Development of a cellulose nanofiber composite film containing CuO/ZnO nanoparticles and its human norovirus inactivation properties in clams

Human norovirus (HuNoV) threatens human health worldwide, highlighting the critical need for antiviral materials. In this study, copper and zinc nanoparticles (NPs) as inorganic antiviral materials and a cellulose nanofiber (CNF) were selected to construct a hybrid inorganic-organic composite film (CuO/ZnO NPs-CNF) against MS2 and murine norovirus 1 (MNV-1) (surrogates for HuNoV) viruses. Selected NPs concentrations (50 µM of CuO NPs and 5 mM of ZnO NPs) showed non-toxicity (89.88 % of cell viability) through the MTT assay. Upon the development and characterization of the CuO/ZnO NPs-CNF film, high compatibility between CuO/ZnO NPs and CNF was identified, along with consistent thickness, low moisture content, and water solubility. A synergistic antiviral effect was exhibited in vitro and on the film against MS2 and MNV-1 in the mentioned co-treated inorganic matrix within 1 h. The antiviral stability of the film was also maintained at various temperatures (−18 °C, 5 °C, and 25 °C) and humidities (20–30 %, 50–60 %, and 70–80 %). When a synergistic antiviral effect was exhibited on the CuO/ZnO NPs-CNF film against both MS2 and MNV-1, mechanisms analysis by SDS-PAGE and RT-qPCR revealed that there were damages to both capsid protein and RdRP. The application of CuO/ZnO NPs-CNF film on clams resulted in a significant decrease in the survival rate of MS2 by up to 65.67 % and MNV-1 by up to 78.62 % when treated at low temperatures compared to the CNF film. Taken together, these results suggest that the CuO/ZnO NPs-CNF film could be used as potential anti-HuNoV agent in the food industry.