The surface of polyvinyl alcohol/chitosan (PVA/Cs) films was modified utilizing facing-dielectric barrier discharge argon plasma driven by alternating current in an atmospheric pressure condition. The surface properties of the films were analyzed using the scanning electron microscope, water contact angle, X-ray photoelectron spectroscopy, and atomic force microscope. The surface energy and corresponding components were evaluated by the Owens–Wendt technique. Additionally, the antibacterial properties of samples were studied using the plate counting method against Escherichia coli and Staphylococcus aureus. After the plasma treatment, a new oxygen-containing hydrophilic group was introduced, and the surface roughness of the film significantly increased. The wettability of PVA/Cs films improved owing to a new polar Osingle bondCdouble bondO functional group and increased surface roughness. With increasing plasma treatment time, up to 10 min, more significant antibacterial efficiency of the films against E. coli (98%) and S. aureus (96%) was observed. The results demonstrate that the increased surface wettability of PVA/Cs films enhanced their antibacterial efficiency against the two microscopic organisms. Additionally, the plasma treatment enabled the strong hydrogen bonds formed between the two polymer chains of the film, resulting in slightly increased of elastic and plastic behaviors.