Plasma-activated water(PAW) was prepared by using atmospheric pressure plasma jet technology with compressed air at 0.15 MPa as the working gas. PAW was activated for 30 s， 60 s， and 90 s at a power of 380 W. The physicochemical properties of PAW were analyzed， and the inhibitory effects of different plasma-activated waters on various spoilage bacteria (Hafnia alvei， Hafnia paralvei， and Serratia grimesii) isolated from spoiled salmon as well as Escherichia coli and Staphylococcus aureus were studied. The inactivation kinetics model of PAW was analyzed using the GInaFIT Tool. The integrity of cell membrane and changes in intracellular nucleic acids after sterilization were determined. Furthermore， the inhibitory and dispersion effects of PAW on the biofilm of H. alvei， H. paralvei， and S.grimesii were investigated. It was found that the pH value of PAW gradually decreased with increasing discharge time， while the redox potential， conductivity， and content of H2O₂， O3， NO₂-， and NO3- increased. PAW activated for 60 s could completely kill the pathogenic bacteria after 6 min of treatment. PAW60 and PAW90 exhibited a better fit to the logarithmic model when treating H. alvei A2， H. paralvei A3， and S.grimesii A5 for 6 min. Cell integrity was disrupted， and DNA leakage occurred after PAW treatment. The formation of biofilms by the pathogenic bacteria mentioned above could be inhibited by PAW treatment for approximately 2 min， and the dispersion of mature biofilms required 20-30 min. This study provides a theoretical basis for using plasma-activated water to inhibit pathogenic and spoilage bacteria in seafood products.