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Volatiles produced by Bacillus subtilis strain Tpb55 inhibited sporulation, mycelial growth and pigment biosynthesis of Alternaria alternata, a pathogen of tobacco brown spot. Under enclosed experiment conditions, the volatiles of Tpb55 showed 100% efficiency in control of tobacco brown spot. Compared with the control, volatiles of Tpb55 significantly increased the plant height, leaf blade length and fresh weight of tobacco, and no obvious adverse effect on tobacco was found. Results from this research showed that volatiles produced by Tpb55 effected well on the inhibition of pathogen, the control of disease and the promotion of tobacco growth. Bacterial volatiles provide a new source of compounds with antibiotic and growth-promoting features.