The aim of this study was to acquire endospore-former(s) for aquatic animal feed based on the probiotic potential in vitro, including the anti-pathogen spectrum, gastrointestinal fluid tolerance, antioxidant activity, enzyme-producing ability, and basic safety assessment. The strain Bacillus velezensis FJAT-57093 was found to exhibit the strongest antibacterial ability against Aeromonas hydrophila in the agar well diffusion inhibition assays from 111 Bacillus-like strains. Moreover, the results showed that the compounds of the strain FJAT-57093 enriched by acid precipitation might be the main antibacterial metabolites. The strain FJAT-57093 also exhibited antibacterial effects against the aquatic pathogens Photobacterium damselae, Edwardsiella tarda, Vibrio parahaemolyticus, and Vibrio vulnificus. The safety assessment revealed that the strain FJAT-57093 was non-hemolytic and susceptible to ten antibiotics. The putative virulence and antibiotic resistance genes predicted were predominantly intrinsic to the FJAT-57093 genome. Furthermore, the strain FJAT-57093 demonstrated a tolerance of acid and bile salt under the simulated gastrointestinal tract conditions, extracellular enzyme-producing abilities, as well as an auto-aggregation rate of 45.88% at 24 h and co-aggregation rates with the aforementioned five aquatic pathogens, ranging from 14.87% to 58.55%. Additionally, its extracellular metabolites displayed strong antioxidant activities, with ABTS⁺ and DPPH radical scavenging rates of up to 99.82% and 42.74%, respectively. In summary, the strain B. velezensis FJAT-57093 was found to possess strong antibacterial activities against multiple aquatic pathogens and desirable in vitro probiotic properties.