Tobacco mosaic virus (TMV) is a major factor leading to the yield reduction of Solanaceae crops. In order to explore edible fungus-derived polysaccharide materials for the prevention and control of Tobacco mosaic virus, the control effect of six crude polysaccharides against TMV was determined by the half-leaf method, five-point injection test and systematic host infection activity evaluation. Among them, the extraction rate of crude polysaccharide from Agaricus blazei, 24.32%, was the highest, followed by Pleurotus ostreatus 18.56%, and that from Pleurotus eryngii, 10.04% was the lowest. The total sugar content of A. blazei was significantly higher than that of other species, reaching 37.71%, followed by that of P. igniarius and G. Lucidum, exceeding 32.00%. In addition, the sulfate content of A. blazei crude polysaccharide was also as high as 2.10%. In this research, the inhibitory effect of the crude polysaccharides was determined by the half leaf method. Among the 6 tested polysaccharide treatments, the polysaccharides that were inoculated after treated for 48 h had a good inhibition on TMV. The preventive effects of P. ostreatus, C. versicolor and A. blazei reached more than 60.00%. Especially, the preventive effect of 24 mg/mL A. blazei crude polysaccharide was up to 92.04%, and the number of dead spots on the leaves was significantly less than those in the clear water control group. In the five-point injection test, the inhibition rate of A. blazei crude polysaccharide was 38.18%. The brightness of the fluorescent spots was weaker than that in the control group, the fluorescent area did not change much, and did not appeared in the new leaves. A. blazei crude polysaccharide had an inhibitory effect in the early stage of virus infection, which could delay the moving speed of TMV and slow down the system invasion process. On NC-89, the control effect of agaricus antler polysaccharide on TMV was as high as 76.81%, which was better than that of 0.5% lentinan water agent and 1.8% octinamine acetate water agent. When the concentration was 48 mg/mL, the plants were damaged, and the excessive concentration of polysaccharide formed precipitation on the surface of the plants, causing the plants to yellow. Field test showed that the treatment effect of A. blazei crude polysaccharides on TMV was 70.31%. The plants with twisted, curled leaves and stopped growing had an obvious improvement. At the same time, it had a certain inhibitory effect on TMV, the onset time of sprayed tobacco was delayed, the tobacco grew well, and the leaves were extended. The results of this study would lay a foundation for further study on the action mode and mechanism of fungal polysaccharides against Tobacco mosaic virus.