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.

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.

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.

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.

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.

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.

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.

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.

Integrative taxonomy reveals a new but common Neotropical treefrog, hidden under the name Boana xerophylla.

Boana xerophylla is a common treefrog widely distributed in northern Brazil, Colombia, French Guiana, Guyana, Panama, Suriname, Trinidad and Tobago, and Venezuela. A recent study found molecular, acoustic, and morphometric differences between the populations located on opposite sides of the Orinoco River. Here, we carry out an updated molecular phylogenetic analysis, including new samples from all the countries along the distribution area, and analyzed additional call recordings from Brazil, Colombia, French Guiana, Panama, Trinidad and Tobago, and Venezuela. Our phylogenetic inference reveals three geographically restricted lineages: one in the eastern Guiana Shield (corresponding to B. xerophylla sensu stricto), another in the western Guiana Shield, and a third one north of the Orinoco River. Morphological and acoustic data agree with the differentiation between the populations north of the Orinoco River and the eastern Guiana Shield despite the low genetic p-distances observed (16S rRNA: 0.72.2 %). We argue that the populations north of the Orinoco River correspond to a new species, sister of B. xerophylla. We name and describe Boana platanera sp. nov. from the southern versant of the Cordillera de Mérida (08º48'26'' N, 70º30'46'' W, WGS 84; 947 m asl), Venezuela, and refer all the populations north of the Orinoco River currently identified as B. xerophylla to this species. The new species can be readily diagnosed from B. xerophylla (characters of the latter in parentheses) by a pale orange-yellow or light brown dorsal coloration (dark brown to green), palpebral membrane with dark pigments (pigments absent); pericloacal region dark brown (cream), advertisement call with shorter first note length than B. xerophylla. This study represents an empirical example regarding false negatives behind genetic thresholds for species discovery, appraising the use of integrative taxonomic approaches.

Exploring the vertebrate fauna of the Bird's Head Peninsula (Indonesia, West Papua) through DNA barcodes.

Biodiversity knowledge is widely heterogeneous across the Earth's biomes. Some areas, due to their remoteness and difficult access, present large taxonomic knowledge gaps. Mostly located in the tropics, these areas have frequently experienced a fast development of anthropogenic activities during the last decades and are therefore of high conservation concerns. The biodiversity hotspots of Southeast Asia exemplify the stakes faced by tropical countries. While the hotspots of Sundaland (Java, Sumatra, Borneo) and Wallacea (Sulawesi, Moluccas) have long attracted the attention of biologists and conservationists alike, extensive parts of the Sahul area, in particular the island of New Guinea, have been much less explored biologically. Here, we describe the results of a DNA-based inventory of aquatic and terrestrial vertebrate communities, which was the objective of a multidisciplinary expedition to the Bird's Head Peninsula (West Papua, Indonesia) conducted between 17 October and 20 November 2014. This expedition resulted in the assembly of 1005 vertebrate DNA barcodes. Based on the use of multiple species-delimitation methods (GMYC, PTP, RESL, ABGD), 264 molecular operational taxonomic units (MOTUs) were delineated, among which 75 were unidentified and an additional 48 were considered cryptic. This study suggests that the diversity of vertebrates of the Bird's Head is severely underestimated and considerations on the evolutionary origin and taxonomic knowledge of these biotas are discussed.

Two new species of Anomaloglossus (Anura: Aromobatidae) of the stepheni group from Suriname.

Anomaloglossus is a species-rich genus of frogs endemic to the Guiana Shield that still harbours several unnamed species. According to a recent integrative taxonomic survey, the A. stepheni species group includes five valid nominal species and at least four putatively unnamed species, two in Brazil and two in Suriname. In this paper, we describe the two species from Suriname based on adult and tadpole morphology as well as their calls and natural history. Both have exotrophic tadpoles transported by the male to small water bodies. These two new species differ from each other and from other congeners in body size, colouration pattern, call characteristics and breeding sites. Both have narrow distributions and should be considered Endangered according to IUCN criteria.

One step beyond a broad molecular phylogenetic analysis: Species delimitation of Adenomera marmorata Steindachner, 1867 (Anura: Leptodactylidae).

Taxonomists always have had intense discussions about how species should be delimited and recently many studies have used integrative approaches by combining molecular, morphological, and bioacoustic data. Although these studies are paramount for understanding species diversity, few of them actually formalize species delimitations to the final step of nomenclatural acts. Historically, the Neotropical frog genus Adenomera has been considered as a difficult taxonomic group because it comprises many morphologically similar species exhibiting high levels of intraspecific polymorphism. A recent work using molecular data shed light on the phylogenetic relationships within the genus and identified several lineages that may correspond to undescribed species but did not delimit species boundaries. In the Atlantic Forest, a clade formed by A. marmorata and two putative species (Adenomera sp. J and Adenomera sp. K) were identified. In this paper, we combine morphological, acoustic, and molecular data in order to evaluate species limits within this Atlantic Forest Adenomera clade. We provide a redescription of A. marmorata and restrict its type locality to the Tijuca Massif, in the Municipality of Rio de Janeiro, Brazil. Our results do not support A. marmorata and the two candidate species as diagnosable distinct species. Therefore A. marmorata corresponds to a species with pronounced morphological and acoustic variation in the genus and a complex phylogeographic structure.

The complete mitochondrial genome of Iphisa elegans (Reptilia: Squamata: Gymnophthalmidae).

The complete mitogenome of the lizard Iphisa elegans Gray, 1851 was sequenced using a shotgun approach on an Illumina HiSeq 3000 platform, providing the first mitogenome for Gymnophthalmidae. The genome was 18,622 bp long, with 13 protein-coding genes, two rRNA (12S and 16S), and 22 tRNA, as well as the control region. A maximum likelihood phylogenetic analysis including I. elegans and all other available mitogenomes of Squamata provided a tree in accordance with previous phylogenetic relationships inferred for Squamata.

A new species of Anomaloglossus (Anura: Aromobatidae) of the stepheni group with the redescription of A. baeobatrachus (Boistel and de Massary, 1999), and an amended definition of A. leopardus Ouboter and Jairam, 2012.

Anomaloglossus is a species-rich genus of frogs endemic to the Guiana Shield that still harbors several unnamed species. Within the A. stepheni species group (which includes four valid nominal species), A. baeobatrachus has an uncertain taxonomic status, notably because the holotype was an unvouchered specimen depicted in a popular journal. Another member of this group, A. leopardus, was only superficially described, lacking information on the sex of specimens in the type series and on advertisement call. Therefore, these two taxa need clarifications in order to allow the description of the extant undescribed species. In this paper, we redescribe A. baeobatrachus based on newly collected material from the species type locality and provide information about its reproductive ecology. We also provide an amended definition of A. leopardus using newly collected material from its type locality. These two species form a clade along with a third species from the Eastern Guiana Shield, which is also described herein. The reproductive biology of A. baeobatrachus and A. stepheni is very similar. Both species have endotrophic and nidicolous tadpoles, despite being distantly related, suggesting independent evolution of this breeding mode. The new species and A. leopardus, on the other hand, have exotrophic tadpoles.

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.

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.