Species boundaries in <italic>Philaethria</italic> butterflies: an integrative taxonomic analysis based on genitalia ultrastructure, wing geometric morphometrics, DNA sequences, and amplified fragment length polymorphisms

Abstract

Neotropical passion-vine butterflies in the tribe Heliconiini (Lepidoptera: Nymphalidae) are a major focus of research in ecology and evolution because of their diverse, aposematic wing patterns, extensive Müllerian mimicry, and coevolution with their Passifloraceae host-plants. However, the basic taxonomy of this group, which is essential to evolutionary ecology research, has been built over the last two centuries using primarily gross morphological comparisons, with most species identification being based on wing colour pattern variation. For some taxa, such as the genus Philaethria Billberg, even the most basic information, such as species limits and geographical distributions, remains uncertain. Furthermore, descriptions of new species, within Philaethria and beyond, have generally been based on small sample sizes collected from a restricted area of the full geographical distribution. To address these issues in the genus Philaethria, here we used an integrative taxonomic approach involving both morphology (genitalia ultrastructure; linear and geometric morphometric analyses of wing shape) and molecular data (multilocus DNA sequence data and amplified fragment length polymorphisms). Specifically, we tested the taxonomic validity of two Philaethria species, Philaethria pygmalion and Philaethria wernickei, described in the literature as having disjunct distributions, corresponding to the Amazon Basin and the Atlantic Rain Forest of Brazil, respectively. Our analyses revealed that these two Philaethria species cannot be delimited and diagnosed using metric and nonmetric morphological characters. Furthermore, they occur in sympatry in the Cerrado biome of central Brazil, and appear to form a latitudinal cline in wing colour variation across their combined distribution. These results are further supported by limited genetic differentiation and a lack of reciprocal monophyly between Amazon and Atlantic Rain Forest populations based on DNA sequence data, and unstructured amplified fragment length polymorphism variation. Our combined results allow us to clarify species-level limits within the genus Philaethria, whereby we propose that P. pygmalion is conspecific with P. wernickei (new synonym), and reassess the spatial range of P. wernickei by providing a refined mapping of its geographical distribution. Beyond clarifying the taxonomy of Philaethria, our results provide a solid, integrative framework that could be applied to fully characterize the taxonomy of other species in the Heliconiini and beyond.