Chitin and chitosan have been proved to have enormous applications in biomedical, pharmaceutical, and industrial fields. The horse mussel, Modiolus modiolus, a refuse of the fishery industries at Thondi, is a reserve of rich chitin. The aim of this work is to extract chitosan from the horse mussel and its further characterization using Fourier transform infrared spectroscopy (FTIR), micro-Raman spectroscopy, X-ray diffraction (XRD), and elemental analysis. The result of FTIR studies revealed different functional groups of organic compounds such as out-of-plane bending (564 cm–1), C–O–C stretching (711 cm–1), and CH2 stretching (1174 cm–1) in chitosan. The degree of acetylation of the extracted chitosan was observed to be 57.43%, which makes it suitable as a biopolymer for biomedical applications. Prominent peaks observed with micro-Raman studies were at 484 cm–1 (14,264 counts/s), 2138 cm–1 (45,061 counts/s), and 2447 cm–1 (45,636 counts/s). XRD studies showed the crystalline nature of the polymer, and the maximum peak was observed at 20.04°. Elemental analysis showed a considerable decrease in the percentage of nitrogen and carbon upon the conversion of chitin to chitosan, while chitosan had a higher percentage of hydrogen and sulfur. The antibacterial activities of chitosan from the horse mussel were found to be efficient at a 200 μg/mL concentration against all the bacterial strains tested with a comparatively higher antibacterial activity against Escherichia coli (9 mm) and Bacillus subtilis (8 mm).