Bacillus subtilis strain Ydj3 was applied to sweet peppers to understand the influence of this bacterium on the growth, fruit quality, and rhizosphere microbial composition of sweet pepper. The promotion of seed germination was observed for sweet pepper seeds treated with the Ydj3 strain, indicating that Ydj3 promoted seed germination and daily germination speed (131.5 ± 10.8 seeds/day) compared with the control (73.8 ± 2.5 seeds/day). Strain Ydj3 displayed chemotaxis toward root exudates from sweet pepper and could colonize the roots, which enhanced root hair growth. Following the one-per-month application of strain Ydj3 to sweet pepper grown in a commercial greenhouse, the yield, fruit weight, and vitamin C content significantly increased compared with those of the control. Additionally, the composition of the rhizosphere bacterial community of sweet pepper changed considerably, with the Bacillus genus becoming the most dominant bacterial genus in the treated group. These results suggested that B. subtilis Ydj3 promotes seed germination and enhances fruit quality, particularly the vitamin C content, of sweet pepper. These effects may be partly attributed to the B. subtilis Ydj3 colonization of sweet pepper roots due to Ydj3 chemotaxis toward root exudates, resulting in the modulation of the rhizosphere bacterial community.