Poison frogs (Dendrobatidae) are famous for their aposematic species, having a combination of diverse color patterns and defensive skin toxins, yet most species in this family are inconspicuous (cryptic) and considered non-aposematic.Epipedobatesis among the youngest genus-level clades of Dendrobatidae that includes both aposematic and cryptic species. Using mitochondrial and nuclear markers, we demonstrate deep genetic divergences among cryptic species ofEpipedobatesbut relatively shallow genetic divergences among conspicuous species. Our phylogenetic analysis includes broad geographic sampling of the cryptic lineages typically identified asE. boulengeriandE. espinosai, which reveals two putative new species, one in west-central Colombia (E.sp. 1) and the other in north-central Ecuador (E.aff.espinosai). We conclude thatE. darwinwallaceiis a junior subjective synonym ofE. espinosai. We also clarify the geographic distributions of crypticEpipedobatesspecies including the widespreadE. boulengeri. We provide a qualitative assessment of the phenotypic diversity in each nominal species, with a focus on the color and pattern of cryptic species. We conclude thatEpipedobatescontains eight known valid species, six of which are cryptic. A relaxed molecular clock analysis suggests that the most recent common ancestor ofEpipedobatesis ~11.1 million years old, which nearly doubles previous estimates. Last, genetic information points to a center of species diversity in the Chocó at the southwestern border of Colombia with Ecuador.

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During acoustic communication, an audible message is transmitted from a sender to a receiver, often producing changes in behavior. In a system where evolutionary changes of the sender do not result in a concomitant adjustment in the receiver, communication and species recognition could fail. However, the possibility of an evolutionary decoupling between sender and receiver has rarely been studied. Frog populations in the Allobates femoralis cryptic species complex are known for their extensive morphological, genetic and acoustic variation. We hypothesized that geographic variation in acoustic signals of A. femoralis was correlated with geographic changes in communication through changes in male-male recognition. To test this hypothesis, we quantified male call recognition using phonotactic responses to playback experiments of advertisement calls with two, three and four notes in eight localities of the Amazonian basin. Then, we reconstructed the ancestral states of call note number in a phylogenetic framework and evaluated whether the character state of the most recent common ancestor predicted current relative responses to two, three and four notes. The probability of a phonotactic response to advertisement calls of A. femoralis males was strongly influenced by the call mid-frequency and the number of notes in most populations. Positive phonotaxis was complete for calls from each individual's population, and in some populations, it was also partial for allotopic calls; however, in two populations, individuals equally recognized calls with two, three or four notes. This evidence, in conjunction with our results from phylogenetic comparative methods, supports the hypothesis of decoupled evolution between sender and receiver in the male-male communication system of the A. femoralis complex. Thus, signal recognition appears to evolve more slowly than the calls.

Restriction-site-associated DNA sequencing (RADseq) has become an accessible way to obtain genome-wide data in the form of single-nucleotide polymorphisms (SNPs) for phylogenetic inference. Nonetheless, how differences in RADseq methods influence phylogenetic estimation is poorly understood because most comparisons have largely relied on conceptual predictions rather than empirical tests. We examine how differences in ddRAD and 2bRAD data influence phylogenetic estimation in two nonmodel frog groups. We compare the impact of method choice on phylogenetic information, missing data, and allelic dropout, considering different sequencing depths.Given that researchers must balance input (funding, time) with output (amount and quality of data), we also provide comparisons of laboratory effort, computational time, monetary costs, and the repeatability of library preparation and sequencing. Both 2bRAD and ddRAD methods estimated well-supported trees, even at low sequencing depths, and had comparable amounts of missing data, patterns of allelic dropout, and phylogenetic signal. Compared to ddRAD, 2bRAD produced more repeatable datasets, had simpler laboratory protocols, and had an overall faster bioinformatics assembly. However, many fewer parsimony-informative sites per SNP were obtained from 2bRAD data when using native pipelines, highlighting a need for further investigation into the effects of each pipeline on resulting datasets. Our study underscores the importance of comparing RADseq methods, such as expected results and theoretical performance using empirical datasets, before undertaking costly experiments.

Territoriality and parental care are complex reproductive behaviors found in many taxa from insects to mammals. Parental care can be carried out by the female, the male, or both, depending on the species. Territoriality, in contrast, is predominantly displayed by males. Different selective pressures imposed on individuals from the sex performing territorial or parental care behaviors may also lead to sexual differentiation in other life-history traits. Due to their territorial behavior and their diversity of parental care behaviors, Neotropical poison frogs are an excellent study system to investigate whether behavioral traits can influence sexual differentiation in intrinsic or extrinsic traits of individuals. Here, we evaluate whether territorial and parental care behaviors mediate sexual differentiation in ecological (habitat use) and phenotypic (coloration, morphology) traits in the critically endangered Lehmann’s poison frog (Oophaga lehmanni), a species in which males defend territories while females provide parental care. We found sex differences in habitat use and morphological traits, but not in coloration. Males use trunks and green leaves as perches more frequently and are found on higher substrates, than females. We found no sex differences in body size, but females have longer arms than males, which is probably associated with their parental duties (climbing trees to feed the tadpoles). Altogether, our results provide evidence that selection pressures act differently on male and female traits, and that territoriality and parental care may promote the evolution of sexual differentiation in dendrobatids. Long-term wildlife observations are essential to identify important life-history traits and to evaluate hypotheses about the behavioral ecology and conservation of this and other vertebrate species.