Purpose: The purpose of this study was to validate a new one-step multiplex RT-Real-time PCR-kit for simultaneous detection and differentiation of HAV, NoV (GI, GII) and MS2 phage as a process control. The kit was developed and optimized for the special needs in food and water analytics, regarding inhibiting substances and low-dose contamination in soft fruits, minced meat, seafood and drinking water.
Methods: Virus concentrations from food (berries, mussels, and minced meat) and water samples were carried out according to ISO/TS 15216. RNA was prepared by the foodproof Sample Preparation Kit IV. Virus detection was performed with the foodproof Hepatitis A virus plus Norovirus (GI, GII) Detection Kit.
Results: Norovirus- and hepatitis A virus-positive samples were tested. Specificity was controlled by several norovirus and hepatitis A virus genotypes. The sensitivity of the assay was analyzed by quantified full-length viral RNA and the LOD was set at 10 copies per reaction for all targets. No quenching effects were observed, in case of possible mixed infection. The integrated process control enabled the calculation of recovery rates up to 0.01%.
Significance: The foodproof Hepatitis A virus plus Norovirus (GI, GII) Detection Kit is the first for easy detection in a single assay including a process control. The kit is based on primers, probes, and methods which are mentioned in the ISO/TS 15216.