The characteristics of fatigue crack growth were investigated in Ti-6Al-4V alloy with three kinds of microstructure prepared with different heat treatments. The effect of microstructure on the crack growth behavior was attributed to the development of crack tip shielding, primarily resulting from the role of the crack path morphology in inducing crack closure and crack deflection. Roughness induced crack closure played an important role on the near-threshold growth behavior at the load ratio of 0.05, but the growth rate data plotted in terms of effective stress intensity factor range ΔKeff (after allowing for closure) still exhibited the effect of microstructure. Fractographic observations were performed using a scanning electron microscope (SEM) with the aid of image processing, which enables the three dimensional observation using a stereo pair of SEM micrographs. The roughness was evaluated quantitatively by the ratio of the total area of the three dimensional fracture surface to its projected area. As the fracture surface roughness was taken into account in re-evaluating the crack growth characteristics, the effect of microstructure disappeared, indicating that the intrinsic crack growth resistance was the same for all microstructure. Thus, fracture surface roughness is a controlling parameter of the fatigue crack growth in Ti-6Al-4V alloy. Furthermore, the crack growth rate were compared with those of other structural materials such as steels, nodular cast iron and an aluminum alloy.