Abstract: In order to explore the potential antibacterial medicinal value of insect fungi and find new compounds with novel structures and bioactivity. In this study, the single-cell multi-product strategy(OSMAC)activates were used in liquid culture of Metarhizium guizhouense. The effects of different carbon sources, nitrogen sources, inoculation amount and inoculation time on antibacterial activity of liquid culture extracts of Metarhizium guizhouense were detected. The results showed that the extracts of Metarhizium guizhouense could inhibit the growth of Bacillus cereus, Bacillus lentus and Straphylococcus haemolyticus of 13 common pathogenic bacteria. The minimum inhibitory concentrations of the extracts to 3 pathogenic bacteria were 6.25 mg/mL, 1.56 mg/mL, 6.25 mg/mL, respectively. The effects of different carbon and nitrogen sources on the antibacterial activity of liquid culture extracts of Metarhizium guizhouense was significant, and the extracts of Metarhizium guizhouense grown in starch and potato culture medium showed strongest antibacterial activity. Different inoculation amount had little effects on the antibacterial of extracts of Metarhizium guizhouense. The effects of different culture time on the antibacterial effect of mycelium extract was significant. The antibacterial activity of extract of Metarhizium guizhouense increased with the increase of culture time.