The design of a new drug material through modification of some well-known antibiotics to combat pathogenic bacteria must include a complete understanding of matrix regulation because the human body consists of primarily three types of matrices, that is, solid, semisolid, and liquid, all of which have a tendency to regulate antibacterial efficacy along with the bactericidal mechanism of several antimicrobial agents. Herein, matrix-dependent action of ciprofloxacin-based polymeric hydrogel scaffold was explored against a new species of Vibrio, namely, Vibrio chemaguriensis Iso1 ( V. chemaguriensis), which is resistant to most of the common antibiotics and possess genes that can be linked to pathogenicity. Ciprofloxacin was attached to the polymeric system consisting of hydrophilic polyethylene glycol methyl ether methacrylate (PEGMA) and zwitterionic sulphabetaine methacrylate (SBMA) with an antifouling nature via covalent linkage leading to effective polymer antibiotic conjugates (PACs) with linear and hyperbranched architectures. The hyperbranched PAC was transformed to a polymeric gel exhibiting greater antibacterial efficacy in solid matrix than that of the liquid one with a complete bactericidal effect and rod to spherical switching of bacterial cell followed by chain formation via "dual" contact activity and release mechanism through sustained removal of thiol-terminated ciprofloxacin fragment along with an equilibrium swelling and deswelling process. Lower killing efficacy was displayed in the liquid matrix with an intact cell morphology that is due to lack of forced contact between the cell wall and gel surface as well as entrapment of released bioactive fragment via an additional thick exopolysaccharide (EPS) layer, which represents a great challenge to modern medical sciences.