In spite of scientific evidence demonstrating the antiviral activity of lactic-acids bacteria, little is known about the mechanism of their action. Previously, several bacteriocins isolated from lactic acid bacteria (LAB) and some other microorganisms were reported as having antiviral activity in vitro. In the present study, chemically synthetized enterocin B (EntB) and the strain E. faecium L3, known as the producer of this peptide, were tested for activity against influenza viruses. The inhibition of cytopathic effect of А/Perth/16/2009(H3N2) and A/South Africa/3626/2013(H1N1) pdm influenza viruses in MDCK cells by chemically synthetized EntB was revealed. The EntB demonstrated antiviral activity at a concentration of 2.5–5 μg/ml depending on the dose of viruses. This peptide exhibited low toxicity in MDCK cells, causing partial damage of the monolayer of the cells only at a concentration above 10 μg/ml. It was also shown, that strain E. faecium L3-protected mice from lethal A/South Africa/3626/2013(H1N1) pdm infection. We speculate that this protective effect of enterococci may be associated with the specific action of enterocin B, which possesses antiviral activity in vitro.