Systematics of the Osteocephalus buckleyi species complex (Anura, Hylidae) from Ecuador and Peru

We present a new phylogeny, based on DNA sequences of mitochondrial and nuclear genes, for frogs of the genus Osteocephalus with emphasis in the Osteocephalus buckleyi species complex. Genetic, morphologic, and advertisement call data are combined to define species boundaries and describe new species. The phylogeny shows strong support for: (1) a basal position of Osteocephalus taurinus + Osteocephalus oophagus, (2) a clade containing phytotelmata breeding species, and (3) a clade that corresponds to the Osteocephalus buckleyi species complex. Our results document a large proportion of hidden diversity within a set of populations that were previously treated as a single, widely distributed species, Osteocephalus buckleyi. Individuals assignable to Osteocephalus buckleyi formed a paraphyletic group relative to Osteocephalus verruciger and Osteocephalus cabrerai and contained four species, one of which is Osteocephalus buckleyisensu stricto and three are new. Two of the new species are shared between Ecuador and Peru (Osteocephalus vilmaesp. n. and Osteocephalus cannatellaisp. n.) and one is distributed in the Amazon region of southern Peru (Osteocephalus germanisp. n.) We discuss the difficulties of using morphological characters to define species boundaries and propose a hypothesis to explain them.