Bletilla striata extracts were prepared by methanol circumfluence extraction. Five different polarity fractions were obtained by extraction in turn with n-hexane, dichloromethane, ethyl acetate, n-butanol and water. The inhibiting effects on E. coli, S.aureus, B.subtilis and P.aeruginos were measured by using filter paper disc diffusion method and minimum inhibitory concentration (MIC) method. The chemical constituents of the active fraction were analyzed by gas chromatography-mass spectrometry (GC-MS). The results showed that the methanol extract, n-hexane and dichloromethane fractions exhibited significant inhibition against four common bacterial strains. Dichloromethane fraction presented the best antibacterial effect than other fractions (P＜0.05). Dichloromethane fraction exerted potent inhibitory effects on E. coli, S.aureus, B.subtilis and P.aeruginos in a dose-dependent manner. Its MIC against four common bacterial strains was 2 mg/mL. Eleven compounds representing 94.16% of the total constituents were identified by GC-MS analysis. The main constituents of dichloromethane fraction were 2-methyl-3,4-benzophenanthrene (39.97%), ethyl homovanillate (25.25%), trans-4′-fluoro-4-(methylthio) chalcone (9.44%), dioctyl sebacate (7.34%), and linoleic acid (3.86%). In conclusion, B.striata fractions exerted potent antibacterial activity. Dichloromethane fraction was found to be the most active, and aromatic series, esters, organic acids might be potential bioactive components.