Ligilactobacillus salivarius UMNPBX2 Cell-Free Extract Has Antiviral Effects on H4N6 Low-Pathogenic Avian Influenza Virus Subtype in Madin–Darby Canine Kidney Cell Line and Embryonated Chicken Eggs

Avian influenza (AI) is a highly infectious disease affecting birds. Some strains of AI virus (AIV) have zoonotic potential, posing a threat to humans. The H4N6 subtype is a low-pathogenic virus and causes mild infection in poultry. However, it has raised increasing concern due to its capability to infect pigs and its high potential for reassortment when co-infected with other strains. This study investigated the antiviral properties of turkey-derived Ligilactobacillus salivarius UMNPBX2 (L. salivarius UMNPBX2) cell-free extract (CFE) using both cell culture and in ovo methods. We assessed the growth kinetics of the H4N6 virus and the cytotoxicity of L. salivarius UMNPBX2 CFE in Madin–Darby Canine Kidney (MDCK) cells. The results revealed that the CFE from the 109 CFU/mL L. salivarius UMNPBX2 overnight culture had strong antiviral activities (p < 0.05). The CFE obtained from 107 to 105 CFU/mL of overnight culture also significantly reduced viral replication (p < 0.05), demonstrating dose-dependent inhibition of viral replication. Additionally, CFEs did not increase pro-inflammatory cytokine gene expression of IL-1β and IL-6 but rather tended to decrease it (IL-6). The embryo survivability experiments revealed a significant dose-dependent increase in survival rate (p < 0.05). The findings of this study highlight the antiviral properties of L. salivarius UMNPBX2 CFE, which contain potential postbiotics against the H4N6 virus, warranting in vivo studies.