Fusarium wilt caused by Fusarium oxysporum f. sp. niveum is one of the most devastating fungal diseases affecting watermelon (Citrullus lanatus L.). The present study aimed to identify potent antagonistic bacterial strains with substantial antifungal activity against F. oxysporum f. sp. niveum and to explore their potential for biocontrol of Fusarium wilt in watermelon. Out of 77 isolates from watermelon rhizosphere, six bacterial strains—namely, DHA4, DHA6, DHA10, DHA12, DHA41, and DHA55—exhibited significant antifungal activity against F. oxysporum f. sp. niveum, as well as other phytopathogenic fungi, including Didymella bryoniae, Sclerotinia sclerotiorum, Fusarium graminearum, and Rhizoctonia solani. These Gram-positive, rod-shaped, antagonistic bacterial strains were able to produce exo-enzymes (e.g., catalase, protease, and cellulase), siderophore, and indole-3-acetic acid and had the ability to solubilize phosphate. In greenhouse experiments, these antagonistic bacterial strains not only promoted plant growth but also suppressed Fusarium wilt in watermelon. Among these strains, DHA55 was the most effective, achieving the highest disease suppression of 74.9%. Strain DHA55 was identified as Bacillus amyloliquefaciens based on physiological, biochemical, and molecular characterization. B. amyloliquefaciens DHA55 produced various antifungal lipopeptides, including iturin, surfactin, and fengycin, that showed significant antifungal activities against F. oxysporum f. sp. niveum. Microscopic observations revealed that B. amyloliquefaciens DHA55 exhibited an inhibitory effect against F. oxysporum f. sp. niveum on the root surface of watermelon plants. These results demonstrate that B. amyloliquefaciens DHA55 can effectively promote plant growth and suppress the development of watermelon Fusarium wilt, providing a promising agent for the biocontrol of Fusarium wilt in watermelon.