Spectral and temporal features of echolocation calls produced by 15 insectivorous bat species in three families from Madagascar are described. In addition we provide a library of bat vocalizations that can be used for acoustic inventories involving heterodyne and time-expansion bat detectors. Time-expanded recordings of calls from 153 bats from 15 species were analyzed using five commonly used temporal and frequency variables measured from spectrographs. Echolocation calls for six species (Scotophilus tandrefana, S. marovaza, Emballonura tiavato, Neoromicia spp., N. malagasyensis and Triaenops auritus) are described for the first time. A discriminant function analysis revealed that a function based on the five measured variables provided a correct overall classification of 82.2%. Three groups of echolocation calls based upon the temporal and frequency characteristics of calls are recognized. The Constant Frequency group consists of hipposiderids and Emballonura spp., the Frequency Modulated/Quasi-Constant Frequency group is dominated by vespertilionids, and one species, Myotis goudoti, is in the Frequency Modulated group. Further we describe the utility of using acoustic sampling in inventory and monitoring studies, and in investigations of habitat use.