In the present study, extraction of chitin from shrimp (Metapenaeopsis  stridulans) wastes was performed chemically or biologically (using Bacillus subtilis (B) and Lactobacillus plantarum (L) via single-step (L or B) or successive co-fermentation techniques (L/B or B/L)). The proximate composition, physicochemical properties, and antibacterial activity of the prepared chitosan samples were determined. Although the bio-extracted chitosan samples (3.59-6.64%) yields were lower than the chemically extracted samples (8.44%), the bio-extracted samples showed better quality. The DDA and CrI % of the biologically and chemically extracted chitosan samples were (79.5-80.35%) and (71.0-77.7%), respectively. The intrinsic viscosity and molecular weights of the bio-extracted chitosan (B, L, B/L, and L/B) samples (η= 0.0779, 0.0847, 0.0943 and 0.0909 dl/g, Mw=26.37, 30.17, 35.09 and 33.32 kDa, respectively) were lower than the chemically extracted samples (η=0.1903 dl/g, Mw=94.34 kDa). XRD revealed the crystalline nature of all the polymers. The bio-extracted chitosan samples showed greater inhibition zones (IZ) against B. cereus. Chitosan sample (B) exhibited the highest antimicrobial activity against B. cereus, Salmonella sp. and Vibrio sp. The present findings proved the potential use of bio-extracted chitosan as an effective, safer, and more natural preservative in the food industry to combat foodborne pathogens.