The effects of microstructure on the fatigue crack growth behavior of surface‐ and middle‐layer materials of high‐strength bridge steel (including steel Q370qE, Q420qE, and Q500qE) plates were investigated. The microstructures are captured via an optical microscope, and fracture morphologies are examined by scanning electron microscopy. The results indicate that for materials with different strength grades of steel plates at the same sampling locations, the higher the strength grade, the lower the fatigue crack growth rate (FCGR). In addition, materials with smaller equivalent grain sizes or higher volume fractions of the second hard phase exhibit lower FCGR for the surface‐ and middle‐layer materials of the identical strength‐grade steel plate. The fatigue fracture morphological examination results show that typical fatigue striations along with secondary cracks are observed in the stable crack growth region of three high‐strength bridge steel plates. Moreover, fatigue steps are observed in steel Q500qE, which has the lowest FCGR, indicating that the fatigue steps contribute significantly to resisting the fatigue crack propagation.