From June to August in 2004 and 2005, we conducted the studies on spontaneous vocalization development of greater horseshoe bat, Rhinolophus ferrumequinum, in Zhi’an Village (Jilin province, Northeast China). In contrast to adult bats, infant bats of the greater horseshoe bat emitted calls characterized by multiharmonics and variable harmonic patterns. With the physical growth of infants, the dominant frequency, pulse duration and frequency of each harmonic of spontaneous calls increased, the number of harmonics decreased from 5–8 to 1–2 and dominant harmonic switched from first to the second with peak frequency increasing. Vocalizations of infant bats of the greater horseshoe bat could be categorized to those serving as precursors of echolocation sounds and those serving as isolation calls used to attract their mothers. According to observation on mother-infant reunion, the female adult bats only suckled their own babies, but not other pups in the same colony. And the mother recognized their own infants through both odor and vocal cues indicating that the isolation calls emitted by infant bats played an important role in mother-infant communication.