| Summary: | <i>Fusarium oxysporum</i>, a primary soil-borne fungus that affects rice seedlings globally, is responsible for rice seedling blight (RSB), which reduces seedling quality and survival rates. The synthetic fungicides used to treat this disease negatively affect human health and the environment. A biocontrol bacterial isolate, <i>Bacillus velezensis</i> Bv S3, isolated from the rice rhizosphere, showed a strong antagonistic effect on RSB-causing <i>F. oxysporum</i>. The ratio of the longest to the shortest radius of <i>F</i>. <i>oxysporum</i> following Bv S3 inoculation was 2.52 (cm/cm) in a plate standoff experiment. This was different from the other biocontrol strains. Bv S3 exhibits a wide spectrum of antifungal activity against various pathogenic fungi that cause RSB. When 10% Bv S3 liquid culture filtrate was applied, it dramatically reduced <i>F. oxysporum</i> spore germination and mycelial growth, with inhibition rates of 66.7%, and 45.7%, respectively, and caused hyphal malformations. Furthermore, the Bv S3 suspension (1 × 10<sup>8</sup> CFU/mL) reduced RSB by 65.5% and 76.5% in pot experiments, effectively promoted the growth of rice seedlings, and improved the activities of neutral phosphatase, urease, invertase, and catalase in rice rhizosphere soil. The active substances produced by Bv S3 were sensitive to temperature and ultraviolet irradiation, and the antifungal effect significantly increased after 90 min of exposure, with antifungal effect observed at pH 7. Bv. S3 effectively reduced the incidence of RSB and showed potential as a biocontrol agent.
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