reaction mixture with each phenolic compound at its natural concentration in raspberries showed that ellagic acid, hydroxybenzoic acid, caffeic acid and cyanidin-3-glucoside inhibited amplification, but only ellagic acid remained inhibitory in the ISO method. HAV recovery from frozen strawberries was undetectable (0%) without additional treatment but reached 39% with the OneStep PCR Inhibitor Removal Kit. For frozen blueberries, MobiSpin S-400 performed better, yielding about 52% recovery compared to 23% with OneStep. Sample dilution further enhanced HAV detection across most berry types.
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