Using whole cells as antigens, specific egg yolk antibodies (IgY) were generated respectively against Shewanella putrefaciens and Pseudomonas fluorescens, two important spoilage bacteria in aquatic products. The obtained IgY exhibited a high and specific affinity to corresponding antigen bacteria. A concentration-dependent antimicrobial activity of the specific IgY was observed in liquid medium. Compared to control groups, the cell number of S. putrefaciens and P. fluorescens was reduced approximately 84.7% and 88.1%, respectively, in the presence of specific IgY (contain salts) at a concentration of 200 mg/mL after 8 h of incubation. Similar growth inhibitory effects were also observed in solid medium, in which the inhibition ratio was calculated as 52.8% and 62.5% for S. putrefaciens and P. fluorescens, respectively, in the presence of specific IgY at a concentration of 100 mg/mL. These results indicated a great potential of specific IgY as a safe and natural antimicrobial agent for aquatic food preservations.