BackgroundAdvancements in portable scientific instruments provide promising avenues to expedite field work in order to understand the diverse array of organisms that inhabit our planet. Here, we tested the feasibility for in situ molecular analyses of endemic fauna using a portable laboratory fitting within a single backpack in one of the world's most imperiled biodiversity hotspots, the Ecuadorian Chocó rainforest. We used portable equipment, including the MinION nanopore sequencer (Oxford Nanopore Technologies) and the miniPCR (miniPCR), to perform DNA extraction, polymerase chain reaction amplification, and real-time DNA barcoding of reptile specimens in the field.FindingsWe demonstrate that nanopore sequencing can be implemented in a remote tropical forest to quickly and accurately identify species using DNA barcoding, as we generated consensus sequences for species resolution with an accuracy of >99% in less than 24 hours after collecting specimens. The flexibility of our mobile laboratory further allowed us to generate sequence information at the Universidad Tecnológica Indoamérica in Quito for rare, endangered, and undescribed species. This includes the recently rediscovered Jambato toad, which was thought to be extinct for 28 years. Sequences generated on the MinION required as few as 30 reads to achieve high accuracy relative to Sanger sequencing, and with further multiplexing of samples, nanopore sequencing can become a cost-effective approach for rapid and portable DNA barcoding.ConclusionsOverall, we establish how mobile laboratories and nanopore sequencing can help to accelerate species identification in remote areas to aid in conservation efforts and be applied to research facilities in developing countries. This opens up possibilities for biodiversity studies by promoting local research capacity building, teaching nonspecialists and students about the environment, tackling wildlife crime, and promoting conservation via research-focused ecotourism.

Comparative phylogeography allow us to understand how shared historical circumstances have shaped the formation of lineages, by examining a broad spectrum of co-distributed populations of different taxa. However, these types of studies are scarce in the Neotropics, a region that is characterized by high diversity, complex geology, and poorly understood biogeography. Here, we investigate the diversification patterns of five lineages of amphibians and reptiles, co-distributed across the Choco and Andes ecoregions in northwestern Ecuador. Mitochondrial DNA and occurrence records were used to determine the degree of geographic genetic divergence within species. Our results highlight congruent patterns of parapatric speciation and common geographical barriers for distantly related taxa. These comparisons indicate similar biological and demographic characteristics for the included clades, and reveal the existence of two new species of Pristimantis previously subsumed under P. walkeri, which we describe herein. Our data supports the hypothesis that widely distributed Chocoan taxa may generally experience their greatest opportunities for isolation and parapatric speciation across thermal elevational gradients. Finally, our study provides critical information to predict which unstudied lineages may harbor cryptic diversity, and how geology and climate are likely to have shaped their evolutionary history.

A molecular phylogeny of the Neotropical snail-eating snakes (tribe Dipsadini) is presented including 43 (24 for the first time) of the 77 species, sampled for both nuclear and mitochondrial genes. Morphological and phylogenetic support was found for four new species of Dipsas and one of Sibon, which are described here based on their unique combination of molecular, meristic, and color pattern characteristics. Sibynomorphus is designated as a junior subjective synonym of Dipsas. Dipsas latifrontalis and D. palmeri are resurrected from the synonymy of D. peruana. Dipsas latifasciata is transferred from the synonymy of D. peruana to the synonymy of D. palmeri. A new name, D. jamespetersi, is erected for the taxon currently known as Sibynomorphus petersi. Re-descriptions of D. latifrontalis and D. peruana are presented, as well as the first photographic voucher of an adult specimen of D. latifrontalis, along with photographs of all known Ecuadorian Dipsadini species. The first country record of D. variegata in Ecuador is provided and D. oligozonata removed from the list of Peruvian herpetofauna. With these changes, the number of Dipsadini reported in Ecuador increases to 22, 18 species of Dipsas and four of Sibon.

Within Dipsadinae, some recent authors have recognized a tribe Nothopsini containing the genera Diaphorolepis, Emmochliophis, Nothopsis, Synophis, and Xenopholis, on the basis of a number of putative morphological synapomorphies. However, molecular results suggest that Nothopsis, Synophis, and Xenopholis do not form a monophyletic group, while the remaining taxa are unsampled in recent molecular phylogenies. Here, DNA-sequence data for some Diaphorolepis and Synophis species are provided for the first time, as well as additional new sequences for Nothopsis and some Synophis species. Including these and other existing data for nothopsine species, previous studies showing that Nothopsini is not a natural group are corroborated. Nothopsini Cope, 1871 is restricted to Nothopsis. Diaphorolepidini Jenner, 1981 is resurrected and re-delimited to include only Diaphorolepis, Emmochliophis, and Synophis. Finally, Xenopholis remains Dipsadinae incertae sedis. Known material of Diaphorolepidini is reviewed to generate revised and expanded descriptions and diagnoses at the tribe, genus, and species level. Numerous cryptic species are likely present in Synophis bicolor and Synophis lasallei. Finally, a new population from the low-elevation cloud forests of SW Ecuador is reported upon, which is genetically and morphologically distinct from all other species, that is here named Synophis zaheri sp. n.

We present a molecular phylogeny of snake genus Atractus, with an improved taxon sampling that includes 30 of the 140 species currently recognized. The phylogenetic tree supports the existence of at least three new species in the Pacific lowlands and adjacent Andean slopes of the Ecuadorian Andes, which we describe here. A unique combination of molecular, meristic and color pattern characters support the validity of the new species. With the newly acquired data, we propose and define the Atractus iridescens species group, as well as redefine the Atractus roulei species group. The species Atractus iridescens is reported for the first time in Ecuador, whereas Atractus bocourti and Atractus medusa are removed from the herpetofauna of this country. We provide the first photographic vouchers of live specimens for Atractus multicinctus, Atractus paucidens and Atractus touzeti, along with photographs of 19 other Ecuadorian Atractus species. The current status of Atractus occidentalis and Atractus paucidens is maintained based on the discovery of new material referable to these species. With these changes, the species number reported in Ecuador increases to 27, a number that is likely to increase as material not examined in this work becomes available and included in systematic studies.

Geographic barriers and elevational gradients have long been recognized as important in species diversification. Here, we illustrate an example where both mechanisms have shaped the genetic structure of the Neotropical rainfrog, Pristimantis ornatissimus, which has also resulted in speciation. This species was thought to be a single evolutionary lineage distributed throughout the Ecuadorian Chocó and the adjacent foothills of the Andes. Based on recent sampling of P. ornatissimus sensu lato, we provide molecular and morphological evidence that support the validity of a new species, which we name Pristimantis ecuadorensis sp. nov. The sister species are elevational replacements of each other; the distribution of Pristimantis ornatissimus sensu stricto is limited to the Ecuadorian Chocó ecoregion (< 1100 m), whereas the new species has only been found at Andean localities between 1450–1480 m. Given the results of the Multiple Matrix Regression with Randomization analysis, the genetic difference between P. ecuadorensis and P. ornatissimus is not explained by geographic distance nor environment, although environmental variables at a finer scale need to be tested. Therefore this speciation event might be the byproduct of stochastic historic extinction of connected populations or biogeographic events caused by barriers to dispersal such as rivers. Within P. ornatissimus sensu stricto, morphological patterns and genetic structure seem to be related to geographic isolation (e.g., rivers). Finally, we provide an updated phylogeny for the genus, including the new species, as well as other Ecuadorian Pristimantis.

We present a new molecular phylogeny of the stream treefrog genus Hyloscirtus, with an improved taxon sampling in the Hyloscirtus bogotensis group. The tree supports the existence of three clades within the genus (Hyloscirtus armatus group, H. bogotensis group and Hyloscirtus larinopygion group) in congruence with previous studies, and suggests the presence of at least three new species in the H. bogotensis group. Herein, we describe one of these species, Hyloscirtus mashpi n. sp. from the Pacific slope of the Ecuadorian Andes. The validity of the latter is supported by molecular, morphological and acoustic data. We also tested individuals of the new species for the chytrid fungus Batrachochytrium dendrobatidis, finding a prevalence of 17.6% (6 positives and 28 negatives). However, at sampled streams, frog densities were high, suggesting that H. mashpi n. sp. may be tolerant to the infection.

We report the recent finding of four adults of Atelopus longirostris, a Critically Endangered species that was last seen in 1989, when catastrophic Atelopus declines occurred. The rediscovery of A. longirostris took place in a new locality, Junín, 1250-1480 m asl, Provincia Imbabura, Ecuador, on 28-31 March 2016. The four frogs were found in two isolated small patches of native forest in a fragmented area heavily modified for agriculture and livestock; one patch protected by the Junín Community Reserve, and another non-protected private patch near the reserve. We found high prevalence of Batrachochytrium dendrobatidis (Bd) in the amphibian community of Junín, but A. longirostris tested negative. The finding of A. longirostris after 27 years is surprising and fits an apparent pattern of mild conditions that might be promoting either the recovery or persistence in low numbers of some relict amphibian populations. The frogs are the first founders of an ex situ assurance colony in Jambatu Research and Conservation Center. Expansion of the Junín Community Reserve is urgently needed to add the currently non-protected patch of forest, where A. longirostris also occurs. The restoration of the forest in degraded areas between both forest patches and in the related river margins is also necessary. This restoration will grant the connectivity between both isolated metapopulations and the normal movement of individuals to the breeding sites in the Chalguayacu and Junín River basins. The latter should be protected to prevent any kind of water pollution by the opencast copper exploitation of the mining concession Llurimagua, which is underway. Atelopus longirostris belongs to a group of at least 29 species of Ecuadorian Atelopus that are critically endangered, 15 of which remain unsighted for at least one decade, and most of them might be extinct. Further synchronous, multidisciplinary and integrative research is needed, aiming to understand the most aspects of the biology of species of Atelopus to support in situ and ex situ conservation actions.