The objective of this study was to develop and characterize hydrogel completes consisting of polyvinyl alcohol (PVA), sodium alginate (SA), and cellulose fiber (CF) incorporated with various concentrations of gamat (sea cucumber) gel and evaluate their antibacterial performance for potential wound dressing applications. Comprehensive characterization of the hydrogels involved incorporating gamat gel at concentrations of 0%, 3%, 5%, and 7% of the total polymer weight, which was followed by measurements including moisture content, swelling ratio, gel fraction, Fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), scanning electron microscopy (SEM), and bacterial penetration tests against Pseudomonas aeruginosa. The hydrogel samples demonstrated a moisture retention of approximately 93%, a swelling ratio ranging from 18% to 23%, and a gel fraction between 44% and 47%. Analysis by FTIR revealed intermolecular interactions and crosslinking between the polymeric components, and XRD patterns showed semicrystalline characteristics within the hydrogel matrix. Micrographs obtained by SEM showed a well-defined porous structure with larger pores and increased interconnection at higher gamat gel concentrations. Results from the bacterial penetration test showed that the incorporation of gamat gel effectively enhanced the hydrogel’s antibacterial barrier, preventing P. aeruginosa for up to 2 days, with optimum performance observed at a concentration of 7%. The results indicate that gamat gel-enriched PVA/SA/CF hydrogels exhibit desirable physicochemical and antibacterial characteristics, positioning them as promising materials for advanced wound dressings with enhanced moisture balance, structural integrity, and resistance to bacterial penetration. © Iran Polymer and Petrochemical Institute 2026.