To obtain effective biocontrol strains for downy mildew of grape, 38 endophytic bacteria were isolated from fruits, seeds, and old stems of six grape varieties. Using spot inoculation mixtures of sporangial suspensions of Plasmopara viticola and biocontrol bacterial suspension, this screen yielded three strains (G1, G5, and G9) with good antagonistic effects against P. viticola. The growth inhibition rate was 100%, which was comparable to the effect of the positive control Bacillus subtilis strain CN181. The enzyme activity and the metabolites of strain G1 were examined on casein hydrolysate medium, sodium carboxymethyl cellulose agar plates, and chrome azurol sulfonate (CAS) agar plates. The antifungal protein component was identified by liquid chromatography–mass spectrometry (LC–MS). The results showed that strain G1 was more effective against Plasmopara viticola after two field trials, and the inhibition rates of strain G1 on the seventh day of the two field trials were 47.5% and 36.9%, respectively. Strain G1 was identified as Bacillus amyloliquefaciens based on morphological examination and 16S rDNA sequence analysis. It produced proteases, cellulases, and siderophores. Crude protein of the strain mainly included the putative segregation protein SpoVG, which inhibited P. viticola.