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The aromobatid frog Allobates amissibilis sp. nov. is described from a very limited area in the Iwokrama Mountains at elevations between 160 and 950 m, in central Guyana, South America. The new taxon is diagnosed from other Allobates species by morphology, bioacoustics, and genetics, and can be distinguished readily from known congeners by cryptic colouration, small size (16.3–17.8 mm snout-vent length), absence of distinct dorsal pattern in living adult individuals, presence of a distinctly enlarged tubercle on each eyelid, diffuse broad oblique lateral stripe extending from groin to about midbody length, sexually dimorphic throat colouration (pinkish grey, suffused with dark pigment in male, immaculate cream to yellow in female), belly cream to yellow in both sexes, and distinct vocalisation formed by calls of usually 9–12 notes emitted between silent intervals of usually ca . 2–6 s, with a dominant frequency ranging from 5,064 to 5,845 Hz. The new species is the third microendemic reported from the Iwokrama Mountains together with the caecilian Caecilita iwokramae and the lizard Gonatodes timidus. We recommend considering the conservation status of the new species as Data Deficient, and highlight that environmental impact assessments are needed to refine its IUCN threat status. [ABSTRACT FROM AUTHOR]

Spiny-backed tree frogs of the genus Osteocephalus are conspicuous components of the tropical wet forests of the Amazon and the Guiana Shield. Here, we revise the phylogenetic relationships of Osteocephalus and its sister group Tepuihyla, using up to 6134 bp of DNA sequences of nine mitochondrial and one nuclear gene for 338 specimens from eight countries and 218 localities, representing 89% of the 28 currently recognized nominal species. Our phylogenetic analyses reveal (i) the paraphyly of Osteocephalus with respect to Tepuihyla, (ii) the placement of ' Hyla' warreni as sister to Tepuihyla, (iii) the non-monophyly of several currently recognized species within Osteocephalus and (iv) the presence of low (<1%) and overlapping genetic distances among phenotypically well-characterized nominal species (e.g. O. taurinus and O. oophagus) for the 16 S gene fragment used in amphibian DNA barcoding. We propose a new taxonomy, securing the monophyly of Osteocephalus and Tepuihyla by rearranging and redefining the content of both genera and also erect a new genus for the sister group of Osteocephalus. The colouration of newly metamorphosed individuals is proposed as a morphological synapomorphy for Osteocephalus. We recognize and define five monophyletic species groups within Osteocephalus, synonymize three species of Osteocephalus ( O. germani, O. phasmatus and O. vilmae) and three species of Tepuihyla ( T. celsae, T. galani and T. talbergae) and reallocate three species ( Hyla helenae to Osteocephalus, O. exophthalmus to Tepuihyla and O. pearsoni to Dryaderces gen. n.). Furthermore, we flag nine putative new species (an increase to 138% of the current diversity). We conclude that species numbers are largely underestimated, with most hidden diversity centred on widespread and polymorphic nominal species. The evolutionary origin of breeding strategies within Osteocephalus is discussed in the light of this new phylogenetic hypothesis, and a novel type of amplexus (gular amplexus) is described. [ABSTRACT FROM AUTHOR]

The article focuses on a research work conducted to assure the presence of the sphaerodactylid lizard, gonatodes vittatus in Guyana. It mentions that the rediscovery of gonatodes vittatus have been made according to the specimens collected in the urban area of Georgetown, Guyana. It also mentions that this lizard primarily inhabits seashores, thorn scrubland and forest edges. It reports on the specimens deposited in the herpetological collections of the Royal Belgian Institute of Natural Sciences, Brussels, Belgium. It also reports that several authors have assured the occurrence of this species in Guyana on the basis of various voucher specimens.

Iphisa elegans Gray, 1851 is a ground-dwelling lizard widespread over Amazonia that displays a broadly conserved external morphology over its range. This wide geographical distribution and conservation of body form contrasts with the expected poor dispersal ability of the species, the tumultuous past of Amazonia, and the previously documented prevalence of cryptic species in widespread terrestrial organisms in this region. Here we investigate this homogeneity by examining hemipenial morphology and conducting phylogenetic analyses of mitochondrial ( CYTB) and nuclear ( C-MOS) DNA sequence data from 49 individuals sampled across Amazonia. We detected remarkable variation in hemipenial morphology within this species, with multiple cases of sympatric occurrence of distinct hemipenial morphotypes. Phylogenetic analyses revealed highly divergent lineages corroborating the patterns suggested by the hemipenial morphotypes, including co-occurrence of different lineages. The degrees of genetic and morphological distinctness, as well as instances of sympatry among mtDNA lineages/morphotypes without nuDNA allele sharing, suggest that I. elegans is a complex of cryptic species. An extensive and integrative taxonomic revision of the I. elegans complex throughout its wide geographical range is needed. © 2012 The Linnean Society of London, Zoological Journal of the Linnean Society, 2012, 166, 361-376. [ABSTRACT FROM AUTHOR]

ABSTRACT Male frogs emit stereotypical advertisement calls to attract mates and deter conspecific rivals. The evolution of these calls is thought to be linked to anatomical constraints and the acoustic characteristics of their surroundings. The acoustic adaptation hypothesis (AAH) posits that species evolve calls that maximize propagation distance and reduce signal degradation in the environment where they are emitted. We applied phylogenetic comparative analyses to study the association of body size, vegetation density, type of aquatic ecosystem, and calling site on the evolution of acoustic traits in Cophomantini, a large radiation of Neotropical treefrogs (Hylidae). We obtained and analyzed body size, acoustic, and habitat data from a total of 112 species (58% of Cophomantini), using the most inclusive available phylogeny. We found a significant negative correlation between peak frequency, body size, and calling site, but contrary to the predictions of the AAH, we did not find support for associations among call traits and environmental characteristics. Although spectral allometry is explained by an anatomical constraint, it could also be maintained by female choice. We recommend that future studies strive to incorporate factors such as female mate preferences, eavesdropping by predators or parasites, and genetic drift. [ABSTRACT FROM AUTHOR]

Males of the advanced salamanders (Salamandroidea) attain internal fertilization without a copulatory organ by depositing a spermatophore on the substrate in the environment, which females subsequently take up with their cloaca. The aquatically reproducing modern Eurasian newts (Salamandridae) have taken this to extremes, because most species do not display close physical contact during courtship, but instead largely rely on females following the male track at spermatophore deposition. Although pheromones have been widely assumed to represent an important aspect of male courtship, molecules able to induce the female following behaviour that is the prelude for successful insemination have not yet been identified. Here, we show that uncleaved sodefrin precursor-like factor (SPF) protein pheromones are sufficient to elicit such behaviour in female palmate newts (Lissotriton helveticus). Combined transcriptomic and proteomic evidence shows that males simultaneously tail-fan multiple ca 20 kDa glycosylated SPF proteins during courtship. Notably, molecular dating estimates show that the diversification of these proteins already started in the late Palaeozoic, about 300 million years ago. Our study thus not only extends the use of uncleaved SPF proteins outside terrestrially reproducing plethodontid salamanders, but also reveals one of the oldest vertebrate pheromone systems. [ABSTRACT FROM AUTHOR]

Coralsnakes of the genus Micrurus are a diverse group of venomous snakes ranging from the southern United States to southern South America. Much uncertainty remains over the genus diversity, and understanding Micrurus systematics is of medical importance. In particular, the widespread Micrurus nigrocinctus spans from Mexico throughout Central America and into Colombia, with a number of described subspecies. This study provides new insights into the phylogenetic relationships within M. nigrocinctus by examining sequence data from a broad sampling of specimens from Mexico, Guatemala, Honduras, Nicaragua, Costa Rica, and Panama. The recovered phylogenetic relationships suggest that M. nigrocinctus is a species complex originating in the Pliocene and composed of at least three distinct species-level lineages. In addition, recovery of highly divergent clades supports the elevation of some currently recognized subspecies to the full species rank while others may require synonymization. [ABSTRACT FROM AUTHOR]

Many anuran species remain to be formally named and described in Amazonia, notably in the Guiana Shield, and particularly in megadiverse groups such as Pristimantis. Several species in the Guiana Shield region have been confused with Pristimantis marmoratus and P. ockendeni. Hylodes grandoculis, a taxon previously placed in the synonymy of P. marmoratus, may be available for one of these species. To disentangle this confusing situation, we examined the external morphology and osteology (via µ-CT scans) of the holotype of H. grandoculis, the holotype of Pristimantis marmoratus, and of recently collected material for which we also analyzed molecular, acoustic, and morphological variation. We concluded that some populations from Suriname and northern Pará, Brazil, are distinct from P. marmoratus and correspond to Pristimantis grandoculis. Other populations, from French Guiana, are closely related to P. grandoculis but their status remains ambiguous. Finally, some populations, from French Guiana and Amapá, Brazil, are conspicuously distinct from both P. marmoratus and P. grandoculis and are described herein as P. crepitaculus sp. nov. A third species, belonging to a "trans-amazon complex", occurs in southern Suriname, Guyana, and Brazil and remains undescribed. Pristimantis grandoculis and related populations from French Guiana lack external tympanum, columella, pharyngeal ostia, vocal slits and do not vocalize. This represents a rare, perhaps unique, example of a deaf and mute species of frogs from the Amazonian lowlands. [ABSTRACT FROM AUTHOR]

Major historical landscape changes have left significant signatures on species diversification. However, how these changes have affected the build-up and maintenance of Amazonia's megadiversity continues to be debated. Here, we addressed this issue by focusing on the evolutionary history of a pan-Amazonian toad genus that has diversified throughout the Neogene (Amazophrynella). Based on a comprehensive spatial and taxonomic sampling (286 samples, all nominal species), we delimited operational taxonomic units (OTUs) from mitochondrial DNA sequences. We delimited 35 OTUs, among which 13 correspond to nominal species, suggesting a vast underestimation of species richness. Next, we inferred time-calibrated phylogenetic relationships among OTUs based on complete mitogenomic data, which confirmed an ancient divergence between two major clades distributed in eastern and western Amazonia, respectively. Ancestral area reconstruction analyses suggest that the Andean foothills and the Brazilian Shield region represent the ancient core areas for their diversification. These two clades, probably isolated from one other by lacustrine ecosystems in western Amazonia during the Miocene, display a pattern of northward and eastward dispersals throughout the Miocene‒Pliocene. Given the ecological association of Amazophrynella with non-flooded forests, our results reinforce the perception that ancient Amazonian landscape changes had a major impact on the diversification of terrestrial vertebrates. [ABSTRACT FROM AUTHOR]

Aim: We investigate the spatiotemporal context of the diversification of Allobates, a widespread genus of Amazonian frogs with high species diversity particularly in western Amazonia. We tested if that diversity originated in situ or through repeated dispersals from other Amazonian areas and if this diversification took place during or after the Pebas system, a vast lacustrine system occupying most western Amazonia between 23 and 10 million years ago (Mya). Location: Amazonia. Taxon: Allobates (Anura: Aromobatidae). Methods: We gathered a spatially and taxonomically extensive sampling of mitochondrial DNA sequences from 932 Allobates specimens to delimit Operational Taxonomic Units (OTUs). Complete mitogenomes of these OTUs were assembled to reconstruct a time‐calibrated phylogeny used to infer the historical and spatial origin of the Amazonian Allobates lineages. Results: Phylogenetic analyses and ancestral area reconstruction suggested that most of the western Amazonian lineages resulted from in situ diversification and that these events occurred between the inferred origin of the genus (25 Mya) and 10 Mya, with a possible peak between 14 and 10 Mya. Dispersal among areas mostly occurred from western Amazonia towards the Brazilian and the Guiana Shields. Closely related Allobates OTUs display an allopatric pattern of distribution, matching interfluves delimited by modern Amazonian rivers. Main Conclusions: In western Amazonia, diversification of Allobates appears to have been simultaneous with the last stages of the Pebas system (14–10 Mya). Subsequently (within the last 10 Mya), modern Amazonian river courses shaped the distribution pattern of Allobates species and possibly promoted allopatric speciation. [ABSTRACT FROM AUTHOR]

Aim: Mapping Amazonian biodiversity accurately is a major challenge for integrated conservation strategies and to study its origins. However, species boundaries and their respective distribution are notoriously inaccurate in this region. Here, we generated a georeferenced database of short mtDNA sequences from Amazonian frogs, revised the species richness and the delimitation of bioregions of the Eastern Guiana Shield and estimated endemism within these bioregions. Location: Amazonia, with a focus on the Eastern Guiana Shield. Taxon studied: Amphibia: Anura. Methods: We used an extensive DNA‐based sampling of anuran amphibians of Amazonia using next‐generation sequencing to delimit Operational Taxonomic Units (OTU) and their distribution. We analysed this database to infer bioregions using Latent Dirichlet Allocation modelling. We then compared endemism within these bioregions based on our results and the current IUCN database, and inferred environmental variables that contributed the most to the biogeographic pattern. Results: The recognized anuran species richness within the focal area increased from 440 species currently listed by the IUCN Red List to as much as 876 OTUs with our dataset. We recovered eight bioregions, among which three lie within the Eastern Guiana Shield. We estimated that up to 82% of the OTUs found in this area are endemic, a figure three times higher than the previous estimate (28%). Environmental features related to seasonal precipitations are identified as playing an important role in shaping Amazonian amphibian bioregions. Main conclusions: Our results have major implications for defining future conservation priorities of this vast area given that endemism in most Amazonian bioregions is vastly underestimated, and might therefore hide a large portion of threatened species. Moreover, these findings raise concern about meta‐analyses based on public databases within Amazonia dealing with poorly known groups. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • A comprehensive phylogeny of Chironius (19 of 22 species) confirms its monophyly. • Genetic diversity suggests multiple species within some widespread species. • C. grandisquamis (Chocoan Rainforest) + C. challenger (Pantepui) are sister to all other species. • Current distribution patterns resulted from multiple range expansions. Abstract Neotropical sipo snakes (Chironius) are large diurnal snakes with a long tail and big eyes that differ from other Neotropical snakes in having 10 or 12 dorsal scale rows at midbody. The 22 currently recognized species occur from Central America south to Uruguay and northeastern Argentina. Based on the largest geographical sampling to date including ∼90% of all species, we analyzed one nuclear and three mitochondrial genes using phylogenetic methods to (1) test the monophyly of Chironius and some of its widely distributed species; (2) identify lineages that could represent undescribed species; and (3) reconstruct ancestral distributions. Our best hypothesis placed C. grandisquamis (Chocoan Rainforest) + C. challenger (Pantepui) as sister to all other species. Based on phylogeny and geographic distribution, we identified 14 subclades as putative species within Chironius fuscus , C. multiventris (including C. foveatus and C. laurenti), C. monticola , and C. exoletus. Under current taxonomy, these species show nearly twice as much genetic diversity as other species of Chironius for ND4. Biogeographical analyses using BioGeoBEARS suggest that current distribution patterns of Chironius species across South America resulted from multiple range expansions. The MRCA of the clade C. challenger + C. grandisquamis was most likely distributed over the Pantepui region, the Andes, and the Chocoan Rainforest, whereas the remaining lineages probably evolved from an Amazonian ancestor. [ABSTRACT FROM AUTHOR]

The Cercosaurini tribe stands out from other Gymnophthalmidae lizards for including several species with morphological adaptations to aquatic lifestyle (“Crocodile‐Like Morphology” – CLM). Recent molecular phylogenies of Cercosaurini demonstrated the paraphyly of CLM species, implicitly suggesting that adaptations to the aquatic life evolved more than once. However, CLM species have remained poorly sampled, and a number of uncertainties persist, such as the monophyly of Neusticurus and the placement of Potamites apodemus within the tribe. Based on a more extensive molecular and morphological data set, we propose a phylogenetic hypothesis for Neusticurus and investigate, for the first time, the phylogenetic position of P. apodemus. We recovered a monophyletic Neusticurus clade; however, Neusticurus rudis as currently understood was recovered as paraphyletic, with two lineages that also show consistent morphological diagnosis; as a result, we resurrect and provide a taxonomic redescription of Neusticurus surinamensis Müller, . Moreover, because P. apodemus was recovered as sister to all other Echinosaura, we propose a new combination for this species (Echinosaura apodema). We also review the distribution of Neusticurus species, offering a comprehensive view of their biogeography. Lastly, our ancestral character reconstruction and dating analyses indicate that the CLM phenotype evolved four times independently during Cercosaurini′s evolutionary history. We hypothesize that the CLM phenotype and subsequent adaptation to aquatic life may be linked to the development of the Pebas Lake in western Amazonia during the Miocene. [ABSTRACT FROM AUTHOR]

[Display omitted] • Molecular data suggest the existence of 15 species within Pipa. • Saturation of mtDNA causes overestimated divergence times compared to nuDNA. • Diversification started during the early Neogene probably in Amazonia. • Amazonian lineages have probably acquired endotrophic development independently. 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. [ABSTRACT FROM AUTHOR]

Abstract: 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 >5.8kb 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 7Ma minimum, while the northern Amazonian Adelophryne diversified much earlier, some 13Ma minimum. This striking biogeographic pattern coincides with major events that have shaped the face of the South American continent, as we know it today. [Copyright &y& Elsevier]

Background: Few animal populations have been studied under the framework of the OCBIL theory, which addresses the ecology and evolution of biodiversity on old climatically buffered infertile landscapes. Available genetic data challenge the low connectivity and high genetic differentiation predicted for isolated tepui-summit vertebrate communities, suggesting potential dispersal among summits. However, the OCBIL theory posits reduced dispersibility, enhanced resilience to habitat fragmentation and inbreeding due to small populations. We tested these hypotheses by conducting the first analytic evaluation of the spatial ecology and population biology of a tepui-summit vertebrate at multiple spatial scales., Results: We used harmonic radar tracking (100 individuals/448 points of contact) and capture-mark-recapture data (596 individuals captured/52 recaptured) to reveal the temporal niche, microhabitat use, population size, and dispersal abilities of the tepui-summit endemic toad Oreophrynella quelchii on Roraima-tepui. Abundance was determined using a closed population model incorporating sources of variation in capture probability. We tested the relative influence of biotic and abiotic variables on distances moved through model selection. Our data indicate that the population size of O. quelchii is remarkably large (ca. 12 million individuals), with strong seasonal demographic fluctuations. Ecology and observed limited spatial movements challenge the likelihood of active dispersal among tepui tops in this species. Our results are counter to those predicted by the available genetic data but support two hypotheses of the OCBIL theory: reduced dispersibility and enhanced resilience. However, they do not support the expectation of a small refugial population size., Conclusion: We postulate that the insular, hostile tepui-summit environment tends to produce robust demographic populations, likely to buffer stochastic adverse environmental effects, rather than diversity, as observed in much younger post-Pleistocene Neotropical landscapes. Our results draw attention to the value of faunal studies using an OCBIL framework to better understand the ecology and evolution of this unique biota worldwide., (© 2024. The Author(s).)

Tadpoles serve as crucial evidence for testing systematic and taxonomic hypotheses. Suctorial tadpoles collected in Guyana were initially assigned to Rhaebo nasicus through molecular phylogeny. Subsequent analysis of larval and adult morphological traits revealed synapomorphies within the clade encompassing R. nasicus and R. ceratophrys, prompting the recognition of a new genus described herein as Adhaerobufo. The new genus is distinguished from other bufonids by specific phenotypic traits including an enlarged, suctorial oral disc with distinct papillae arrangements, and the presence of certain muscles and narial vacuities at the larval stage. However, only a few adult external characteristics (e.g., enlarged eyelids, infraocular cream spot), seem to be reliably discriminative from related genera. This study underscores the significance of larval morphology in anuran systematics and offers new insights into the evolution of suctorial and gastromyzophorous larvae within bufonids., (© 2024. The Author(s).)

In addition to the type locality (the summit of Aprada-tepui, Bolívar State of Venezuela), the distribution of the egg-brooding frog Stefania satelles was long thought to include several isolated tabletop mountain (tepui) summits surrounding the large Chimantá Massif in Bolívar State (hence the Latin name "satelles"). However, multilocus molecular phylogenetic analyses have revealed that this taxon includes several undescribed morphologically cryptic species, and that S. satelles should be restricted to its type locality. Two tepui-summit species confused under that name in the literature remain to be named, and the present paper aims at describing these populations previously referred to as Stefania sp. 3 and S. sp. 5. Stefania sp. 3 is only known from the small summit of Angasima-tepui, while S. sp. 5 is only reported from the small summit of Upuigma-tepui, both mountains being located south of the Chimantá Massif. These new, phylogenetically distinct species are described based on external morphology and osteology and in comparison to close relatives in the S. ginesi clade, which consists exclusively of tepui summit species. Both new species have highly restricted geographic ranges (less than 3 km 2 ) and should be listed as Critically Endangered according to IUCN criteria., (© 2024. The Author(s).)

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., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

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., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)