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The gray wolf (Canis lupus) is among the few large carnivores that survived the Late Pleistocene megafaunal extinctions. Thanks to their complex history of admixture and extensive geographic range, the number of gray wolf subspecies and their phylogenetic relationships remain poorly understood. Here, we perform whole-genome sequencing of a gray wolf collected from peninsular India that was phenotypically distinct from gray wolves outside India. Genomic analyses reveal that the Indian gray wolf is an evolutionarily distinct lineage that diverged from other extant gray wolf lineages ∼110 thousand years ago. Demographic analyses suggest that the Indian wolf population declined continuously decline since separating from other gray wolves and, today, has exceptionally low genetic diversity. We also find evidence for pervasive and mosaic gene flow between the Indian wolf and African canids including African wolf, Ethiopian wolf, and African wild dog despite their current geographical separation. Our results support the hypothesis that the Indian subcontinent was a Pleistocene refugium and center of diversification and further highlight the complex history of gene flow that characterized the evolution of gray wolves.

A new bush frog species, Raorchestes cangyuanensis sp. nov., from Cangyuan, Yunnan Province, China, is described based on morphological and molecular analyses. It differs from all known congeners by a combination of the following characters: body size small, adult snout-vent length (SVL) 16.1–20.0 mm in males (n=3); tympanum indistinct; tips of all fingers and toes expanded into discs with circummarginal grooves; rudimentary webbing between toes; fingers and toes with lateral dermal fringes; inner and outer metacarpal tubercles present; heels meeting when limbs held at right angles to body; crotch with a distinct black patch; discs of fingers and toes orange; male with external single subgular vocal sac and reddish nuptial pad at the base of first finger.

By introducing different ancillary ligands and solvent molecules, four lanthanide complexes [Yb2(L)2(acac)2(CH3OH)2]·CH3OH·H2O (1), [Lu2(L)2(acac)2(CH3OH)2]·2CH3OH (2), [Yb2(L)2(dbm)2(CH3OH)2]·CH3OH (3), and [Lu2(L)2(dbm)2(C2H5OH)2] (4) (where H2L = 2-(hydroxyimino)-2-[(5-methyl-2-hydroxyphenyl)methylene]hydrazide, Hacac = acetylacetonate and Hdbm = 1,3-diphenyl-1,3-propanedione) have been synthesized and characterized. The X-ray structural analyses show that complexes 1 and 2 are isostructural in the Triclinic system, space group Pī, complex 3 crystallizes in Monoclinic system, space group P21/c, and complex 4 crystallizes in Orthorhombic system, space group Pbca. The metal ions in complexes 1–4 are all eight-coordinated and show a similar distorted triangular dodecahedron coordination geometry. The fluorescence properties of complexes 1–4 were also investigated. Complexes 1 and 3 showed characteristic emission peaks of Yb(III) ion, while complexes 2 and 4 exhibited emission peaks consistent with that of the ligand when they were excited at the same wavelength.

Viviparous (live-bearing) vertebrates have evolved repeatedly within otherwise oviparous (egg-laying) clades. Over two-thirds of these changes in vertebrate reproductive parity mode happened in squamate reptiles, where the transition has happened between 98 and 129 times. The transition from oviparity to viviparity requires numerous physiological, morphological, and immunological changes to the female reproductive tract, including eggshell reduction, delayed oviposition, placental development for supply of water and nutrition to the embryo by the mother, enhanced gas exchange, and suppression of maternal immune rejection of the embryo. We performed genomic and transcriptomic analyses of a closely related oviparous–viviparous pair of lizards ( Phrynocephalus przewalskii and Phrynocephalus vlangalii ) to examine these transitions. Expression patterns of maternal oviduct through reproductive development of the egg and embryo differ markedly between the two species. We found changes in expression patterns of appropriate genes that account for each of the major aspects of the oviparity to viviparity transition. In addition, we compared the gene sequences in transcriptomes of four oviparous–viviparous pairs of lizards in different genera ( Phrynocephalus , Eremias , Scincella , and Sphenomorphus ) to look for possible gene convergence at the sequence level. We discovered low levels of convergence in both amino acid replacement and evolutionary rate shift. This suggests that most of the changes that produce the oviparity–viviparity transition are changes in gene expression, so occasional reversals to oviparity from viviparity may not be as difficult to achieve as has been previously suggested.

A new bush frog species, Raorchestes cangyuanensis sp. nov. , from Cangyuan, Yunnan Province, China, is described based on morphological and molecular analyses. It differs from all known congeners by a combination of the following characters: body size small, adult snout-vent length (SVL) 16.1–20.0 mm in males (n=3); tympanum indistinct; tips of all fingers and toes expanded into discs with circummarginal grooves; rudimentary webbing between toes; fingers and toes with lateral dermal fringes; inner and outer metacarpal tubercles present; heels meeting when limbs held at right angles to body; crotch with a distinct black patch; discs of fingers and toes orange; male with external single subgular vocal sac and reddish nuptial pad at the base of first finger.

The Himalaya are among the youngest and highest mountains in the world, but the exact timing of their uplift and origins of their biodiversity are still in debate. The Himalayan region is a relatively small area but with exceptional diversity and endemism. One common hypothesis to explain the rich montane diversity is uplift-driven diversification—that orogeny creates conditions favoring rapid in situ speciation of resident lineages. We test this hypothesis in the Himalayan region using amphibians and reptiles, two environmentally sensitive vertebrate groups. In addition, analysis of diversification of the herpetofauna provides an independent source of information to test competing geological hypotheses of Himalayan orogenesis. We conclude that the origins of the Himalayan herpetofauna date to the early Paleocene, but that diversification of most groups was concentrated in the Miocene. There was an increase in both rates and modes of diversification during the early to middle Miocene, together with regional interchange (dispersal) between the Himalaya and adjacent regions. Our analyses support a recently proposed stepwise geological model of Himalayan uplift beginning in the Paleocene, with a subsequent rapid increase of uplifting during the Miocene, finally giving rise to the intensification of the modern South Asian Monsoon.

Despite the substantial role that chickens have played in human societies across the world, both the geographic and temporal origins of their domestication remain controversial. To address this issue, we analyzed 863 genomes from a worldwide sampling of chickens and representatives of all four species of wild jungle fowl and each of the five subspecies of red jungle fowl (RJF). Our study suggests that domestic chickens were initially derived from the RJF subspecies Gallus gallus spadiceus whose present-day distribution is predominantly in southwestern China, northern Thailand and Myanmar. Following their domestication, chickens were translocated across Southeast and South Asia where they interbred locally with both RJF subspecies and other jungle fowl species. In addition, our results show that the White Leghorn chicken breed possesses a mosaic of divergent ancestries inherited from other subspecies of RJF. Despite the strong episodic gene flow from geographically divergent lineages of jungle fowls, our analyses show that domestic chickens undergo genetic adaptations that underlie their unique behavioral, morphological and reproductive traits. Our study provides novel insights into the evolutionary history of domestic chickens and a valuable resource to facilitate ongoing genetic and functional investigations of the world’s most numerous domestic animal.

Genetic introgression not only provides material for adaptive evolution but also confounds our understanding of evolutionary history. This is particularly true for canids, a species complex in which genome sequencing and analysis has revealed a complex history of admixture and introgression. Here, we sequence 19 new whole genomes from high-altitude Tibetan and Himalayan wolves and dogs and combine these into a larger data set of 166 whole canid genomes. Using these data, we explore the evolutionary history and adaptation of these and other canid lineages. We find that Tibetan and Himalayan wolves are closely related to each other, and that ∼39% of their nuclear genome is derived from an as-yet-unrecognized wolf-like lineage that is deeply diverged from living Holarctic wolves and dogs. The EPAS1 haplotype, which is present at high frequencies in Tibetan dog breeds and wolves and confers an adaptive advantage to animals living at high altitudes, was probably derived from this ancient lineage. Our study underscores the complexity of canid evolution and demonstrates how admixture and introgression can shape the evolutionary trajectories of species.

Myanmar, a biodiversity hotspot, harbors a striking diversity and endemism of species. Despite this, its herpetofauna remains one of the least explored in continental Asia due to restrictions of crossing political boundaries and infrastructure in remote regions. Many species in adjacent China and India are hypothesized to occur in Myanmar but records are wanting. Recent fieldwork found the frogs Polypedates braueri, Nasutixalus jerdonii and Oreolalax jingdongensis there, and the latter two species represent new generic records for Myanmar. All major morphological characters of these populations match the original descriptions. In addition, our matrilineal genealogy based on DNA barcoding confirms their identities. Overall, these findings confirm that the amphibian diversity is underestimated and this has important implications for conservation. Analyses indicate that northern Myanmar is a biogeographic corridor for the Himalayas, southern China, and northeastern India.

Seven new dinuclear centrosymmetric complexes, [Ln2(dbm)2L2(DMF)2]·2DMF (Ln = Nd (1), Eu (2), Tb (3), Dy (4), Er (5), Yb (6)) and [Dy2(dbm)2L2(CH3OH)2]·2CH3OH (7) (Hdbm = 1,3-diphenyl-1,3-propanedione, H2L = 2-[(2-(amino)benzhydrazide)-methyl]-8-hydroxylquinoline, DMF = dimethyl-formamide) have been synthesized and structurally characterized. Dynamic magnetic studies reveal the slow magnetic relaxation of complexes 4 and 7 are influenced by a fast quantum tunneling relaxation of the magnetization (QTM). However, the different magnetic relaxation behaviors were observed apparently in the two Dy2 complexes under a 3000 Oe dc field and modulated by the subtle change of solvents from DMF to CH3OH. For complex 7, the temperature dependence curves of in-phase (χ′) and out-of-phase (χ″) show better frequency-dependent signals, suggesting a slow relaxation of magnetization, typical of SMM behavior. Fitting the dynamic magnetic data of complex 7 to the Arrhenius law gives the energy barrier Ueff/kB = 44.06 K with the pre-exponential factor τ0 = 2.17 × 10−7 s under a 3000 Oe dc field.

The synthesis and characterization of a new family of neutral tetranuclear Ln(III) complexes, [Eu4L4(CO3)(acac)2(H2O)2(CH3OH)2]·5H2O·2CH3OH (1), [Gd4L4(CO3)(acac)2 (CH3OH)4]·2CH3OH·CH2Cl2 (2), [Tb4L4(CO3)(acac)2(H2O)2(CH3OH)2]·3H2O·CH3OH (3), and [Er4L4(CO3)(acac)2(H2O)2(CH3OH)(DMF)]·H2O·CH3OH (4) (where acac− is acetylacetonate, H2L is 2-(hydroxyimino)-2-[(5-methyl-2-hydroxyphenyl)methylene]hydrazide and DMF = N,N-dimethyl formamide) are reported. For complexes 1–4, by absorbing and fixing CO2 spontaneously under atmospheric conditions, with CO32− as a bridging tetradentate ligand, the four LnIII ions are bridged by two μ2-O oxygen atoms from CO32− to form a Ln4 cluster. X-ray diffraction data reveal that the non-centrosymmetric (1, 3 and 4) or centrosymmetric (2) unit has two distinct LnIII ions, nine-coordinated LnIII ions in distorted spherical capped square antiprism geometry and eight-coordinated LnIII ions in distorted triangular dodecahedron geometry. Magnetic study indicates that complex 2 can act as a cryogenic magnetic refrigerant and the magnetocaloric effect (MCE) was detected with the magnetic entropy change of −ΔSm = 21.43 J kg−1 K−1 at ΔH = 7 T and 5 K. Furthermore, the fluorescence properties of complexes 1 and 3 were also investigated. The results prove that complex 3 can show TbIII characteristic emission peaks, while for complex 1, EuIII typical emission peaks were not observed.

Montane frogs of the genus Quasipaa Dubois, 1992 occur from southern China to Southeast Asia (Frost 2021). Analyses of mtDNA (Cytb) and nuDNA data (Rag1, Rag2, Rhod, Tyr) for samples from 93 localities throughout its distribution yield a phylogeny. Clades A and B occur in Southeast Asia, clade C in northern Yangtze River, China, clade D in southwestern China, and clades E and F in southeastern China. Results place Q. yei within monophyletic Quasipaa and identify two new species. Based on nuDNA data, the basal split of clade A and B indicates an Indochinese origin of Quasipaa. The west–east diversification of five species across South China (Q. spinosa, Q. exilispinosa, Q. jiulongensis, Q. shini, Q. boulengeri) corresponds to topographic terrains II and III of China. Divergence of species from southeastern China (Q. shini, Q. jiulongensis, Q. spinosa, Q. exilispinosa) and southwestern China (Q. boulengeri) dates to 15.30–16.56 Ma (million years ago). A principal component analysis (PCA) and t-test involving 19 bioclimatic variables identifies significantly different environmental conditions between the two regions. Species’ distribution models (SDM) for Q. spinosa and Q. boulengeri identify the best areas to be eastern and western South China, respectively. Thus, environmental variation appears to have influenced the genetic divergence and distributions of Quasipaa in South China. Mito-nuclear discordance indicates that some individuals of Q. exilispinosa and Q. spinosa hybridized historically.

Genetic introgression provides material for adaptive evolution, but also confounds our understanding of evolutionary history. This is particularly true for canids, a species complex including dogs, wolves, coyotes, jackals, and other wolf-like carnivores, in which genomic analyses have revealed a complex history of admixture and introgression. Among the most enigmatic canid lineages whose origin and evolutionary relationships remain unresolved are the high-altitude wolves of the Tibetan Plateau, the Tibetan grey wolf and the Himalayan wolf. Here, we use newly assembled genomes of high-altitude Tibetan and Himalayan wolves to explore the evolutionary history and adaptation of this charismatic group. We find that Tibetan and Himalayan wolves are closely related to each other, and that as much as 38-43% of their nuclear genome is derived from an as-yet unrecognized wolf-like lineage that is deeply diverged from living Holarctic wolves and dogs. The EPAS1 haplotype, which is present at high frequencies in Tibetan dog breeds and wolves and confers an adaptive advantage to animals living at high altitudes, was probably derived from this ancient lineage. Our study underscores the complexity of canid evolution and demonstrates how admixture and introgression shape the evolutionary trajectories and adaptation of species.

Pleistocene glacial-interglacial climatic oscillations greatly shaped the current genetic structure of many species. However, geographic features may influence the impact of climatic cycling. Distinct geographic and environmental characters between northern and southern parts of the eastern Qinghai-Tibetan Plateau (EQTP) facilitate explorations into the impacts of geographic features on species. The northern parts of EQTP contain large areas of marsh, and the environment is rather homogeneous. In contrast, the southern EQTP harbors complex alpine valleys and a much more heterogeneous setting. We evaluate DNA sequence variation from both the mitochondrial and nuclear genomes in Nanorana pleskei, a species endemic to the EQTP. Hypothesis testing on the evolutionary history of N. pleskei indicates that northern populations can disperse freely, but alpine valleys isolate southern populations. Demographic histories between northern and southern populations also differ. Northern populations appear to have experienced population expansions, while southern frogs exhibit a far more stable demographic history. By combining climatic analyses and species' distribution models, our study suggests that geographic and environmental features drive the differences between the northern and southern EQTP.

South China and Indochina host striking species diversity and endemism. Complex tectonic and climatic evolutions appear to be the main drivers of the biogeographic patterns. In this study, based on the geologic history of this region, we test 2 hypotheses using the evolutionary history of Microhyla fissipes species complex. Using DNA sequence data from both mitochondrial and nuclear genes, we first test the hypothesis that the Red River is a barrier to gene flow and dispersal. Second, we test the hypothesis that Pleistocene climatic cycling affected the genetic structure and population history of these frogs. We detect 2 major genetic splits that associate with the Red River. Time estimation suggests that late Miocene tectonic movement associated with the Red River drove their diversification. Species distribution modeling (SDM) resolves significant ecological differences between sides of the Red River. Thus, ecological divergence also probably promoted and maintained the diversification. Genogeography, historical demography, and SDM associate patterns in southern China with climate changes of the last glacial maximum (LGM), but not Indochina. Differences in geography and climate between the 2 areas best explain the discovery. Responses to the Pleistocene glacial-interglacial cycling vary among species and regions.

Nine dinuclear Ln(III) complexes, [Ln(dbm)2(L)]2 (Ln = Eu (1), Tb (2), Dy (3), Ho (4), Er (5)) and [Ln(dbm)2(L′)]2 (Ln = Tb (6), Dy (7), Ho (8), Er (9)) (dbm = 1,3-diphenyl-1,3-propanedione, HL = 2-[[(4-methoxy-phenyl)imino]methyl]-8-hydroxy-quinoline and HL′ = 2-[[(4-ethoxyphenyl)imino]methyl]-8-hydroxyquinoline) have been synthesized, and structurally and magnetically characterized. The nine complexes are all phenoxo-O bridged binuclear complexes, in which Ln1 and Ln1a are in an eight-coordinated environment bridged by two phenoxido oxygen atoms of two 8-hydroxyquinoline Schiff base ligands. Although complexes 3 and 7 have very similar structures, magnetic studies reveal that they exhibit different magnetic relaxation behaviors with the effective barriers (ΔE/kB) of 34.5 K for 3 and 67.6 K for 7. The dissimilar dynamic magnetic behaviors of 3 and 7 mostly result from the different electron-donating effect induced by the two alkoxy (–OCH3 and –OC2H5) of the 8-hydroxyquinoline Schiff base ligands. Meanwhile, for complexes 2, 5, 6 and 9, there are no observed magnetic relaxation behaviors under a zero dc field. In addition, the luminescence properties of 1, 2 and 6 were studied.

The genus Amolops ("torrent frogs") is one of the most species-rich genera in Ranidae, with 59 recognized species. This genus currently includes six species groups diagnosed mainly by morphology. Several recent molecular studies indicated that the classification of species groups within Amolops remains controversial, and key nodes in the phylogeny have been inadequately resolved. In addition, the diversity of Amolops remains poorly understood, especially for those from incompletely sampled regions. Herein, we investigate species-level diversity within the genus Amolops throughout southern China and Southeast Asia, and infer evolutionary relationships among the species using mtDNA data (16S, COI, and ND2). Molecular analyses indicate nine unnamed species, mostly distributed in the Himalayas. We then utilized anchored hybrid enrichment to generate a dataset representing the major mitochondrial lineages to resolve phylogenetic relationships, biogeography, and pattern of species diversification. Our resulting phylogeny strongly supports the monophyly of four previously identified species groups (the A. ricketti, A. daiyunensis, A. hainanensis, and A. monticola groups), but paraphyly for the A. mantzorum and A. marmoratus groups, as previously defined. We erect one new species group, the A. viridimaculatus group, and recognize Dubois' (1992) subgenus Amo as the A. larutensis species group. Biogeographic analysis suggests that Amolops originated on the Indo-Burma/Thai-Malay Peninsula at the Eocene/Oligocene boundary, and dispersed outward, exemplifying a common pattern observed for the origin of Asian biodiversity. The early divergence within Amolops coincides with the Himalayan uplift and the lateral extrusion of Indochina at the Oligocene/Miocene boundary. Our results show that paleoclimatic and geomorphological events have profoundly influenced the patterns of lineage diversification within Amolops.

Rhacophoridae are one of the most speciose and ecologically diverse families of amphibians. Resolution of their evolutionary relationships is key to understanding the accumulation of biodiversity, yet previous hypotheses based on Sanger sequencing exhibit much discordance amongst generic relationships. This conflict precludes the making of sound macroevolutionary conclusions. Herein, we conduct the first phylogenomic study using broad-scale sampling and sequences of 352 nuclear DNA loci obtained using anchored hybrid enrichment targeted sequencing. The robust time-calibrated phylogenetic hypothesis clarifies several long-disputed relationships and facilitates the testing of evolutionary hypotheses on spatiotemporal diversification and reproductive modes. The major extant lineages of Rhacophoridae appear to have radiated in mainland Asia, and the spatiotemporal process corresponds with several common accumulations of biodiversity in Asia. Analyses do not detect any case of “Out of Himalaya” in Rhacophoridae. All transitions of reproductive modes appear to have evolved in an ordered, gradual sequence associated with gaining independence of standing water for larval development. The different reproductive modes are phylogenetically conserved and the completion of their transitions appear to have occurred over a period of ~30 Ma, which does not fit a pattern of a rapid burst of diversification. Innovations in reproductive modes associate statistically with the uneven distribution of species-richness between clades, where higher diversification is linked to increased terrestrial modes of reproduction. These results strengthen the hypothesis that breeding innovations drive diversification by providing new opportunities for ecological release and dispersion.

Significance Central topics in evolutionary biology include uncovering the processes and genetic bases of speciation and documenting environmental adaptations and processes responsible for them. The challenging environment of the Qinghai-Tibetan Plateau (QTP) facilitates such investigations, and the Tibetan frog, Nanorana parkeri, offers a unique opportunity to investigate these processes. A cohort of whole-genome sequences of 63 individuals from across its entire range opens avenues for incorporating population genomics into studies of speciation. Natural selection plays an important role in maintaining and driving the continuing divergence and reproductive isolation of populations of the species. The QTP is a natural laboratory for studying how selection drives adaptation, how environments influence evolutionary history, and how these factors can interact to provide insight into speciation., Tibetan frogs, Nanorana parkeri, are differentiated genetically but not morphologically along geographical and elevational gradients in a challenging environment, presenting a unique opportunity to investigate processes leading to speciation. Analyses of whole genomes of 63 frogs reveal population structuring and historical demography, characterized by highly restricted gene flow in a narrow geographic zone lying between matrilines West (W) and East (E). A population found only along a single tributary of the Yalu Zangbu River has the mitogenome only of E, whereas nuclear genes of W comprise 89–95% of the nuclear genome. Selection accounts for 579 broadly scattered, highly divergent regions (HDRs) of the genome, which involve 365 genes. These genes fall into 51 gene ontology (GO) functional classes, 14 of which are likely to be important in driving reproductive isolation. GO enrichment analyses of E reveal many overrepresented functional categories associated with adaptation to high elevations, including blood circulation, response to hypoxia, and UV radiation. Four genes, including DNAJC8 in the brain, TNNC1 and ADORA1 in the heart, and LAMB3 in the lung, differ in levels of expression between low- and high-elevation populations. High-altitude adaptation plays an important role in maintaining and driving continuing divergence and reproductive isolation. Use of total genomes enabled recognition of selection and adaptation in and between populations, as well as documentation of evolution along a stepped cline toward speciation.

Southeast Asia and southern China (SEA-SC) harbor a highly diverse and endemic flora and fauna that is under increasing threat. An understanding of the biogeographical history and drivers of this diversity is lacking, especially in some of the most diverse and threatened groups. The Asian leaf-litter frog genus Leptolalax Dubois 1980 is a forest-dependent genus distributed throughout SEA-SC, making it an ideal study group to examine specific biogeographic hypotheses. In addition, the diversity of this genus remains poorly understood, and the phylogenetic relationships among species of Leptolalax and closely related Leptobrachella Smith 1928 remain unclear. Herein, we evaluate species-level diversity based on 48 of the 53 described species from throughout the distribution of Leptolalax. Molecular analyses reveal many undescribed species, mostly in southern China and Indochina. Our well-resolved phylogeny based on multiple nuclear DNA markers shows that Leptolalax is not monophyletic with respect to Leptobrachella and, thus, we assign the former to being a junior synonym of the latter. Similarly, analyses reject monophyly of the two subgenera of Leptolalax. The diversification pattern of the group is complex, involving a high degree of sympatry and prevalence of microendemic species. Northern Sundaland (Borneo) and eastern Indochina (Vietnam) appear to have played pivotal roles as geographical centers of diversification, and paleoclimatic changes and tectonic movements seem to have driven the major divergence of clades. Analyses fail to reject an "upstream" colonization hypothesis, and, thus, the genus appears to have originated in Sundaland and then colonized mainland Asia. Our results reveal that both vicariance and dispersal are responsible for current distribution patterns in the genus.

The horned toad assemblage, genus Megophrys sensu lato, currently includes three groups previously recognized as the genera Atympanophrys, Xenophrys and Megophrys sensu stricto. The taxonomic status and species composition of the three groups remain controversial due to conflicting phenotypic analyses and insufficient phylogenetic reconstruction; likewise, the position of the monotypic Borneophrys remains uncertain with respect to the horned toads. Further, the diversity of the horned toads remains poorly understood, especially for widespread species. Herein, we evaluate species-level diversity based on 45 of the 57 described species from throughout southern China, Southeast Asia and the Himalayas using Bayesian inference trees and the Generalized Mixed Yule Coalescent (GMYC) approach. We estimate the phylogeny using both mitochondrial and nuclear DNA data. Analyses reveal statistically significant mito-nuclear discordance. All analyses resolve paraphyly for horned toads involving multiple strongly supported clades. These clades correspond with geography. We resurrect the genera Atympanophrys and Xenophrys from the synonymy of Megophrys to eliminate paraphyly of Megophrys s.l. and to account for the morphological, molecular and biogeographic differences among these groups, but we also provide an alternative option. Our study suggests that Borneophrys is junior synonym of Megophrys sensu stricto. We provide an estimation of timeframe for the horned toads. The mitochondrial and nuclear trees indicate the presence of many putative undescribed species. Widespread species, such as Xenophrys major and X. minor, likely have dramatically underestimated diversity. The integration of morphological and molecular evidence can validate this discovery. Montane forest dynamics appear to play a significant role in driving diversification of horned toads.

South China and Indochina host striking species diversity and endemism. Complex tectonic and climatic evolutions appear to be the main drivers of the biogeographic patterns. In this study, based on the geologic history of this region, we test 2 hypotheses using the evolutionary history of Microhyla fissipes species complex. Using DNA sequence data from both mitochondrial and nuclear genes, we first test the hypothesis that the Red River is a barrier to gene flow and dispersal. Second, we test the hypothesis that Pleistocene climatic cycling affected the genetic structure and population history of these frogs. We detect 2 major genetic splits that associate with the Red River. Time estimation suggests that late Miocene tectonic movement associated with the Red River drove their diversification. Species distribution modeling (SDM) resolves significant ecological differences between sides of the Red River. Thus, ecological divergence also probably promoted and maintained the diversification. Genogeography, historical demography, and SDM associate patterns in southern China with climate changes of the last glacial maximum (LGM), but not Indochina. Differences in geography and climate between the 2 areas best explain the discovery. Responses to the Pleistocene glacial–interglacial cycling vary among species and regions.

True frogs of the genus Rana are widely used as model organisms in studies of development, genetics, physiology, ecology, behavior, and evolution. Comparative studies among the more than 100 species of Rana rely on an understanding of the evolutionary history and patterns of diversification of the group. We estimate a well-resolved, time-calibrated phylogeny from sequences of six nuclear and three mitochondrial loci sampled from most species of Rana, and use that phylogeny to clarify the group's diversification and global biogeography. Our analyses consistently support an "Out of Asia" pattern with two independent dispersals of Rana from East Asia to North America via Beringian land bridges. The more species-rich lineage of New World Rana appears to have experienced a rapid radiation following its colonization of the New World, especially with its expansion into montane and tropical areas of Mexico, Central America, and South America. In contrast, Old World Rana exhibit different trajectories of diversification; diversification in the Old World began very slowly and later underwent a distinct increase in speciation rate around 29-18 Ma. Net diversification is associated with environmental changes and especially intensive tectonic movements along the Asian margin from the Oligocene to early Miocene. Our phylogeny further suggests that previous classifications were misled by morphological homoplasy and plesiomorphic color patterns, as well as a reliance primarily on mitochondrial genes. We provide a phylogenetic taxonomy based on analyses of multiple nuclear and mitochondrial gene loci. [Amphibians; biogeography; diversification rate; Holarctic; transcontinental dispersal.

The crab-eating frog, Fejervarya cancrivora, is the only frog that lives near seas. It tolerates increased environmental concentrations of sodium, chloride and potassium partly by raising ion and urea levels in its blood plasma. The molecular mechanism of the adaptation remains rarely documented. Herein, we analyze transcriptomes of the crab-eating frog and its closely related saline-intolerant species, F. limnocharis, to explore the molecular basis of adaptations to such extreme environmental conditions. Analyses reveal the potential genetic mechanism underlying the adaptation to salinity for the crab-eating frog. Genes in categories associated with ion transport appear to have evolved rapidly in F. cancrivora. Both positively selected and differentially expressed genes exhibit enrichment in the GO category regulation of renal sodium excretion. In this category, the positively selected sites of ANPEP and AVPR2 encode CD13 and V2 receptors, respectively; they fall precisely on conserved domains. More differentially expressed rapidly evolved genes occur in the kidney of F. cancrivora than in F. limnocharis. Four genes involved in the regulation of body fluid levels show signs of positive selection and increased expression. Significant up-regulation occurs in several genes of F. cancrivora associated with renin-angiotensin system and aldosterone-regulated sodium reabsorption pathways, which relate to osmotic regulation.

The development of efficient sequencing techniques has resulted in large numbers of genomes being available for evolutionary studies. However, only one genome is available for all amphibians, that of Xenopus tropicalis, which is distantly related from the majority of frogs. More than 96% of frogs belong to the Neobatrachia, and no genome exists for this group. This dearth of amphibian genomes greatly restricts genomic studies of amphibians and, more generally, our understanding of tetrapod genome evolution. To fill this gap, we provide the de novo genome of a Tibetan Plateau frog, Nanorana parkeri, and compare it to that of X. tropicalis and other vertebrates. This genome encodes more than 20,000 protein-coding genes, a number similar to that of Xenopus. Although the genome size of Nanorana is considerably larger than that of Xenopus (2.3 vs. 1.5 Gb), most of the difference is due to the respective number of transposable elements in the two genomes. The two frogs exhibit considerable conserved whole-genome synteny despite having diverged approximately 266 Ma, indicating a slow rate of DNA structural evolution in anurans. Multigenome synteny blocks further show that amphibians have fewer interchromosomal rearrangements than mammals but have a comparable rate of intrachromosomal rearrangements. Our analysis also identifies 11 Mb of anuran-specific highly conserved elements that will be useful for comparative genomic analyses of frogs. The Nanorana genome offers an improved understanding of evolution of tetrapod genomes and also provides a genomic reference for other evolutionary studies.

Anthropogenic global climate changes are one of the greatest threats to biodiversity. Distribution modeling can predict the effects of climate changes and potentially their effects on genetic diversity. DNA barcoding quickly identifies patterns of genetic diversity. As a case study, we use DNA barcodes and distribution models to predict threats under climate changes in the frog Nanorana parkeri, which is endemic to the Qinghai-Tibetan Plateau. Barcoding identifies major lineages W and E. Lineage W has a single origin in a refugium and Lineage E derives from three refugia. All refugia locate in river valleys and each greatly contributes to the current level of intraspecific genetic diversity. Species distribution models suggest that global climate changes will greatly influence N. parkeri, especially in the level of genetic diversity, because two former refugia will fail to provide suitable habitat. Our pipeline provides a novel application of DNA barcoding and has important implications for the conservation of biodiversity in southern areas of the Qinghai-Tibetan Plateau.

DNA barcoding is a proven tool for the rapid and unambiguous identification of species, which is essential for many activities including the vouchering tissue samples in the genome 10K initiative, genealogical reconstructions, forensics and biodiversity surveys, among many other applications. A large-scale effort is underway to barcode all amphibian species using the universally sequenced DNA region, a partial fragment of mitochondrial cytochrome oxidase subunit I COI. This fragment is desirable because it appears to be superior to 16S for barcoding, at least for some groups of salamanders. The barcoding of amphibians is essential in part because many species are now endangered. Unfortunately, existing primers for COI often fail to achieve this goal. Herein, we report two new pairs of primers (➀, ➁) that in combination serve to universally amplify and sequence all three orders of Chinese amphibians as represented by 36 genera. This taxonomic diversity, which includes caecilians, salamanders and frogs, suggests that the new primer pairs will universally amplify COI for the vast majority species of amphibians.