The study aimed to investigate the optimal preparation process of clove essential oil-polyvinyl alcohol (PVA) slow-release bacteriostatic microspheres and their preservation effect on blueberries. Taking Escherichia coli, Staphylococcus aureus, and Penicillium as target strains commonly found in blueberries after picking, the essential oils with the best antibacterial effect were screened out by using the two-fold dilution method, and the best preparation process conditions of the microspheres were obtained by optimising the microspheres through response surface experiments using the microsphere embedding rate as an index. The structure and performance of the microspheres were characterised and applied to blueberries to test the effect of theirfreshness preservation. The results showed that clove essential oil had a better inhibitory effect on the three strains of bacteria; the optimal preparation process: 4% PVA solution, 2g Span 80,emulsification temperature 48 ℃, the microsphere embedding rate of up to 72.69%; scanning electron microscopy showed that the microsphere surface was smooth and regular spherical, withan average particle size of 43.16 μm, and the comparison of infrared spectral graphs of the PVA microspheres, the PVA raw material and the essential oil of clove shows that the clove essential oil was wrapped by PVA; in anhydrous ethanol and 3% acetic acid solution, the release rate of the microspheres was above 60%, which possessed the slow-release property; compared with the control group, the changes of various quality indexes of blueberry fruits in the microsphere-treated group were better than those in the control group. In conclusion, the clove essential oil-PVA slow-release bacteriostatic microspheres could delay the spoilage of blueberries and effectively maintain the quality of blueberries.