Ongoing declines for the world's amphibians in the face of emerging threats.

Systematic assessments of species extinction risk at regular intervals are necessary for informing conservation action1,2. Ongoing developments in taxonomy, threatening processes and research further underscore the need for reassessment3,4. Here we report the findings of the second Global Amphibian Assessment, evaluating 8,011 species for the International Union for Conservation of Nature Red List of Threatened Species. We find that amphibians are the most threatened vertebrate class (40.7% of species are globally threatened). The updated Red List Index shows that the status of amphibians is deteriorating globally, particularly for salamanders and in the Neotropics. Disease and habitat loss drove 91% of status deteriorations between 1980 and 2004. Ongoing and projected climate change effects are now of increasing concern, driving 39% of status deteriorations since 2004, followed by habitat loss (37%). Although signs of species recoveries incentivize immediate conservation action, scaled-up investment is urgently needed to reverse the current trends.

Relicts in the mist: Two new frog families, genera and species highlight the role of Pantepui as a biodiversity museum throughout the Cenozoic.

The iconic mountains of the Pantepui biogeographical region host many early-diverging endemic animal and plant lineages, concurring with Conan Doyle's novel about an ancient "Lost World". While this is the case of several frog lineages, others appear to have more recent origins, adding to the controversy around the diversification processes in this region. Due to its remoteness, Pantepui is challenging for biological surveys, and only a glimpse of its biodiversity has been described, which hampers comprehensive evolutionary studies in many groups. During a recent expedition to the Neblina massif on the Brazil-Venezuela border, we sampled two new frog species that could not be assigned to any known genus. Here, we perform phylogenetic analyses of mitogenomic and nuclear loci to infer the evolutionary relationships of the new taxa and support their description. We find that both species represent single lineages deeply nested within Brachycephaloidea, a major Neotropical clade of direct-developing frogs. Both species diverged >45 Ma from their closest relatives: the first is sister to all other Brachycephaloidea except for Ceuthomantis, another Pantepui endemic, and the second is sister to Brachycephalidae, endemic to the Brazilian Atlantic Forest. In addition to these considerable phylogenetic and biogeographic divergences, external morphology and osteological features support the proposition of two new family and genus-level taxa to accommodate these new branches of the amphibian tree of life. These findings add to other recently described ancient vertebrate lineages from the Neblina massif, providing a bewildering reminder that our perception of the Pantepui's biodiversity remains vastly incomplete. It also provides insights into how these mountains acted as "museums" during the diversification of Brachycephaloidea and of Neotropical biotas more broadly, in line with the influential "Plateau theory". Finally, these discoveries point at the yet unknown branches of the tree of life that may go extinct, due to global climate change and zoonotic diseases, before we even learn about their existence, amphibians living at higher elevations being particularly at risk.

Diversification of the terrestrial frog genus Anomaloglossus (Anura, Aromobatidae) in the Guiana Shield proceeded from highlands to lowlands, with successive loss and reacquisition of endotrophy.

Two main landscapes emerge from the Guiana Shield: the highlands to the west called the Pantepui region and the Amazonian lowlands to the east, both harbouring numerous endemic species. With 32 currently recognized species, the genus Anomaloglossus stands out among Neotropical frogs as one that diversified only within the Guiana Shield both in the highlands and the lowlands. We present a time-calibrated phylogeny obtained by using combined mitogenomic and nuclear DNA, which suggests that the genus originates from Pantepui where extant lineages started diversifying around 21 Ma, and subsequently (ca. 17 Ma) dispersed during the Miocene Climatic Optimum to the lowlands of the eastern Guiana Shield where the ability to produce endotrophic tadpoles evolved. Further diversification within the lowlands in the A. stepheni group notably led to an evolutionary reversal toward exotrophy in one species group during the late Miocene, followed by reacquisition of endotrophy during the Pleistocene. These successive shifts of reproductive mode seem to have accompanied climatic oscillations. Long dry periods might have triggered evolution of exotrophy, whereas wetter climates favoured endotrophic forms, enabling colonization of terrestrial habitats distant from water. Acquisition, loss, and reacquisition of endotrophy makes Anomaloglossus unique among frogs and may largely explain the current species diversity. The micro evolutionary processes involved in these rapid shifts of reproductive mode remain to be revealed.

The small and inconspicuous majority: Revealing the megadiversity and historical biogeography of the Pristimantis unistrigatus species group (Anura, Strabomantidae).

With more than 600 recognized species, the genus Pristimantis is already the most diverse among vertebrates, but described species only represent a fraction of the actual diversity in this clade. This genus is widely distributed throughout the Neotropics and represents an interesting model for biogeographic studies because Pristimantis spp. are direct developing and generally have narrow ecological niches and low dispersal abilities. The P. unistrigatus species group is one of the most important components in the genus (ca. 200 recognized species) and has been supported by morphological but not by molecular evidence. We assessed the species boundaries and distribution in the P. unistrigatus species group and infer spatiotemporal patterns of diversification related to historical landscape changes in the Neotropics. We gathered three mitochondrial, and two nuclear DNA loci from 416 specimens throughout the range of the group, and including 68 nominal species. We redefine the group based on the obtained phylogeny and found 151 candidate species that composes it, with 83 of these remaining undescribed. We recovered 11 major clades within the group that diverged before 13 Ma. The diversification of the group started during the early Miocene most likely in northwestern South America, currently corresponding to western Amazonia and northern Andes. The other neotropical areas subsequently acted as sinks, receiving lineages mostly during the last 10 Ma, after the demise of the Pebas System and the setup of the modern Amazonian hydrographic system.

Linking Predator Responses to Alkaloid Variability in Poison Frogs.

Many chemically-defended/aposematic species rely on diet for sequestering the toxins with which they defend themselves. This dietary acquisition can lead to variable chemical defenses across space, as the community composition of chemical sources is likely to vary across the range of (an aposematic) species. We characterized the alkaloid content of two populations of the Dyeing Poison Frog (Dendrobates tinctorius) in northeastern French Guiana. Additionally, we conducted unpalatability experiments with naive predators, Blue Tits (Cyanistes caeruleus), using whole-skin secretion cocktails to assess how a model predator would respond to the defense of individuals from each population. While there was some overlap between the two D. tinctorius populations in terms of alkaloid content, our analysis revealed that these two populations are markedly distinct in terms of overall alkaloid profiles. Predator responses to skin secretions differed between the populations. We identified 15 candidate alkaloids (including three previously undescribed) in seven classes that are correlated with predator response in one frog population. We describe alkaloid profile differences between populations for D. tinctorius and provide a novel method for assessing unpalatability of skin secretions and identifying which toxins may contribute to the predator response. In one population, our results suggest 15 alkaloids that are implicated in predator aversive response. This method is the first step in identifying the causal link between alkaloids and behavioral responses of predators, and thus makes sense of how varying alkaloid combinations are capable of eliciting consistent behavioral responses, and eventually driving evolutionary change in aposematic characters (or characteristics).

Diversity, biogeography, and reproductive evolution in the genus Pipa (Amphibia: Anura: Pipidae).

The genus Pipa is a species-poor clade of Neotropical frogs and one of the most bizarre-looking due to many highly derived anatomical traits related to their fully aquatic lifestyle. With their African relatives, they form the Pipidae family, which has attracted much attention, especially regarding its anatomy, reproductive biology, paleontology and biogeography. However, the actual diversity and phylogenetic relationships within Pipa remain poorly understood, and thus so do their historical biogeography and the evolution of striking features, such as the absence of teeth and endotrophy in some species. Using short mtDNA sequences across the distribution of the genus, we identified 15 main lineages (Operational Taxonomic Units - OTUs). This more than doubles the number of the currently seven valid nominal species. Several closely related OTUs do not share nuDNA alleles, confirming species divergence. Time-calibrated phylogenies obtained from mitogenomes and from 10 nuclear loci provide highly similar topologies but strikingly distinct node ages for Pipa. High dN/dS ratios and the variation of substitution rates across the trees suggest a strong effect of saturation on fast evolving positions of mtDNA, producing a substantially shorter stem branch of Pipa. Focusing on the nuDNA topology, we inferred an early Neogene Amazonian origin of the diversification of Pipa, with an initial split between the Guiana-Brazilian Shields and Western Amazonia, a pattern observed in many other co-distributed groups. All the western species are edentate, suggesting a single loss in the genus. Each of these groups diversified further out of Amazonia, toward the Atlantic Forest and toward trans-Andean forests, respectively. These events are concomitant with paleogeographic changes and match patterns observed in other co-distributed taxonomic groups. The two Amazonian lineages have probably independently acquired endotrophic larval development.

Weak warning signals can persist in the absence of gene flow.

Aposematic organisms couple conspicuous warning signals with a secondary defense to deter predators from attacking. Novel signals of aposematic prey are expected to be selected against due to positive frequency-dependent selection. How, then, can novel phenotypes persist after they arise, and why do so many aposematic species exhibit intrapopulation signal variability? Using a polytypic poison frog (Dendrobates tinctorius), we explored the forces of selection on variable aposematic signals using 2 phenotypically distinct (white, yellow) populations. Contrary to expectations, local phenotype was not always better protected compared to novel phenotypes in either population; in the white population, the novel phenotype evoked greater avoidance in natural predators. Despite having a lower quantity of alkaloids, the skin extracts from yellow frogs provoked higher aversive reactions by birds than white frogs in the laboratory, although both populations differed from controls. Similarly, predators learned to avoid the yellow signal faster than the white signal, and generalized their learned avoidance of yellow but not white. We propose that signals that are easily learned and broadly generalized can protect rare, novel signals, and weak warning signals (i.e., signals with poor efficacy and/or poor defense) can persist when gene flow among populations, as in this case, is limited. This provides a mechanism for the persistence of intrapopulation aposematic variation, a likely precursor to polytypism and driver of speciation.

When the tail shakes the snake: phylogenetic affinities and morphology of Atractus badius (Serpentes: Dipsadidae), reveals some current pitfalls on the snake's genomic age.

Atractus badius has a long and controversial nomenclatural history due to both its antiquity and the brevity of its original description. This species was described based on two syntypes from Java. Later, a lectotype was designated and the distribution range restricted to the Guiana Shield. Although this species has been repeatedly recorded throughout Amazonia and the Andes, these records have been erroneously assigned to A. badius because of a considerable level of confusion in the literature. We found 13 additional specimens of this poorly known snake, expanding our knowledge on its morphological variability, phylogenetic relationships and distribution. In this paper, we report new localities and data on meristic, morphometric, coloration, scales micro-ornamentation, osteology and hemipenial morphology to A. badius. We also determine the phylogenetic position of A. badius as nested in a composite Guiana Shield clade. In addition, we discuss recent advances on the systematics of Atractus and comment on some 'new arrangements' with respect to previously recognized species groups and available morphological evidence. Finally, we highlight the importance of accurate voucher identification before promoting taxonomic changes or implementing nomenclatural acts derived from new phylogenetic hypotheses, and notably propose the synonymy of the recently described A. pyroni with A. roulei.

Phenotypic and life-history diversification in Amazonian frogs despite past introgressions.

The advent of genomics in phylogenetics and population genetics strengthened the perception that conflicts among gene trees are frequent and often due to introgression. However, hybridization occurs mostly among species that exhibit little phenotypic differentiation. A recent study delineating species in Anomaloglossus, a frog genus endemic to the Guiana Shield, identified an intriguing pattern in the A. baeobatrachus species complex. This complex occurs in French Guiana and Amapá (Brazil) and comprises two sympatric phenotypes contrasting not only in body size, habitat, and advertisement call, but also in larval development mode (endotrophic vs exotrophic tadpoles). However, molecular and phenotypic divergences are, in some cases, incongruent, i.e specimens sharing mtDNA haplotypes are phenotypically distinct, suggesting a complex evolutionary history. Therefore, we genotyped 106 Anomaloglossus individuals using ddRADseq to test whether this phenotype/genotype incongruence was a product of phenotypic plasticity, incomplete lineage sorting, multiple speciation events, or admixture. Based on more than 16,000 SNPs, phylogenetic and population genetic approaches demonstrated that exotrophic populations are paraphyletic. Species tree and admixture analyses revealed a strikingly reticulate pattern, suggesting multiple historical introgression events. The evolutionary history of one exotrophic population in northern French Guiana is particularly compelling given that it received genetic material from exotrophic ancestors but shows very strong genetic affinity with the nearby endotrophic populations. This suggests strong selection on larval development and mating call after secondary contact and hybridization. The case of A. baeobatrachus represents a striking example of introgression among lineages that are phenotypically distinct, even in their larval development mode, and highlights how high-resolution genomic data can unravel unexpectedly complex evolutionary scenarios.

Explosive breeding in tropical anurans: environmental triggers, community composition and acoustic structure.

BACKGROUND: Anurans largely rely on acoustic communication for sexual selection and reproduction. While multiple studies have focused on the calling activity patterns of prolonged breeding assemblages, species that concentrate their reproduction in short-time windows, explosive breeders, are still largely unknown, probably because of their ephemeral nature. In tropical regions, multiple species of explosive breeders may simultaneously aggregate leading to massive, mixed and dynamic choruses. To understand the environmental triggers, the phenology and composition of these choruses, we collected acoustic and environmental data at five ponds in French Guiana during a rainy season, assessing acoustic communities before and during explosive breeding events. RESULTS: We detected in each pond two explosive breeding events, lasting between 24 and 70 h. The rainfall during the previous 48 h was the most important factor predicting the emergence of these events. During explosive breeding events, we identified a temporal factor that clearly distinguished pre- and mid-explosive communities. A common pool of explosive breeders co-occurred in most of the events, namely Chiasmocleis shudikarensis, Trachycephalus coriaceus and Ceratophrys cornuta. Nevertheless, the species composition was remarkably variable between ponds and for each pond between the first and the second events. The acoustic structure of explosive breeding communities had outlying levels of amplitude and unexpected low acoustic diversity, significantly lower than the communities preceding explosive breeding events. CONCLUSIONS: Explosive breeding communities were tightly linked with specific rainfall patterns. With climate change increasing rainfall variability in tropical regions, such communities may experience significant shifts in their timing, distribution and composition. In structurally similar habitats, located in the same region without obvious barriers, our results highlight the variation in composition across explosive breeding events. The characteristic acoustic structure of explosive breeding events stands out from the circadian acoustic environment being easily detected at long distance, probably reflecting behavioural singularities and conveying heterospecific information announcing the availability of short-lived breeding sites. Our data provides a baseline against which future changes, possibly linked to climate change, can be measured, contributing to a better understanding on the causes, patterns and consequences of these unique assemblages.

Cryptic diversity in Amazonian frogs: Integrative taxonomy of the genus Anomaloglossus (Amphibia: Anura: Aromobatidae) reveals a unique case of diversification within the Guiana Shield.

Lack of resolution on species boundaries and distribution can hamper inferences in many fields of biology, notably biogeography and conservation biology. This is particularly true in megadiverse and under-surveyed regions such as Amazonia, where species richness remains vastly underestimated. Integrative approaches using a combination of phenotypic and molecular evidence have proved extremely successful in reducing knowledge gaps in species boundaries, especially in animal groups displaying high levels of cryptic diversity like amphibians. Here we combine molecular data (mitochondrial 16S rRNA and nuclear TYR, POMC, and RAG1) from 522 specimens of Anomaloglossus, a frog genus endemic to the Guiana Shield, including 16 of the 26 nominal species, with morphometrics, bioacoustics, tadpole development mode, and habitat use to evaluate species delineation in two lowlands species groups. Molecular data reveal the existence of 18 major mtDNA lineages among which only six correspond to described species. Combined with other lines of evidence, we confirm the existence of at least 12 Anomaloglossus species in the Guiana Shield lowlands. Anomaloglossus appears to be the only amphibian genus to have largely diversified within the eastern part of the Guiana Shield. Our results also reveal strikingly different phenotypic evolution among lineages. Within the A. degranvillei group, one subclade displays acoustic and morphological conservatism, while the second subclade displays less molecular divergence but clear phenotypic divergence. In the A. stepheni species group, a complex evolutionary diversification in tadpole development is observed, notably with two closely related lineages each displaying exotrophic and endotrophic tadpoles.

Unexpected phylogenetic positions of the genera Rupirana and Crossodactylodes reveal insights into the biogeography and reproductive evolution of leptodactylid frogs.

Despite major progress in deciphering the amphibian tree of life by molecular phylogenetics, we identified two questions remaining to be answered regarding relationships within Hyloidea, the clade of South American origin that comprises most extant anuran diversity. A few genera like Rupirana and Crossodactylodes have enigmatic phylogenetic positions, and relationships among major lineages within some families like Leptodactylidae remain ambiguous. To resolve these specific questions we used two approaches (1) a complete matrix approach representing >6.6 kb, including most major Hyloidea lineages (61 terminals) combining different methods of phylogenetic reconstruction and measures of node support; and (2) a supermatrix approach >11.6 kb with a focus on Leptodactylidae. Both Rupirana and Crossodactylodes are unambiguously grouped with Paratelmatobius and Scythrophrys. The clade comprising these four genera is named Crossodactylodinae and embedded within Leptodactylidae. Crossodactylodinae is moderately supported as sister group of Leptodactylinae from (1) and as the sister group of the other Leptodactylidae from (2) with low support. Genera within Crossodactylodinae are scattered along a north-south axis in the Atlantic forest and their origins are very ancient (Paleocene). Such results stress the importance of the northern Atlantic forest in terms of conservation. Moreover, the position of Pseudopaludicola, which is well supported as the sister group to all other Leiuperinae, suggests that foam-nest building may have arisen independently in Leptodactylinae and Leiuperinae. Moreover, in spite of being of similar age, foam-nest builders are more widespread than nonfoam-nest breeders and have higher species diversity. Nevertheless, the bulk of the diversity within foam-nest breeders arose some 20 Myr later than the character itself.

Multiple quaternary refugia in the eastern Guiana shield revealed by comparative phylogeography of 12 frog species.

The Guiana Shield (GS) is one of the most pristine regions of Amazonia and biologically one of the richest areas on Earth. How and when this massive diversity arose remains the subject of considerable debate. The prevailing hypothesis of Quaternary glacial refugia suggests that a part of the eastern GS, among other areas in Amazonia, served as stable forested refugia during periods of aridity. However, the recently proposed disturbance-vicariance hypothesis proposes that fluctuations in temperature on orbital timescales, with some associated aridity, have driven Neotropical diversification. The expectations of the temporal and spatial organization of biodiversity differ between these two hypotheses. Here, we compare the genetic structure of 12 leaf-litter inhabiting frog species from the GS lowlands using a combination of mitochondrial and nuclear sequences in an integrative analytical approach that includes phylogenetic reconstructions, molecular dating, and Geographic Information System methods. This comparative and integrated approach overcomes the well-known limitations of phylogeographic inference based on single species and single loci. All of the focal species exhibit distinct phylogeographic patterns highlighting taxon-specific historical distributions, ecological tolerances to climatic disturbance, and dispersal abilities. Nevertheless, all but one species exhibit a history of fragmentation/isolation within the eastern GS during the Quaternary with spatial and temporal concordance among species. The signature of isolation in northern French Guiana (FG) during the early Pleistocene is particularly clear. Approximate Bayesian Computation supports the synchrony of the divergence between northern FG and other GS lineages. Substructure observed throughout the GS suggests further Quaternary fragmentation and a role for rivers. Our findings support fragmentation of moist tropical forest in the eastern GS during this period when the refuge hypothesis would have the region serving as a contiguous wet-forest refuge.

A new species of the genus Pristimantis (Amphibia, Craugastoridae) associated with the moderately elevated massifs of French Guiana.

We describe a new Pristimantis from French Guiana, northern South America, which is mainly distinguished from known phenotypically related congeners (i.e. species from the polyphyletic unistrigatus species group) occurring at low and middle elevations in the Guiana Shield by the combination of a distinct tympanum, a lower ratio of tibia vs. hand length, a reddish groin region, and a distinct advertisement call consisting of clusters of generally four short notes. The new species inhabits pristine primary forests on the slopes of isolated massifs reaching more than 400 m elevation, and seems not to occur below ca. 200 m above sea level. Such a sharp altitudinal limit suggests a strong influence of thermal variation on the distribution of the species, and therefore a potential sensitivity to climate change. With only nine isolated populations documented so far, the new species should be prioritized for conservation. Historical climate fluctuations during the Quaternary are likely responsible for the distribution pattern of the new species. 

Two new species of Allobates of the trilineatus clade (Anura: Aromobatidae) from the Eastern Guiana Shield.

The actual diversity of anurans occurring in Amazonia remains only partly described. This is notably the case of the species of the genus Allobates despite a fast rate of species description in this genus during the last decade. This increasing taxonomic effort has been particularly facilitated by the availability of DNA sequences throughout the region. These data suggest for instance the existence of three lineages in the Guiana Shield that may correspond to unnamed species. We examined morphology and calls of these putative species and compared them to related nominal species, which confirmed that two of these lineages not only differ genetically but also morphologically and bioacoustically. These two species are thus named and described herein. Allobates vicinus sp. nov. from northern Suriname forms a clade with another candidate species from southern Suriname and Northern Pará, (Brazil) and with A. granti that occurs in French Guiana. Allobates ripicolus sp. nov. occurs throughout the eastern Guiana Shield and is nested within the A. tapajos complex which otherwise occurs in southeastern Amazonia and comprises A. tapajos and A. kamilae, along with three additional candidate species.

A new species of Pristimantis (Anura: Strabomantidae) from white-sand forests of central Amazonia, Brazil.

The white-sand ecosystems in the Solimões-Negro Interfluve are among the less studied in Amazonia. Recent herpetological surveys conducted west of Manaus, Brazil (central Amazonia) indicate that white-sand forests host a unique anuran fauna comprising habitat specialized and endemic species. In the present study we describe a new species of rain frog belonging to the Pristimantis unistrigatus species group from the white-sand forest locally called "campinarana" (thin-trunked forests with canopy height below 20 m). The new species is phylogenetically close to rain frogs from western Amazonian lowlands (P. delius, P. librarius, P. matidiktyo and P. ockendeni). It differs from its closest relatives mainly by its size (male SVL of 17.3-20.1 mm, n = 16; female SVL of 23.2-26.5 mm, n = 6), presence of tympanum, tarsal tubercles and dentigerous processes of vomers, its translucent groin without bright colored blotches or marks, and by its advertisement call (composed of 5-10 notes, call duration of 550-1,061 ms, dominant frequency of 3,295-3,919 Hz). Like other anuran species recently discovered in the white-sand forests west of Manaus, the new species seems to be restricted to this peculiar ecosystem.

Molecular phylogeny and morphometric analyses reveal deep divergence between Amazonia and Atlantic Forest species of Dendrophryniscus.

Dendrophryniscus is an early diverging clade of bufonids represented by few small-bodied species distributed in Amazonia and the Atlantic Forest. We used mitochondrial (414 bp of 12S, 575 bp of 16S genes) and nuclear DNA (785 bp of RAG-1) to investigate phylogenetic relationships and the timing of diversification within the genus. These molecular data were gathered from 23 specimens from 19 populations, including eight out of the 10 nominal species of the genus as well as Rhinella boulengeri. Analyses also included sequences of representatives of 18 other bufonid genera that were publically available. We also examined morphological characters to analyze differences within Dendrophryniscus. We found deep genetic divergence between an Amazonian and an Atlantic Forest clade, dating back to Eocene. Morphological data corroborate this distinction. We thus propose to assign the Amazonian species to a new genus, Amazonella. The species currently named R. boulengeri, which has been previously assigned to the genus Rhamphophryne, is shown to be closely related to Dendrophryniscus species. Our findings illustrate cryptic trends in bufonid morphological evolution, and point to a deep history of persistence and diversification within the Amazonian and Atlantic rainforests. We discuss our results in light of available paleoecological data and the biogeographic patterns observed in other similarly distributed groups.

From Amazonia to the Atlantic forest: molecular phylogeny of Phyzelaphryninae frogs reveals unexpected diversity and a striking biogeographic pattern emphasizing conservation challenges.

Documenting the Neotropical amphibian diversity has become a major challenge facing the threat of global climate change and the pace of environmental alteration. Recent molecular phylogenetic studies have revealed that the actual number of species in South American tropical forests is largely underestimated, but also that many lineages are millions of years old. The genera Phyzelaphryne (1 sp.) and Adelophryne (6 spp.), which compose the subfamily Phyzelaphryninae, include poorly documented, secretive, and minute frogs with an unusual distribution pattern that encompasses the biotic disjunction between Amazonia and the Atlantic forest. We generated >5.8 kb sequence data from six markers for all seven nominal species of the subfamily as well as for newly discovered populations in order to (1) test the monophyly of Phyzelaphryninae, Adelophryne and Phyzelaphryne, (2) estimate species diversity within the subfamily, and (3) investigate their historical biogeography and diversification. Phylogenetic reconstruction confirmed the monophyly of each group and revealed deep subdivisions within Adelophryne and Phyzelaphryne, with three major clades in Adelophryne located in northern Amazonia, northern Atlantic forest and southern Atlantic forest. Our results suggest that the actual number of species in Phyzelaphryninae is, at least, twice the currently recognized species diversity, with almost every geographically isolated population representing an anciently divergent candidate species. Such results highlight the challenges for conservation, especially in the northern Atlantic forest where it is still degraded at a fast pace. Molecular dating revealed that Phyzelaphryninae originated in Amazonia and dispersed during early Miocene to the Atlantic forest. The two Atlantic forest clades of Adelophryne started to diversify some 7 Ma minimum, while the northern Amazonian Adelophryne diversified much earlier, some 13 Ma minimum. This striking biogeographic pattern coincides with major events that have shaped the face of the South American continent, as we know it today.

A new Chiasmocleis (Anura: Microhylidae) from the eastern Guiana Shield with an amended definition of C. haddadi Peloso, Sturaro, Forlani, Gaucher, Motta, & Wheeler, 2014.

The existence of an unnamed (candidate) species from French Guiana and the state of Amapá (Brazil) closely related to Chiasmocleis haddadi has been hypothesized in previous DNA-based studies. With an integrative use of genetic, morphological and acoustic data we confirm that these populations are indeed distinct from C. haddadi and all other known congeners. We herein describe and name this new species, and amend the definition of C. haddadi since the type-series and its original diagnosis included specimens of the new species. Geographically, the new species occurs eastward of its sister species C. haddadi from which it can be distinguished by having a larger body, a proportionally smaller eye and a distinct dorsal colouration. We suspect that the new species breeds in the leaf litter and that tadpoles undergo endotrophic development whereas co-occurring C. haddadi and C. hudsoni lay clutches in the leaf litter nearby standing water of temporary ponds where exotrophic tadpoles complete their development. This new species is a new addition to the already long list of animals being endemic to the easternmost part of the Guiana Shield, a region identified as a singular bioregion within Amazonia by previous research.