The tympanic middle ear (TME) is a complex morphological adaptation that facilitates auditory reception in multiple clades of tetrapods, including anurans. Despite its utility, it has been partially or completely lost at least 38 times in anurans, and there is no clear explanation for how or why this loss occurred. We used micro-CT scans and 3D segmentation to catalog presence/absence and shape of the columella throughout the Central American toad genus Incilius. We found that the TME has been lost multiple times in Incilius and our outgroup Rhinella, with at least one regaining event. Absence of the columella and the different shapes of the columella both have high phylogenetic signal. Incilius species have a different distribution of columella shapes than many other genera of anurans that have been surveyed, including more blade-shaped columellae than found in other genera and one species with vestigial remnants of a columella. The columella shape most common in Incilius, a blade, may be derived from the rod shape also found in many other bufonids; the adaptive significance of this trait remains unknown. This study succeeds in providing a fine-scale review of earlessness and columellar morphology in Incilius, but the processes underlying the patterns we observe remain unclear.

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