【Background】The red imported fire ant Solenopsis invicta is a dangerous invasive ant species. A possible means of protection against the ant is the entomopathogenic fungus, Metarhizium anisopliae. Environmental factors can impact the toxicity of M.anisopliae against S.invicta. 【Method】This paper investigated the effect of M.anisopliae M09 on the survival of fire ants exposed to different doses and under various soil moisture conditions and composition. 【Result】The survival rate of fire ant workers decreased with an increasing dosage of M.anisopliae M09; the LC50 was 0.37 g after 4 days. M.anisopliae M09 killed nearly all ants in sandy, loamy and clay soils, and had significant difference from the control. However, the treatment reduced ant numbers more effectively in sandy soil than in loamy or clay soils. The mortality of workers treated by M.anisopliae M09 at different soil moisture were significantly different (P<0.01). 【Conclusion and significance】Soil composition and moisture have significant effect on M.anisopliae against the fire ant, and the soil composition and moisture should be considered when M.anisopliae is applied against the red fire ant.