Social insects communicate by using chemical, visual, tactile, and acoustic signals, including 19 stridulations. Ectatomma ruidum is a mainly Neotropical ant species complex that has faced 20 strong divergence at the genetic level; the species have a highly variable blend of cuticular 21 hydrocarbons and a relatively conserved morphology. Based on evidence for genetic and 22 chemical differentiation, we tested for variation in acoustic traits. We compared the 23 stridulations produced by the species E. ruidum sp. 2, sp. 3-4 and the new putative species E. 24 ruidum sp. 5, as well as the morphology of the stridulatory file. We found that the stridulations 25 produced by E. ruidum sp. 5 were statistically different from those of the other species in a 26 number of traits. The differences in stridulatory traits might rely more on the way the ants 27 produce the sound (rubbed area percentages) than on the morphology of the stridulatory file, 28 for which we did not find variation. Our results highlight the use of acoustic traits as potential 29 taxonomic tools for integrative taxonomic studies and suggest that the acoustic traits of E. 30 ruidum species complex have been subjected to selection.