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The effects of Acidithiobacillus ferrooxidans (At.f) mutated with diethyl sulfate (DES) as a mutagen on the bioleaching of soluble phosphorus (P) from rock phosphate (RP) were investigated. The results show that the oxidative activity of At. f is greatly improved by 1.0% (volume fraction) of DES. Correspondingly, the highest leaching rate of soluble P is also obtained to be 14.9% by the At.f mutated, which is 85.8% higher than that of the adapted At.f without mutation. In addition, the SEM images are significantly performed that the corrosion of RP residue surfaces leached by 1.0% DES-induced At.f is much worse than that of leached by the adapted At. f All the above indicate that the leaching efficiency of soluble P from RP with pyrite can be greatly improved by using DES-induced At.f to a certain extent.