This study was performed to investigate the effect of defect on fatigue strength under the stress of rotary bending. The specimens were made of low carbon steel having artificial microholes, namely, single micro-hole and two adjacent micro-holes as natural defects, and the effects of the diameter of hole and the distance between the holes on fatigue strength have been investigated. The obtained result can be summarized as follows: 1, The critical defect means the largest size of defect that does not affect fatigue limit, and correspondes to the size of defect leading to final fracture under fatigue limit of smooth specimen. The size of defect which has an effect on fatigue limit is larger than that of critical defect. 2, The defect larger than the critical defect affects fatigue strength for as a kind of size effect, and the physical meaning of size effect of defect is considered same as the one of notch effect.