In order to investigate the effects of high voltage atmospheric cold plasma (HVACP) treatment on the number of microorganisms in and the quality of Trachinotus ovatus during refrigerator storage, fresh fish was packaged with gases CO2:O2:N2 (80%:10%:10%) and treated by HVACP at 75 kV for 3 min; then, the samples were stored at 4 ± 1 °C for nine days. The microbial numbers, water content, color value, texture, pH value, thiobarbituric acid reactive substance (TBARS), and total volatile base nitrogen (TVB-N) values of the fish were analyzed during storage. The results showed the growth of the total viable bacteria (TVB), psychrophilic bacteria, Pseudomonas spp., H2S-producing bacteria, yeast, and lactic acid bacteria in the treated samples was limited, and they were 1.11, 1.01, 1.04, 1.13, 0.77, and 0.80 log CFU/g−1 lower than those in the control group after nine days of storage, respectively. The hardness, springiness, and chewiness of the treated fish decreased slowly as the storage time extended, and no significant changes in either pH or water content were found. The lightness (L*) value increased and the yellowness (b*) value decreased after treatment, while no changes in the redness (a*) value were found. The TBARS and TVB-N of the treated samples increased to 0.79 mg/kg and 21.99 mg/100 g, respectively, after nine days of refrigerator storage. In conclusion, HVACP can limit the growth of the main microorganisms in fish samples effectively during nine days of refrigerator storage with no significant negative impact on their quality. Therefore, HVACP is a useful nonthermal technology to extend the refrigerator shelf-life of Trachinotus ovatus.