BACKGROUND: Influenza virus is a significant cause of annual global respiratory disease and death. According to the limited availability of effective drugs and vaccines, innovative antivirals are currently being investigated as possible strategies to contain the spread of infectious agents. Among the various types of nanoparticles, silver nanoparticles (Ag-NPs) have attracted great interest due to their exceptional physicochemical properties. This study aims to investigate the antiviral activity of Ag-NPs against the influenza A virus (IAV)/H1N1. METHODS: The MTT assay was used to determine the possible cytotoxicity of the Ag-NPs. Madin-Darby canine kidney (MDCK) cells were exposed to Ag-NPs extract in conjunction with 100 cell culture infectious dose 50% (CCID50) of virus administered at time intervals during the infection process. The antiviral activity of the extract was evaluated under pre-treatment, post-treatment, and simultaneous assay. Viral titer reduction was assayed using hemagglutination (HA) and CCID50 assays. Viral RNA relative quantification by real-time Polymerase Chain Reaction approach was performed in each experimental condition. RESULTS: The study yielded significant findings regarding the inhibitory effects of Ag-NPs on the IAV/H1N1. Silver nanoparticles showed dose-dependent inhibition of the virus, with the strongest effect observed when administered simultaneously with the virus which the virus titer exhibited a substantial decrease from 5 Log10 in the control group to 1 Log10 in the initial samples, further reducing to 2 Log10 per milliliter at lower concentrations. Notably, Ag-NPs demonstrated a greater reduction in virus titer during the simultaneous stage, showing a statistically significant difference (P <
0.05) between the control and experimental groups). The reduction in viral titer was also evident in both pre- and post-inoculation stages, although the effects were different. CONCLUSION: Silver nanoparticles possess inhibitory effects on the growth of the IAV/H1N1, with a significant reduction in virus titer. These findings suggest the potential of Ag-NPs as effective antiviral agents and highlight opportunities for further research and potential clinical applications in combating IAV (H1N1) infections.