Prothioneazole is a broad-spectrum triazole fungicide at present. In order to clear out the bioactivity of prothionazole to Rhizotonia solani caused by rice sheath blight, the mechanism of action of prothioneazole was preliminarily confirmed by determining the contents of DNA, MDA, ergosterol and protein in the hyphae of R. solani, and the changes of defense enzyme activities in rice, the osmotic pressure of mycelial cells and mycelial lipid peroxidation were studied with the method of physiology, biochemistry and molecular biology. The control effects of prothionazole on rice sheath blight were studied by a field experiment and bioassay in vitro. The results showed that prothionazole had high antifungi activity against 94 strains of rice sheath blight, and the average EC₅₀ value (1.062 9 ± 0.795 9) mg·L^-1 of these strains could be used as the baseline of the sensitivity of rice sheath blight fungi to prothioxazole in this area. The changes in DNA, soluble protein and ergosterol contents in the hyphae of R. solani treated with different concentrations of prothioconazole showed that prothioconazole would lead to the reduction of sterols content, the increase of the osmotic pressure of mycelial cells and degree of the mycelial lipid peroxidation, and the decrease of DNA and soluble protein contents. The activities of POD, CAT, SOD and PAL showed an increasing trend under the action of a series of insecticides, and they had certain inductive disease resistance. The results of field efficacy experiment showed that prothionazole suspension at the dosage of 25, 30 and 35 mL per mu in the concentration of 4.8×10⁵ mg·L^-1 had good control effect on rice sheath blight, reaching 90.40%, 96.29% and 98.17%, respectively, and the control effects were similar to those of the control agent, thiofuramide. The safety evaluation of prothionazole on rice showed that different concentrations of prothionazole had no significant effect on the normal rice growth.