Recombinant Expression and Activity Analysis of a Vibrio alginolyticus Phage-Derived Endolysin LysV039C

Pathogenic Vibrio species are a major cause of disease outbreaks in aquaculture, leading to substantial economic losses and posing risks to food safety. The increasing prevalence of antibiotic resistance among these pathogens has created an urgent need for alternative antimicrobial strategies. Phage-derived endolysins, which specifically degrade bacterial peptidoglycan, have emerged as promising candidates to replace or supplement conventional antibiotics. In this study, we systematically characterized the endolysin LysV039C from the Vibrio alginolyticus phage phiV039C. Whole-genome analysis identified a 462 bp endolysin gene, which was cloned into the pET-28a vector, expressed in E. coli BL21, and purified. The lytic activity of the recombinant enzyme was evaluated using a turbidity reduction assay. LysV039C showed the strongest activity (58.9%) against host bacteria in the logarithmic phase, with peak activity (60.7-64.1%) achieved at 53 μg/mL, 35 °C, and neutral pH. The addition of 2.5 mM EDTA enhanced activity to 63.8%, whereas 10 mM divalent metal ions strongly inhibited the enzyme (<6.5%). Lytic spectrum analysis demonstrated that LysV039C exhibited a broader lytic spectrum against six Vibrio species than its parent phage phiV039C. Overall, LysV039C combines high environmental adaptability (alkaline tolerance, suitability for aquaculture temperature ranges) with efficient and broadened lytic activity against multiple Vibrio species. These findings provide a foundation for developing eco-friendly agents for the prevention and control of pathogenic Vibrio and offer a valuable reference for the mining of other phage-derived antibacterials against aquatic pathogens.