Single-use synthetic plastics that are used as food packaging is one of the major contributors to environmental pollution. Hence, this study aimed to develop a biodegradable edible film incorporated with Limosilactobacillus fermentum. Investigation of the physical and mechanical properties of chitosan (CS), sodium caseinate (NaCas), and chitosan/sodium caseinate (CS/NaCas) composite films allowed us to determine that CS/NaCas composite films displayed higher opacity (7.40 A/mm), lower water solubility (27.6%), and higher Young’s modulus (0.27 MPa) compared with pure CS and NaCas films. Therefore, Lb. fermentum bacteria were only incorporated in CS/NaCas composite films. Comparison of the physical and mechanical properties of CS/NaCas composite films incorporated with bacteria with those of control CS/NaCas composite films allowed us to observe that they were not affected by the addition of probiotics, except for the flexibility of films, which was improved. The Lb. fermentum incorporated composite films had a 0.11 mm thickness, 17.9% moisture content, 30.8% water solubility, 8.69 A/mm opacity, 25 MPa tensile strength, and 88.80% elongation at break. The viability of Lb. fermentum after drying the films and the antibacterial properties of films against Escherichia coli O157:H7 and Staphylococcus aureus ATCC 29213 were also evaluated after the addition of Lb. fermentum in the composite films. Dried Lb. fermentum composite films with 6.65 log10 CFU/g showed an inhibitory effect against E. coli and S. aureus (0.67 mm and 0.80 mm inhibition zone diameters, respectively). This shows that the Lb.-fermentum-incorporated CS/NaCas composite film is a potential bioactive packaging material for perishable food product preservation.