Abstract: In order to investigate the effect of different chemicals and biological fungicides on Valsa ceratosperma, four kinds of biological fungicides: Paenibacillus polymyxa, Bacillus subtilis, polymyxin and matrine, and two kinds of chemicals: tebuconazole and prochloraz, were used as materials to study the inhibition effect of single fungicide and combinations of different fungicides on Valsa ceratosperma mycelial growth and conidial germination in vitro culture of dish way. The results showed that inhibition rate reached 100% in 180 mg/L Bacillus subtilis, 180 mg/L Paenibacillus polymyxa, 225 mg/L polymyxin, 5 mL/L matrine, 0.75 mL/L tebuconazole and 0.75 mL/L prochloraz when a single fungicide was added in the medium. In addition, the combination of 0.02 mL/L prochloraz with 45 mg/L Paenibacillus polymyxa, Bacillus subtilis, 0.045 mg/L polyoxin, as well as the combination of 0.02 mL/L tebuconazole with 45 mg/L Bacillus subtilis showed better inhibition rate than other combinations of fungicides, and the inhibition rates of mycelia and spore were up to 100% in 4 combinations. It is indicated that the 4 combinations of chemicals and biological fungicides were effective and low toxic, and could be used to prevent Valsa ceratosperma.