Expression and characterization of a novel endolysin LysPFX32 as potential biological antimicrobial agent against Pseudomonas fluorescens for pork preservation.

In this study, a novel phage endolysin LysPFX32 was successfully expressed and characterized to investigate its antibacterial activity against P. fluorescens and its biofilm. The molecular docking results identified endolysin LysPFX32 showed an effective binding to peptidoglycan fragment. The minimum inhibitory concentration of LysPFX32 (95 μg/mL) exhibited strong lytic activity against P. fluorescens after EDTA pretreatment. The permeability of cell outer and inner membrane treated with LysPFX32 was increased. Scanning electron microscope analysis revealed that the cell membrane of P. fluorescens was disrupted by LysPFX32, leading to leakage of intracellular contents. Notably, LysPFX32 effectively inhibited biofilm formation and removed mature biofilm of P. fluorescens by inhibiting exopolysaccharides and total protein. LysPFX32 displayed excellent biological safety with negligible hemolysis in mouse red blood cells and lack of cytotoxicity against NIH 3T3 cells. LysPFX32 effectively eradicated P. fluorescens in pork at 28 °C after 24 h. The texture and color difference of pork with added LysPFX32 did not exhibit significant alterations. During 8 d storage, the LysPFX32-treated group exhibited a reduction in amine production and maintained meat quality. This study highlighted the remarkable effectiveness and diverse potential applications of phage endolysin, offering a promising approach for controlling P. fluorescens contamination in food.