Step 1: Virus extraction and concentration
	Sample size	Method
Soft fruits and salad vegetables	25 g/chopped	Elution with agitation followed by precipitation with PEG/NaCl
Bivalve molluscan shellfish	2 g digestive gland from 10 animals	Treatment with a proteinase K solution
Bottled water	Up to 5 L	Adsorption and elution using positively charged membranes followed by concentration by ultrafiltration
Food surfaces	Maximum area 100 cm2	Swabbing
Step 2: RNA extraction
✓ Common to all samples
✓ Reagents should enable processing of 500 μl of extracted virus
✓ Addition of a process control virus
✓ Based on virus capsid disruption with chaotropic reagents and adsorption of RNA to silica particles
Step 3: RTq-PCR
✓ One-step RT-qPCR assay
✓ Reagents should allow processing of 5 μl RNA in 25 μl total volume
✓ Simultaneous monoplex assays for each specific target (NoV GI, NoV GII, HAV and process control virus)
✓ Use of hydrolysis probes
✓ Addition of an external control RNA (purified single-stranded RNA carrying the target sequence for each target virus)
✓ Use of double-stranded DNA control material to make a standard curve
Step 4: Quality control
✓ Virus extraction efficiency should be ≥1%
✓ RT-PCR inhibition should be ≤75%
✓ Amplification efficiencies should range between 90 and 110%