To keep the safety of oyster intended for raw eating, monitoring of norovirus (NoV) contamination by rapid and sensitive method is important. We had developed a rapid method using bead homogenization to recover NoV particles from oyster tissues followed by RNA detection using reverse-transcription, real-time PCR. But there were still remains a need to improve the sensitivity. Therefore we re-evaluated bead homogenization conditions and studied the usefulness of enzymatic digestion after the bead homogenization. In the comparison of homogenization condition (speed and time), the mildest condition (3,500 rpm for 15 sec) showed the highest positive rate (60%) of NoV. On the contrary, when proteinase K digestion was added after bead homogenization, high intensity homogenization at 4,500 rpm for 15 sec showed high positive rate compared with the low intensity homogenization. In the comparison of digestion with two different enzymes, proteinase K and α-amylase, that digest organic compounds in digestive tissues after the bead homogenization, α-amylase digestion showed higher NoV GII RNA copy number (p<0.01) in average. At least 60-min digestion time by α-amylase was required. These results show that the bead homogenization method coupled with α-amylase digestion is useful to recover NoV particles in oyster digestive tissues.
