Abstract: Probiotics, as a potential antibiotic alternative, can improve digestive function, promote growth, and boost the antioxidant capacity and immunity of aquatic animals. To isolate and identify intestinal Bacillus strains from Channa argus and evaluate their potential as probiotics feed additives. Intestinal samples of Channa argus were collected, and bacteria were extracted using the dilution coating method. The species identification was performed using 16S rRNA sequencing and biochemical tests. A variety of experiments were conducted, including high-temperature, acid-base, and bile-salt tolerance assays, adhesion assays, and safety tests. Two Bacillus strains were isolated from the intestinal tract of Channa argus, and after sequencing and BLAST analysis revealed that the strains belonged to the genus Bacillus, which were mainly categorized into Bacillus velezensis and Bacillus subtilis. Strain CA-1 was identified as Bacillus velezensis and strain CA-2 as Bacillus subtilis based on biochemical characterization. Both strains were α-hemolytic, lacked biofilm formation, exhibited hydrophobicity ranging from 57.7% to 73.4%, and showed auto-aggregation from 71.5% to 81.3%. They were able to live in strong acid and alkaline environments as well as in low concentrations of bile salts (0.5%) and withstood high-temperature treatments. Both bacteria were sensitive to the 20 most regularly used antibiotics and inhibited Escherichia coli, Staphylococcus aureus, and Aeromonas hydrophilus. Bacillus velezensis CA-1 and Bacillus subtilis CA-2 demonstrate resilience in harsh conditions, exhibit strong adhesion characteristics, and are relatively safe, making them promising candidates as potential probiotics for Channa argus culture.