Echolocation clicks can reflect the anatomy of the vocalizing animal, enabling the distinction of species. River dolphins from the family Iniidae are formally represented by one species and two subspecies (Inia geoffrensis geoffrensis and I. g. humboldtiana). Additionally, two other species have been proposed (I. boliviensis and I. araguaiaensis) regarding its level of restricted distribution and morph-genetics differences. For the Committee on Taxonomy of the Society for Marine Mammalogy, the specific status of the proposed species relies on further knowledge on morphology, ecology, and genetics. Given that species-specific status is required for conservation efforts, we described and compared the echolocation clicks of Inia spp., searching for specific differences on their vocalizations. The sounds were captured with a Cetacean Research ™ C54XRS (+3/−20 dB, −185 dB re: 1V/μPa) in Guaviare River (Orinoco basin), Madeira River (Madeira basin), Xingu River (Amazon Basin), and Araguaia River (Tocantins-Araguaia basin). We found significant differences in all analyzed parameters (peak frequency, 3 dB bandwidth, 10 dB bandwidth and inter-click interval) for all species and subspecies. Differences in acoustical parameters of clicks are mainly related to the animal’s internal morphology, thus this study may potentially support with information for the species-level classification mostly of I. araguaiaensis (the Araguaian boto). Classifying the Araguaian boto separately from I. geoffrensis has important implications for the species in terms of conservation status, since it is restricted to a highly impacted river system.