Reproductive isolation between geographically separated populations is
generally considered the most common form of speciation. However,
speciation may also occur in the absence of geographic barriers due
phenotypic and genotypic factors such as chemical cue divergence, mating
signal divergence and mitonuclear conflict. Here we performed an
integrative study based on two genome-wide techniques, 3RAD and
ultraconserved elements, coupled with cuticular hydrocarbon and mtDNA
sequence data, to assess the species limits within the E. ruidum
species-complex, a widespread and conspicuous group of Neotropical ants
for which heteroplasmy has been recently discovered in some populations
from southeast Mexico. Our analyses indicate the existence of at least
five distinct species in this complex, two widely distributed along the
Neotropics and three that are restricted to southeast Mexico and that
apparently have high levels of heteroplasmy. We found that species
boundaries in the complex did not coincide with geographic barriers. We
therefore consider possible roles of alternative drivers that may have
promoted the observed patterns of speciation, including mitonuclear
incompatibility, cuticular hydrocarbon differentiation, and colony
structure. Our study highlights the importance of simultaneously
assessing different sources of evidence to disentangle the species
limits of taxa with complicated evolutionary histories.