Synergistic Effects Between Chitooligosaccharide-Epigallocatechin Gallate Conjugate and Nisin on Inhibition of Seafood-Associated Vibrio spp., Aeromonas spp., and Shewanella spp.

Ready-to-cook Asian moon scallop (RAM) may harbor seafood-associated spoilage and opportunistic pathogenic bacteria, leading to quality deterioration and food safety concerns. This study investigated the microbiological diversity, virulence-associated phenotypes, and antibacterial activity of chitooligosaccharide-epigallocatechin gallate conjugate (CEGC) combined with nisin against seafood-associated bacteria isolated from RAM. A total of 102 bacterial isolates were subjected to identification by MALDI-TOF/MS, with Vibrio, Aeromonas, and Shewanella as the predominant genera. Several isolates exhibited proteolytic and hemolytic activities, indicating spoilage potential and virulence-associated traits. Multivariate analysis showed that bacterial isolates differed widely in their antimicrobial susceptibility and virulence-associated phenotypes. Checkerboard assays demonstrated synergistic effects between CEGC and nisin against Vibrio parahaemolyticus and Vibrio fluvialis (FICI = 0.50). The additive effects were observed for the remaining strains, except an indifferent effect (FICI = 1.06) was found for A. hydrophila. The combined treatment markedly inhibited bacterial growth and biofilm formation, achieving approximately 95% biofilm reduction in V. parahaemolyticus. Confocal laser scanning microscopy and scanning electron microscopy revealed severe membrane disruption, increased permeability, cellular deformation, and collapse of biofilm architecture. Furthermore, CEGC-nisin effectively suppressed V. parahaemolyticus growth in cold-stored RAM. These findings demonstrate that CEGC combined with nisin is a promising natural antimicrobial strategy for improving seafood safety and reducing microbial spoilage in ready-to-cook seafood products.